Determination of Optimal Amikacin Dosing Regimens for Pediatric Patients With Burn Wound Sepsis.

This study aimed to develop optimal amikacin dosing regimens for the empirical treatment of Gram-negative bacterial sepsis in pediatric patients with burn injuries. A pharmacodynamic (PD) target in which the peak concentration (Cmax) is ≥8 times the minimum inhibitory concentration (MIC) (Cmax/MIC ≥ 8) is reflective of optimal bactericidal activity and has been used to predict clinical outcomes. Population pharmacokinetic modeling was performed in NONMEM 7.2 for pediatric patients with and without burn injuries. Amikacin pharmacokinetic parameters were compared between the two groups and multiple dosing regimens were simulated using MATLAB to achieve the PD target in ≥90% of patients with burn injuries. The pharmacokinetic analysis included 282 amikacin concentrations from 70 pediatric patients with burn injuries and 99 concentrations from 32 pediatric patients without burns. A one-compartment model with first-order elimination described amikacin pharmacokinetics well for both groups. Clearance (CL) was significantly higher in patients with burn injuries than in patients without (7.22 vs 5.36 L/h, P < .001). The volume of distribution (V) was also significantly increased in patients with burn injuries (22.7 vs 18.7 L, P < .01). Weight significantly influenced amikacin CL (P < .001) and V (P < .001) for both groups. Model-based simulations showed that a higher amikacin dose (≥25 mg/kg) achieved a Cmax/MIC ≥8 in ≥90% of patients with assumed infections of organisms with an MIC = 8 mg/L. Amikacin pharmacokinetics are altered in patients with burn injuries, including a significant increase in CL and V. In simulations, increased doses (≥25 mg/kg) led to improved PD target attainment rates. Further clinical evaluation of this proposed dosing regimen is warranted to assess clinical and microbiological outcomes in pediatric patients with burn wound sepsis.

Trends in bacterial wound isolates and antimicrobial susceptibility in a pediatric burn hospital.

The purpose of this retrospective study was to collate data dealing with organisms cultured from the burn patients and evaluate trends in antimicrobial susceptibility. All cultures collected from each acute admission patient between 2004 and 2011 in the 30-bed pediatric burn hospital were evaluated for their annual frequency and antimicrobial susceptibility. Duplicate cultures were excluded. Staphylococcus aureus was isolated most frequently (25% of total isolates; range, 69-408 isolates/yr), followed by Pseudomonas aeruginosa (13%; range, 40-202 isolates/yr), coagulase-negative staphylococci (9%; range, 2-188 isolates/yr), Enterobacter cloacae (8%; range, 22-128 isolates/yr), and Escherichia coli (6%; range, 19-91 isolates/yr). This rank order remained relatively consistent during the period of study. The emergence of methicillin-resistant S. aureus increased from 20% in 2004 to about 45% in 2009 to 2011. Susceptibility to vancomycin was still 100%. In comparing periods 2004 to 2007 and 2008 to 2011, P. aeruginosa showed increased susceptibility to cefepime (from 76% to 84%) and the aminoglycosides (from 68% to 81%), whereas susceptibility to piperacillin-tazobactam remained high (from 91% to 93%). E. cloacae demonstrated 90 to 100% susceptibility to aminoglycosides, cefepime, and imipenem. E. coli showed an increased rate of resistance to ceftazidime but was still susceptible to imipenem and amikacin. S. aureus and P. aeruginosa continue to be the most prevalent organisms cultured from our pediatric burn population. Almost half of the staphylococcal isolates were methicillin-resistant S. aureus. Despite widespread use of piperacillin-tazobactam, P. aeruginosa susceptibility remained high. Several classes of antimicrobials continued to demonstrate good to excellent activity against the majority of organisms cultured from the burn patients.

A performance improvement initiative to determine the impact of increasing the time interval between changing centrally placed intravascular catheters.

