Albumin reduces the antibacterial activity of polyhexanide-biguanide-based antiseptics against Staphylococcus aureus and MRSA.

BACKGROUND: Wound infection is one of the major complications in acute and chronic wound healing. Antiseptic solutions and wound irrigating agents are routinely used for therapy and prevention in healthcare today. Even if wound exudate contains total protein concentrations up to 9.3% and albumin concentrations up to 2.7% its influence to the antibacterial efficacy of these agents is barely investigated. MATERIALS AND METHODS: This study analyzed the antibacterial effect of polyhexanide biguanide (PHMB) agents (PHMB-concentration 0.005-0.1%) against Staphylococcus aureus and methicillin-resistant-S. aureus (MRSA) after 2min incubation in presents of albumin in different concentrations (0-3%) in a standardized quantitative suspension assay. RESULTS: A significant decrease of the antibacterial activity against S. aureus was shown for a PHMB-concentration of 0.005% from 0.3% albumin (p<0.05), respectively highly significant from 0.75% (p<0.01) on. Thereby the loss of antimicrobial effect was presented as a linear correlation to the rising concentration of albumin. Furthermore a reduction of the antibacterial activity against MRSA in comparison to S. aureus was presented, for albumin concentrations from 3% on highly significant (p<0.01). CONCLUSION: The study showed that albumin causes a significant decrease of the antibacterial potency of PHMB-based antiseptics. Furthermore a diminished potency of the investigated substances for MRSA-contaminated wounds must be taken in consideration. If in vitro experiments show a significant decrease of antibacterial efficacy in the presence of albumin a sufficient activity of PHMB-based agents in clinical practice, especially in cases of exuding wounds or dried-up exudates, cannot be expected.

Maternal intestinal flora and wheeze in early childhood.

BACKGROUND: Increasing evidence links altered intestinal flora in infancy to eczema and asthma. No studies have investigated the influence of maternal intestinal flora on wheezing and eczema in early childhood. OBJECTIVE: To investigate the link between maternal intestinal flora during pregnancy and development of wheeze and eczema in infancy. METHODS: A total of 60 pregnant women from the Boston area gave stool samples during the third trimester of their pregnancy and answered questions during pregnancy about their own health, and about their children's health when the child was 2 and 6 months of age. Quantitative culture was performed on stool samples and measured in log(10)colony-forming units (CFU)/gram stool. Primary outcomes included infant wheeze and eczema in the first 6 months of life. Atopic wheeze, defined as wheeze and eczema, was analysed as a secondary outcome. RESULTS: In multivariate models adjusted for breastfeeding, day care attendance and maternal atopy, higher counts of maternal total aerobes (TA) and enterococci (E) were associated with increased risk of infant wheeze (TA: OR 2.32 for 1 log increase in CFU/g stool [95% CI 1.22, 4.42]; E: OR 1.57 [95% CI 1.06, 2.31]). No organisms were associated with either eczema or atopic wheeze. CONCLUSIONS AND CLINICAL RELEVANCE: In our cohort, higher maternal total aerobes and enterococci were related to increased risk of infant wheeze. Maternal intestinal flora may be an important environmental exposure in early immune system development.

Endogenous bacterial flora in pregnant women and the influence of maternal genetic variation.

Preterm labour, premature prelabour rupture of membranes and low birth weight have all been associated with either specific maternal genital tract infections or an altered vaginal microflora during pregnancy. Factors that influence the variation in microbial-host interaction play an important role in individual susceptibility to adverse pregnancy outcomes. The innate immune responses at mucosal surfaces play a crucial role against microbial invasion. Multiple genes are responsible for the regulation of the innate immune system. Genetic polymorphisms that disrupt innate immune recognition or the responses to infectious microorganisms could explain the alterations in microflora and individual susceptibility to pregnancy complications.

Pregnancy disorders that lead to delivery before the 28th week of gestation: an epidemiologic approach to classification.