Existing practice guidelines designed to minimize invasive catheter infections and insertion-related complications in general intensive care unit patients are difficult to apply to the burn population. Burn-specific guidelines for optimal frequency for catheter exchange do not exist, and great variation exists among institutions. Previously, the authors' practice was to follow a new site insertion at 48 hours by an exchange over a guidewire, which was followed 48 hours later by a second guidewire exchange (48h group). As a performance improvement initiative, the authors attempted to determine whether there would be any advantage or disadvantage to extending these intervals to 72 hours (72h). All patients with centrally placed intravascular catheters from October 2007 to August 2008 were included in the 48h group, and all patients with catheters placed from September 2008 to December 2009 comprised the 72h group. Catheter infection rates were determined using the National Healthcare Safety Network definition for central line-associated bloodstream infections (CLABSIs) and calculated as CLABSIs/1000 catheter days. The two groups were not significantly different for age, sex, burn etiology, total burn size, or percent third-degree burn. There were 3.1 CLABSIs/1000 catheter days for the 48h group and 2.8 CLABSIs/1000 catheter days for the 72h group (NS). The authors conclude that increasing the central catheter change interval from 48 to 72 hours did not result in any increase in their CLABSI rate. Implementation of this change in practice is expected to decrease supply costs by $28,000 annually in addition to reducing clinical support services needed to perform these procedures.

Amish burn ointment and burdock leaf dressings: assessments of antimicrobial and cytotoxic activities.

Amish burn wound ointment (ABO) contains honey, lanolin, oils, glycerin, bees wax, and other natural additives. Although there are many anecdotal reports that this ointment covered with a burdock leaf (BL) dressing promotes burn wound healing, little scientific testing of this treatment has occurred. The goal of this study was to evaluate in vitro some of the components of this treatment modality for antimicrobial and cytotoxic activities. The ABO was tested for sterility using standard microbiological techniques. Because of the semisolid, lipid-based nature of the salve, the at-use product could not be tested in bioassays. Samples of BL and the dry ingredients (DI) used in the ointment were provided by the Amish vendor. Aqueous extracts of the DI and of the BL were prepared and freeze dried. The freeze-dried extracts were reconstituted, filtered, and tested separately on keratinocyte and fibroblast cell cultures for cytotoxicity (growth inhibition assay) and against a panel of susceptible and resistant microbes for antimicrobial activity (Nathan's agar-well diffusion assay) in a series of concentrations (% wt/vol). Neither DI nor BL extracts demonstrated antimicrobial activity against any of organisms tested. The DI extract inhibited growth of both keratinocytes and fibroblasts at the 0.1% concentration. The 0.1 and 0.03% concentrations of the BL extract were cytotoxic to both keratinocytes and fibroblasts. Although tests for microbial growth from the at-use preparation of the ABO were negative, extracts of the DI and BL did not demonstrate any antimicrobial activity. Additionally, both extracts inhibited the growth of skin cells in vitro at higher concentrations. These results suggest caution in the use of ABO and BL dressings if there is more than a minimal risk of complications from the burn injury.

Amikacin population pharmacokinetics among paediatric burn patients.

INTRODUCTION: The objectives of this study were to (1) determine the pharmacokinetics of amikacin among children with severe burn and (2) identify influential covariates. METHODS: Population-based pharmacokinetic modelling was performed in NONMEM 7.2 for hospitalized children who received amikacin at 10-20mg/kg divided two, three, or four times per day as part of early empiric treatment of presumed burn-related sepsis. RESULTS: The analysis included data from 70 patients (6 months to 17 years) with 282 amikacin serum concentrations. Amikacin's mean Cmax was 33.2±9.4µg/mL and the mean Cmin was 3.8±4.6µg/mL. The final covariate model estimated clearance as 5.98L/h/70kg (4.97-6.99, 95% CI), the volume of distribution in the central compartment as 16.7L/70kg (14.0-19.4, 95% CI), the volume of distribution in the peripheral compartment as 40.1L/70kg (15.0-80.4, 95% CI), and the inter-compartmental clearance as 3.38L/h/70kg (2.44-4.32, 95% CI). In multivariate analyses, current weight (P<0.001) was a significant covariate, while age, sex, height, serum creatinine, C-reactive protein, platelet count, the extent and type of burn, and concomitant vancomycin administration did not influence amikacin pharmacokinetics. DISCUSSION: Children with burn featured elevated amikacin clearance when compared to healthy adult volunteers. However, peak amikacin concentrations are comparable to those attained in other critically-ill children, suggesting that elevated amikacin clearance may not result in sub-therapeutic antibacterial effects. In this study, we found that amikacin displays two-compartment pharmacokinetics, with weight exerting a strong effect upon amikacin clearance. Further pharmacodynamic studies are needed to establish the optimal dosing regimen for amikacin in paediatric burn patients.