Epidemiologists have grouped the multiple disorders that lead to preterm delivery before the 28th week of gestation in a variety of ways. The authors sought to identify characteristics that would help guide how to classify disorders that lead to such preterm delivery. They enrolled 1,006 women who delivered a liveborn singleton infant of less than 28 weeks' gestation at 14 centers in the United States between 2002 and 2004. Each delivery was classified by presentation: preterm labor (40%), prelabor premature rupture of membranes (23%), preeclampsia (18%), placental abruption (11%), cervical incompetence (5%), and fetal indication/intrauterine growth restriction (3%). Using factor analysis (eigenvalue = 1.73) to compare characteristics identified by standardized interview, chart review, placental histology, and placental microbiology among the presentation groups, the authors found 2 broad patterns. One pattern, characterized by histologic chorioamnionitis and placental microbe recovery, was associated with preterm labor, prelabor premature rupture of membranes, placental abruption, and cervical insufficiency. The other, characterized by a paucity of organisms and inflammation but the presence of histologic features of dysfunctional placentation, was associated with preeclampsia and fetal indication/intrauterine growth restriction. Disorders leading to preterm delivery may be separated into two groups: those associated with intrauterine inflammation and those associated with aberrations of placentation.

Nugent score related to vaginal culture in pregnant women.

OBJECTIVE: To relate Gram-stained smears, using the Nugent criteria, to quantitative and qualitative vaginal cultures in pregnant women. METHODS: Two independent evaluators using the Nugent criteria, a standardized method of Gram-stain interpretation designed to detect bacterial vaginosis, scored 104 vaginal smears from pregnant women. The quantitative and qualitative vaginal cultures were assessed at the same time and the results expressed as log(10) colony-forming units per gram of vaginal secretion. The Nugent scores were compared with the microbiologic findings. RESULTS: The prevalence of normal, intermediate, or bacterial vaginosis vaginal flora as determined by Gram stain was 68%, 21%, and 11%, respectively. A comparison of the mean bacterial counts with the Nugent score showed a weak negative correlation for Lactobacillus species and a positive correlation for gram-variable and gram-negative rods. Additional analysis revealed a strong positive correlation between the mean bacterial counts analyses of Peptostreptococcus, a genus not included in the Nugent scoring system, and the Nugent score. In addition, the Prevotella counts correlated strongly with both the Nugent score and the Peptostreptococcus counts. The quantitative counts for Lactobacillus did not vary significantly among the three defined groups of vaginal microflora; however, significant increases in the concentrations of Gardnerella vaginalis and Prevotella were found as the Nugent score increased. CONCLUSION: A strong correlation was found among the gram-variable and gram-negative genera comprised by the Nugent score. Peptostreptococcus also correlated strongly with the Nugent score and with the Prevotella counts, suggesting that this genus may play a role in determining vaginal health.

Using electronic data to predict the probability of true bacteremia from positive blood cultures.

As part of a project to help physicians make more appropriate treatment decisions, we implemented a clinical prediction rule that computes the probability of true bacteremia for positive blood cultures and displays this information when culture results are viewed online. Prior to implementing the rule, we performed a revalidation study to verify the accuracy of the previously published logistic regression model. We randomly selected 114 cases of positive blood cultures from a recent one-year period and performed a paper chart review with the help of infectious disease experts to determine whether the cultures were true positives or contaminants. Based on the results of this revalidation study, we updated the probabilities reported by the model and made additional enhancements to improve the accuracy of the rule. Next, we implemented the rule into our hospital's laboratory computer system so that the probability information was displayed with all positive blood culture results. We displayed the prediction rule information on approximately half of the 2184 positive blood cultures at our hospital that were randomly selected during a 6-month period. During the study, we surveyed 54 housestaff to obtain their opinions about the usefulness of this intervention. Fifty percent (27/54) indicated that the information had influenced their belief of the probability of bacteremia in their patients, and in 28% (15/54) of cases it changed their treatment decision. Almost all (98% (53/54)) indicated that they wanted to continue receiving this information. We conclude that the probability information provided by this clinical prediction rule is considered useful to physicians when making treatment decisions.

Production of toxic shock syndrome toxin 1 by Staphylococcus aureus requires both oxygen and carbon dioxide.

The effect of O(2) and CO(2) on expression of toxic shock syndrome toxin 1 (TSST-1) by Staphylococcus aureus was investigated under controlled growth conditions with continuous-culture techniques. To stimulate TSST-1 production, air and anaerobic gas were premixed before delivery to the culture vessel. At a growth rate-or mass doubling time (t(d))-of 3 h, production of specific TSST-1 (expressed as micrograms per milligram of cell dry weight) was 5. 9-fold greater at an O(2) concentration of 4% than under anaerobic conditions. Increasing the O(2) concentration to 11% did not result in a significant increase (P> 0.05) in the rate of toxin production over that during growth in 4% O(2) but did result in a significant increase (4.9-fold; P<0.001) in the rate of toxin production over that during anaerobic growth. At a t(d) of 9 h, addition of 3.5% O(2) resulted in a 7.6-fold increase in specific TSST-1 production. When room air was sparged through a culture growing at a t(d) of 9 h, TSST-1 production increased significantly (by 3.4-fold) over that during anaerobic growth. When a growth environment of 4% O(2)-remainder N(2) was studied, no increase in TSST-1 production was observed; this was also the case with 8% O(2) at gas-flow rates of 0.1, 0.2, and 0.4 liters/min. In all experiments, production of biomass (expressed as milligrams of cell dry weight per milliliter) increased, indicating that O(2) was metabolized by S. aureus. Addition of CO(2) to the gas mix (4% O(2), 10% CO(2), 86% N(2)) resulted in a 5.1- to 6.8-fold increase in TSST-1 production over that during anaerobic growth and a 3.6-fold increase over that during growth in an environment of 4% O(2)-remainder N(2). The agr mutant strain tested produced 6.1-fold more specific TSST-1 in a growth environment of 4% O(2)-10% CO(2)-86% N(2) than during anaerobic growth. These data suggest that in this system, O(2) alone does not trigger production of TSST-1; rather, both CO(2) and O(2) are required.

T cells activated by zwitterionic molecules prevent abscesses induced by pathogenic bacteria.

Immunologic paradigms classify bacterial polysaccharides as T cell-independent antigens. However, these models fail to explain how zwitterionic polysaccharides (Zps) confer protection against intraabdominal abscess formation in a T cell-dependent manner. Here, we demonstrate that Zps elicit a potent CD4+ T cell response in vitro that requires available major histocompatibility complex class II molecules on antigen-presenting cells. Specific chemical modifications to Zps show that: 1) the activity is specific for carbohydrate structure, and 2) the proliferative response depends upon free amino and carboxyl groups on the repeating units of these polysaccharides. Peptides synthesized to mimic the zwitterionic charge motif associated with Zps also exhibited these biologic properties. Lysine-aspartic acid (KD) peptides with more than 15 repeating units stimulated CD4+ T cells in vitro and conferred protection against abscesses induced by bacteria such as Bacteroides fragilis and Staphylococcus aureus. Evidence for the biologic importance of T cell activation by these zwitterionic polymers was provided when human CD4+ T cells stimulated with these molecules in vitro and adoptively transferred to rats in vivo conferred protection against intraabdominal abscesses induced by viable bacterial challenge. These studies demonstrate that bacterial polysaccharides with a distinct charge motif activate T cells and that this activity confers immunity to a distinct pathologic response to bacterial infection.

Effect of molecular size on the ability of zwitterionic polysaccharides to stimulate cellular immunity.

The large-molecular-sized zwitterionic capsular polysaccharide of the anaerobe Bacteroides fragilis NCTC 9343, designated polysaccharide (PS) A, stimulates T cell proliferation in vitro and induces T cell-dependent protection against abscess formation in vivo. In the present study, we utilized a modification of a recently developed ozonolytic method for depolymerizing polysaccharides to examine the influence of the molecular size of PS A on cell-mediated immunity. Ozonolysis successfully depolymerized PS A into structurally intact fragments. PS A with average molecular sizes of 129.0 (native), 77.8, 46.9, and 17.1 kDa stimulated CD4+-cell proliferation in vitro to the same degree, whereas the 5.0-kDa fragment was much less stimulatory than the control 129.0-kDa PS A. Rats treated with 129.0-kDa, 46.9-kDa, and 17.1-kDa PS A molecules, but not those treated with the 5.0-kDa molecule, were protected against intraabdominal abscesses induced by challenge with viable B. fragilis. These results demonstrate that a zwitterionic polysaccharide as small as 22 repeating units (88 monosaccharides) elicits a T cell-dependent immune response. These findings clearly distinguish zwitterionic T cell-dependent polysaccharides from T cell-independent polysaccharides and give evidence of the existence of a novel mechanism for a polysaccharide-induced immune response.

Microbial interactions in the vaginal ecosystem, with emphasis on the pathogenesis of bacterial vaginosis.

During bacterial vaginosis (BV), populations of lactobacilli which are generally dominant in the vagina of overtly healthy women are replaced by other facultative and anaerobic microorganisms. Some Lactobacillus strains produce hydrogen peroxide and all produce lactic acid; however, the antagonistic role of these metabolites in vivo remains controversial. Positive interactions among BV-associated organisms may contribute to the pathogenesis of BV and its sequelae.