A survey of invasive catheter practices in U.S. burn centers.

Burn-specific guidelines for optimal catheter rotation, catheter type, insertion methods, and catheter site care do not exist, and practices vary widely from one burn unit to another. The purpose of this study was to define current practices and identify areas of practice variation for future clinical investigation. An online survey was sent to the directors of 123 U.S. burn centers. The survey consisted of 23 questions related to specific practices in placement and maintenance of central venous catheters (CVCs), arterial catheters, and peripherally inserted central catheters (PICCs). The overall response rate was 36%; response rate from verified centers was 52%. Geographic representation was wide. CVC and arterial catheter replacement varied from every 3 days (24% of sites) to only for overt infection (24% of sites); 23% of sites did not use the femoral position for CVC placement. Nearly 60% of units used some kind of antiseptic catheter. Physicians inserted the majority of catheters, and 22% of sites used nonphysicians for at least some insertions. Ultrasound was routinely used by less than 50% of units. A wide variety of post-insertion dressing protocols were followed. PICCs were used in some critically injured patients in 37% of units; the majority of these users did not rotate PICCs. Thus, it can be surmised that wide practice variation exists among burn centers with regard to insertion and maintenance of invasive catheters. Areas with particular variability that would be appropriate targets of clinical investigation are line rotation protocols, catheter site care protocols, and use of PICCs in acute burns.

Pharmacokinetics of colistin in an adolescent boy with extensive burn injury.

Multidrug resistance in Gram-negative bacteria has led to a resurgence in colistin use. No pharmacokinetic data exist for burn patients. A 17-year-old boy suffered a 71% TBSA full-thickness burn with deep necrosis and compartment syndrome. He developed multidrug-resistant Acinetobacter baumannii burn wound sepsis/septic shock with acute renal failure requiring dialysis. The organism was resistant to all tested antibiotics except colistin. He received colistin 2.5 mg/kg every 24 hours. Peak and trough serum concentrations, area under the concentration-time curve, and elimination half-lives of colistin were 3.6 ± 1.0 µg/ml, 0.9 ± 0.5 µg/ml, 47.1 ± 14.4 mg · hr/L, and 12.3 ± 9.4 hours (mean ± SD), respectively. Serum levels were at or above the minimum inhibitory concentration for >90% of therapy. Nevertheless, salvage therapy with colistin proved futile as the patient developed acidosis, coagulopathy, and was vasopressor-dependent without any wound healing. He died on hospital day 52. Microbiologically, the serum levels of colistin were seemingly adequate, as repeat cultures were negative. Given the peak and trough levels of colistin relative to the minimum inhibitory concentration for the organism (0.5 µg/ml), it would seem that the dosage of 2.5 mg/kg administered every 24 hours for this patient on dialysis was appropriate. Patients on dialysis infected with an organism possessing a higher inhibitory concentration (≥1 µg/ml) should probably receive the same dosage every 12 hours to avoid subtherapeutic concentrations. Large-scale study of the pharmacokinetics of colistin in patients with burn injury is urgently needed.

Mice treated with a benzodiazepine had an improved survival rate following Pseudomonas aeruginosa infection.