Effect of surgical adhesion reduction devices on the propagation of experimental intra-abdominal infection.

HYPOTHESIS: The use of certain surgical adhesion reduction devices where there is a risk of concomitant bacterial contamination potentiates intra-abdominal infection. DESIGN: Evaluation of adhesion reduction devices in an experimental model of intra-abdominal infection. SETTING: Experimental animal model. INTERVENTIONS: Adhesion reduction devices were administered at the time of bacterial challenge. MAIN OUTCOME MEASURES: Animal mortality rate, abscess formation, and bacterial counts in peritoneal fluid and blood cultures. RESULTS: The use of bioresorbable membrane adhesion reduction devices in the presence or absence of antibiotic therapy did not alter the disease process as compared with appropriate control groups. However, adhesion reduction gels prepared from sodium hyaluronate and carboxymethylcellulose chemically modified with carbodiimide or ferric ion complexed sodium hyaluronate increased the incidence of peritonitis in treated animals. Gel formulations containing diimide-modified carboxymethylcellulose did not have this effect. CONCLUSIONS: The use of certain adhesion reduction devices resulted in the propagation of intra-abdominal infection in an experimental rat model. This outcome was dependent on the composition of the device employed. The use of adhesion reduction devices should be tested in appropriate models of infection where there is the risk of concomitant bacterial contamination.

IL-2 mediates protection against abscess formation in an experimental model of sepsis.

Little is known regarding the mechanism by which T cells control intraabdominal abscess formation. Treating animals with polysaccharide A (PS A) from Bacteroides fragilis shortly before or after challenge protects against abscess formation subsequent to challenge with different abscess-inducing bacteria. Although bacterial polysaccharides are considered to be T cell-independent Ags, T cells from PS A-treated animals mediate this protective activity. In the present study, we demonstrate that CD4+ T cells transfer PS A-mediated protection against abscess formation, and that a soluble mediator produced by these cells confers this activity. Cytokine mRNA analysis showed that T cells from PS A-treated animals produced transcript for IL-2, IFN-gamma, and IL-10, but not for IL-4. The addition of IL-2-specific Ab to T cell lysates taken from PS A-treated animals abrogated the ability to transfer protection, whereas the addition of Abs specific for IFN-gamma and IL-10 did not affect protection. Finally, administration of rIL-2 to animals at the time of bacterial challenge prevented abscess formation in a dose-dependent manner. These data demonstrate that PS A-mediated protection against abscess formation is dependent upon a CD4+ T cell-dependent response, and that IL-2 is essential to this immune mechanism.

Analysis of a capsular polysaccharide biosynthesis locus of Bacteroides fragilis.

A major clinical manifestation of infection with Bacteroides fragilis is the formation of intra-abdominal abscesses, which are induced by the capsular polysaccharides of this organism. Transposon mutagenesis was used to locate genes involved in the synthesis of capsular polysaccharides. A 24,454-bp region was sequenced and found to contain a 15,379-bp locus (designated wcf) with 16 open reading frames (ORFs) encoding products similar to those encoded by genes of other bacterial polysaccharide biosynthesis loci. Four genes encode products that are similar to enzymes involved in nucleotide sugar biosynthesis. Seven genes encode products that are similar to sugar transferases. Two gene products are similar to O-acetyltransferases, and two products are probably involved in polysaccharide transport and polymerization. The product of one ORF, WcfH, is similar to a set of deacetylases of the NodB family. Deletion mutants demonstrated that the wcf locus is necessary for the synthesis of polysaccharide B, one of the two capsular polysaccharides of B. fragilis 9343. The virulence of the polysaccharide B-deficient mutant was comparable to that of the wild type in terms of its ability to induce abscesses in a rat model of intra-abdominal infection.

Nonlinear models for in vitro kill kinetics of antibiotics.

In this study, we developed nonlinear regression models to analyze the data generated from an in vitro continuous culture system to assess the kinetics of metronidazole and trovafloxacin in inhibiting the growth of Bacteroides fragilis. The model includes parameters describing the initial shock effect of an antibiotic pulse, the overall antibiotic wash-out rate from the system, and the long-term toxicity of the antibiotic in the environment after one pulse and before the next pulse.

A randomized trial of a computer-based intervention to reduce utilization of redundant laboratory tests.