Psychological stress has a high incidence after burn injury, therefore, anxiolytic drugs are often prescribed. Unfortunately, to date, no burn study has investigated the effects of anxiolytic drugs on the ability to fight infection. This study was undertaken to determine if psychological stress, anxiety-modulating drugs, or both, alter survival following an infection. On day 0, 7-week-old male C57Bl/6 mice either received a 15% full-thickness flame burn or were sham treated (anesthesia and shaved), whereas controls received no treatment. Mice received midazolam (1 mg/kg intraperitoneally) or saline daily and were stressed by exposure to rat in a guinea pig cage or placed in an empty cage for 1 hour a day, beginning on postburn day 1. For the survival experiments, mice either received bacteria after 2 or 8 consecutive days of predator exposure and drug treatment, which continued daily for 7 days after inoculation. In a separate set of experiments, after eight daily injections of midazolam, mice were given lipopolysaccharide, bacteria, or saline and were killed 12 hours later. Mice that received midazolam had improved survival rates when compared with their saline-treated counterparts, and the protective effect was more significant the more days they received the drug. For most of the cytokines, the bacteria-induced increase was significantly attenuated by midazolam as was the amount of bacteria in the liver. The protective effect seems to be independent of the drug's anxiolytic activity as there were no significant differences in survival between the predator-stressed and the nonstressed mice. The mechanisms responsible for the protective effect remain to be elucidated.

Methicillin-resistant Staphylococcus aureus furunculitis in the outpatient burn setting.

Methicillin-resistant Staphylococcus aureus (MRSA) is becoming more predominant in the community. We have seen increasing cases of furunculitis in our outpatient burn clinic, which appear to develop weeks after the initial burn injury and in patients with limited inpatient stays. We performed a 3-year retrospective review of all outpatient burn patients who developed furunculitis. Data analyzed included length of hospital stay, type of injury sustained, culture and sensitivity results, and treatment provided. A total of 28 patients were identified with MRSA furunculitis, which presented as painful, hard, indurated boils with minimal purulent drainage. Adults had less extensive burn injuries (mean of 12% TBSA adults vs 20% TBSA children) with shorter hospital stays (mean 8 days adults vs 22 days children). Fifty-seven percent of the patients had multiple furuncules, involving both burned and nonburned areas. Patients with furunculitis had a less resistant MRSA strain than those without furunculitis. Of the 22 patients who received systemic antibiotic coverage, 14 (58%) were successfully treated with 1 antibiotic regimen, whereas 8 (33%) required multiple antibiotics. In this study, furunculitis in the outpatient setting was believed to be consistent with community-acquired MRSA. Incision and drainage was not sufficient in patients with multiple furuncles, and systemic antibiotics were administered. Through increased awareness of the prevalence of community-acquired MRSA in the community, appropriate antibiotic treatment can be initiated, and the discomfort and transmission risk associated with this disease can be minimized.

Antibiotic resistance in clinical isolates of Acinetobacter baumannii, Pseudomonas aeruginosa, and Staphylococcus aureus does not impact sensitivity to human beta defensin 4.

Antibiotic usage is essential for infection control but hastens emergence of antibiotic resistant microbes. In particular, Acinetobacter baumannii is an important pathogen because of its heightened ability to acquire drug resistance. The need for novel antibacterial agents led us to evaluate the sensitivity of drug-resistant bacteria to the antimicrobial activity of human beta defensin 4 (HBD-4). Clinical isolates of A. baumannii (N=14), Pseudomonas aeruginosa (N=15), and Staphylococcus aureus (N=20), including 10 methicillin-resistant (MRSA) isolates, were examined. All bacterial strains were susceptible to HBD-4 antimicrobial activity, with no correlation between antibiotic resistance and HBD-4 sensitivity. The results demonstrate that antibiotic resistant microorganisms, including MRSA, can be inhibited by HBD-4, which may represent an effective therapeutic agent for infections involving drug-resistant microorganisms.

Differential immunological phenotypes are exhibited after scald and flame burns.

A dysfunctional immune system is known to be part of the pathophysiology after burn trauma. However, reports that support this have used a variety of methods, with numerous variables, to induce thermal injury. We hypothesized that, all other parameters being equal, an injury infliction by a scald would yield different immunological responses than one inflicted by a flame. Here, we demonstrated that both burn methods produced a full-thickness burn, yet there was more of an increase in subdermal temperature, hematocrit, mortality, and serum IL-6 concentrations associated with the scald burn. On postinjury day 1, the scald-burned mice showed diminished lymphocyte numbers, interferon gamma production, and lymphocyte T-bet expression as compared with sham- and flame-burned mice. On postburn day 8, spleens from both sets of thermally injured animals showed an increase in proinflammatory myeloid cells as compared with sham-burned mice. Furthermore, the T-cell numbers, T-bet expression, and phenotype were changed such that interferon gamma production was higher in scald-burned mice than in sham- and flame-burned mice. Altogether, the data show that differential immunological phenotypes were observed depending on the thermal injury method used.

Are topical antimicrobials effective against bacteria that are highly resistant to systemic antibiotics?

An increasing number of bacteria are resistant to multiple systemic antibiotics. The purpose of this study was to determine if topical antimicrobials are still effective against multi-drug resistant organisms (MDROs). MDROs, including Acinetobacter, Pseudomonas, Klebsiella, Staphylococcus, and Enterococcus, were collected from four burn hospitals. The sensitivity of 47 MDROs to 11 commonly used topical agents (mafenide acetate, nystatin, mafenide + nystatin, silver nitrate, Dakin's, polymyxin B, neomycin, polymyxin + neomycin, silver sulfadiazine, bacitracin, silver sulfadiazine + bacitracin) was tested using the agar well diffusion assay and compared with the sensitivity of 27 non-MDROs of similar genera. Overall 88% of the tests of the non-MDROs showed susceptibility to the topicals compared with 80% for the MDROs (P < .05). Specific findings included: all of the gram-positive non-MDROs were sensitive to bacitracin compared with only 67% of the MDROs (P < .05); 74% of the non-MDROs were sensitive to neomycin vs 26% of the MDROs (P < .01). Even for the susceptible isolates, the zones of inhibition were smaller for the MDROs than for the non-MDROs (P < .002), indicating decreased susceptibility of the MDROs. Specifically, while the MDRO Acinetobacter were sensitive to most of the topicals, the zones of inhibition for silvadene, silvadene + bacitracin, neomycin, and neomycin + polymyxin were significantly smaller (P < .001) for the Acinetobacter MDROs than the non-MDROs. Although many topicals are still effective against some MDROs, MDROs are more resistant to topicals than are non-MDROs. Some treatment assumptions based historically on the efficacy of topical antimicrobial agents against non-MDROs need to be re-evaluated for MDROs.

Immunization with a Pseudomonas aeruginosa 1244 pilin provides O-antigen-specific protection.

The O antigen is both a major structural outer membrane component and the dominant epitope of most gram-negative bacteria. Pseudomonas aeruginosa 1244 produces a type IV pilus and covalently links an O-antigen repeating unit to each pilin monomer. Here we show that immunization of mice with pure pilin from strain 1244 by use of either the mouse respiratory model or the thermal injury model resulted in protection from challenge with a pilus-null O-antigen-producing 1244 mutant. These results provide evidence that the pilin glycan stimulates a protective response that targets the O antigen, suggesting that this system could be used as the basis for the development of a variety of bioconjugate vaccines protective against gram-negative bacteria.

Can contamination of a patient's allograft be traced back to the allograft donor?

Cultures of allograft sites on burn patients occasionally show microbes that were not previously cultured from that patient. Our purpose was to determine 1) if microbes on allograft sites could have been transferred to the burn patient from the allograft donor and 2) if microbial transfer is different if the allograft was fresh or frozen. All allografts were cultured by the skin bank after recovery (pre-antimicrobial) and after the skin had been in an antimicrobial solution (post-antimicrobial). These culture results were compared with the results of cultures taken at the hospital from the allograft sites of burn patients. All allograft recipients at the burn hospital during 2005 were included in this Institutional Review Board approved study. Sixty-one donors provided 143 allografts for 38 patients. From the 61 donors, 114 precultures were taken; 19.5% were positive for at least one organism. Only 6.8% of 118 post-antimicrobial cultures were positive. Of the 143 allografts on burn patients, 111 were used fresh and 32 had been cryopreserved. During dressing changes, 124 cultures were taken from sites that received fresh allograft and 27 from sites that received frozen; 54.8% of cultures from fresh allograft sites and 41.5% from frozen sites (not significant, chi) were positive for microbes, which were mostly the patient's own flora. None of the microbes isolated from the burn patient allograft sites matched organisms on the pre- or post-antimicrobial cultures from the donor allografts. Regardless of whether the allografts were fresh or frozen, no instances were identified in which donor microorganisms were transferred from a donor to a recipient.

American Burn Association consensus conference to define sepsis and infection in burns.

Because of their extensive wounds, burn patients are chronically exposed to inflammatory mediators. Thus, burn patients, by definition, already have "systemic inflammatory response syndrome." Current definitions for sepsis and infection have many criteria (fever, tachycardia, tachypnea, leukocytosis) that are routinely found in patients with extensive burns, making these current definitions less applicable to the burn population. Experts in burn care and research, all members of the American Burn Association, were asked to review the literature and prepare a potential definition on one topic related to sepsis or infection in burn patients. On January 20, 2007, the participants met in Tucson, Arizona to develop consensus for these definitions. After review of the definitions, a summary of the proceedings was prepared. The goal of the consensus conference was to develop and publish standardized definitions for sepsis and infection-related diagnoses in the burn population. Standardized definitions will improve the capability of performing more meaningful multicenter trials among burn centers.

Intranasal immunization with heterologously expressed polysaccharide protects against multiple Pseudomonas aeruginosa infections.

Surface-expressed bacterial polysaccharides are often immunodominant, protective antigens. However, these antigens are chemically and serologically highly heterogeneous, and conjugation to protein carriers is often necessary to enhance their immunogenicity. Here we show the efficacy of intranasal immunization of mice with attenuated Salmonella enterica serovar Typhimurium expressing the O antigen portion of Pseudomonas aeruginosa lipopolysaccharide. P. aeruginosa is an ideal model system because it can cause a myriad of localized and systemic infections. In particular, this bacterium is a leading cause of hospital-acquired pneumonia and is responsible for infections after burns and after eye injury. In addition, there are mouse models of infection that mimic the clinical manifestations of P. aeruginosa infections. Immunized mice were highly protected against infection, with long-lasting immunity to acute P. aeruginosa pneumonia, whereas mice immunized with Salmonella containing only the cloning vector or PBS were not. Prophylactic and therapeutic administration of sera from vaccinated animals protected naive mice. Intranasal vaccination also provided complete protection from infections after burns and reduced pathology after corneal abrasions. These results indicate that intranasal delivery of heterologously expressed polysaccharide antigens provides protection at distinct sites of infection. This approach for the expression and delivery of polysaccharide antigens as recombinant immunogens could be easily adapted to develop vaccines for many infectious agents, without the need for complicated purification and conjugation procedures.

Expression of human beta defensin 4 in genetically modified keratinocytes enhances antimicrobial activity.

Defensins are cationic peptides of the innate host defense system with antimicrobial activity against many of the microorganisms commonly found in burn units. Beta defensins are variably expressed in the epithelia of skin and other organs. Human beta defensin 4 reportedly has antimicrobial activity against Pseudomonas aeruginosa and is not normally expressed in intact skin. Genetic modification was used to ectopically express human beta defensin 4 in cultured primary epidermal keratinocytes. Keratinocytes expressing human beta defensin 4 showed significantly elevated antimicrobial activity against clinically-isolated P. aeruginosa compared with controls. These results suggest that genetic modification of keratinocytes can increase their resistance to microbial contamination. Bioengineered skin replacements containing human beta defensin 4-modified keratinocytes may be useful for transplantation to contaminated burn wounds.

A microbiologic study of enteral feeding hang time in a burn hospital: can feeding costs be reduced without compromising patient safety?

BACKGROUND: Procedural changes for hospitalized patients must always balance safety with fiscal constraints. Microbiologic contamination of enteral feeding solutions has been previously associated with nosocomial infections. Formula manipulation and hang time contribute to microbial load, and there is considerable variation in hang time recommendations in the medical literature. With cost containment in mind, the purpose of this performance improvement study was to determine if an increase in hang time of a modular tube feeding product would increase microbial load or affect the nosocomial infection rate in pediatric burn patients. METHODS: This biphasic trial initially evaluated the microbial load of the feeding after delivery of two 4-hour aliquots into a container using the same delivery set (total hang time of 8 hours; number of tests = 20). Second, once this feeding procedure was deemed microbiologically safe, tube feedings were administered to patients, and both microbial load and nosocomial infection rate were monitored for 1 year. RESULTS: Contamination levels at the end of the 8-hour period using the same feeding set with 2 consecutive 4-hour feeding aliquots (number of tests = 38) were lower than standard recommendations. The hospital's nosocomial infection rate was not altered by this procedural change, and feeding-set expenses were reduced. CONCLUSIONS: The hang time of our enteral feeding administration set can be increased safely from 4 hours to 8 hours, with the tube feeding preparation added as two 4-hour aliquots without a significant change in microbial load or nosocomial infection rate, thus promoting simultaneous fiscal responsibility and patient safety.

Redesign of portable suction equipment cases: an engineering approach to a disinfection problem.

BACKGROUND: Patients with tracheostomies carry a case for suctioning supplies. A returned case cultured positive for Staphylococcus aureus even after it had been cleaned with a disinfectant. The purposes of this study were to determine (1) the source of the S aureus and (2) how to provide a microbiologically safe case for the next patient. METHODS: Reviews of patient and environmental cultures plus pulsed-field gel electrophoresis (PFGE) were used to determine the source of the S aureus. Repeated wiping with various detergents/disinfectants did not remove the S aureus from the case, which could not be immersed for cleaning because of a chipboard component. We designed a case made of denim with a washable synthetic fiber to provide shape. The new cases were purposefully contaminated and laundered and then cultured. RESULTS: PFGE indicated that the S aureus was from the patient who had most recently used the case. Contamination of the denim case with the original S aureus, followed by laundering, resulted in a case that was free of the S aureus. These results were repeated for other pathogens. CONCLUSION: Commercially available cases for suction equipment can become contaminated with pathogens from the user and may be difficult to disinfect. By making the case out of washable materials, less expensive cases, which could be readily disinfected, resulted.

Analysis of Pseudomonas aeruginosa diguanylate cyclases and phosphodiesterases reveals a role for bis-(3'-5')-cyclic-GMP in virulence.

The opportunistic pathogen Pseudomonas aeruginosa is responsible for systemic infections in immunocompromised individuals and chronic respiratory disease in patients with cystic fibrosis. Cyclic nucleotides are known to play a variety of roles in the regulation of virulence-related factors in pathogenic bacteria. A set of P. aeruginosa genes, encoding proteins that contain putative domains characteristic of diguanylate cyclases (DGCs) and phosphodiesterases (PDEs) that are responsible for the maintenance of cellular levels of the second messenger bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) was identified in the annotated genomes of P. aeruginosa strains PAO1 and PA14. Although the majority of these genes are components of the P. aeruginosa core genome, several are located on presumptive horizontally acquired genomic islands. A comprehensive analysis of P. aeruginosa genes encoding the enzymes of c-di-GMP metabolism (DGC- and PDE-encoding genes) was carried out to analyze the function of c-di-GMP in two disease-related phenomena, cytotoxicity and biofilm formation. Analysis of the phenotypes of DGC and PDE mutants and overexpressing clones revealed that certain virulence-associated traits are controlled by multiple DGCs and PDEs through alterations in c-di-GMP levels. A set of mutants in selected DGC- and PDE-encoding genes exhibited attenuated virulence in a mouse infection model. Given that insertions in different DGC and PDE genes result in distinct phenotypes, it seems likely that the formation or degradation of c-di-GMP by these enzymes is in highly localized and intimately linked to particular targets of c-di-GMP action.