PURPOSE: To determine the impact of giving physicians computerized reminders about apparently redundant clinical laboratory tests. SUBJECTS AND METHODS: We performed a prospective randomized controlled trial that included all inpatients at a large teaching hospital during a 15-week period. The intervention consisted of computerized reminders at the time a test was ordered that appeared to be redundant. Main outcome measures were the proportions of clinical laboratory orders that were canceled and the proportion of the tests that were actually performed. RESULTS: During the study period, there were 939 apparently redundant laboratory tests among the 77,609 study tests that were ordered among the intervention (n = 5,700 patients) and control (n = 5,886 patients) groups. In the intervention group, 69% (300 of 437) of tests were canceled in response to reminders. Of 137 overrides, 41% appeared to be justified based on chart review. In the control group, 51% of ordered redundant tests were performed, whereas in the intervention group only 27% of ordered redundant tests were performed (P <0.001). However, the estimated annual savings in laboratory charges was only $35,000. This occurred because only 44% of redundant tests performed had computer orders, because only half the computer orders were screened for redundancy, and because almost one-third of the reminders were overridden. CONCLUSIONS: Reminders about orders for apparently redundant laboratory tests were effective when delivered. However, the overall effect was limited because many tests were performed without corresponding computer orders, and many orders were not screened for redundancy.

Correlation of cecal microflora of HLA-B27 transgenic rats with inflammatory bowel disease.

Transgenic rats with a high level of expression of the human major histocompatibility complex class I molecule HLA-B27 develop chronic inflammatory bowel disease (IBD) and arthritis. Assessment of the cecal microflora showed a rise in numbers of Escherichia coli and Enterococcus spp., corresponding to the presence and severity of IBD in these rats.

Epidemiology of a dominant clonal strain of vancomycin-resistant Enterococcus faecium at separate hospitals in Boston, Massachusetts.

In 1996, the dominant (43%) strain of vancomycin-resistant enterococci (VRE; type A) at Massachusetts General Hospital was identified at Brigham and Women's Hospital (BWH). To characterize the epidemiology of infection with type A isolates of VRE at BWH, we collected demographic and clinical data for all patients from whom VRE were isolated from a clinical specimen through September 1996. The first clinical isolates from all BWH patients from whom VRE were isolated were typed by pulsed-field gel electrophoresis of SmaI digests of chromosomal DNA. Among patients hospitalized after the first patient at BWH infected with a type A isolate of VRE was identified, exposures were compared between patients who acquired type A isolates of VRE and those who acquired other types of VRE. Isolates from 99 patients identified to have acquired VRE were most commonly from blood (n = 27), urine (n = 19), or wounds (n = 19). Three months after the index patient arrived at BWH and at a time when > or =12 types of strains of VRE were present, type A isolates of VRE became dominant; 39 of 75 (52%) of the study cohort had acquired type A isolates of VRE. We found no association between the acquisition of type A isolates of VRE and transfer from another institution or temporal overlap by service, ward, or floor with patients known to have acquired type A isolates of VRE. By multivariate analysis, only residence in the medical intensive care unit (adjusted odds ratio [OR], 3.2; 95% confidence interval [CI], 1.4 to 107) and the receipt of two or more antibiotics per patient-day (adjusted OR, 12.2; 95% CI, 1.2 to 9.0) were associated with the acquisition of strain A. This strain of VRE, dominant at two Boston hospitals, was associated with intensity of antibiotic exposures (i.e., two or more antibiotics per patient-day). We hypothesize that this strain may have unidentified properties providing a mechanism favoring its spread and dominance over other extant isolates, and further studies are needed to define these properties.

Cellular mechanism of intraabdominal abscess formation by Bacteroides fragilis.

We investigated the cellular mechanism by which Bacteroides fragilis promotes the development of intraabdominal abscesses in experimental models of sepsis. B. fragilis, as well as purified capsular polysaccharide complex (CPC) from this organism, adhered to primary murine mesothelial cells (MMCs) in vitro. The binding of CPC to murine peritoneal macrophage stimulated TNF-alpha production, which when transferred to monolayers of MMCs elicited significant ICAM-1 expression by these cells. This response resulted in enhanced polymorphonuclear leukocyte attachment to MMCs that could be inhibited by Abs specific for TNF-alpha or ICAM-1. Mice treated with TNF-alpha- or ICAM-1-specific Abs failed to develop intraabdominal abscesses following challenge with purified CPC. These results illustrated the role of the CPC in promoting adhesion of B. fragilis to the peritoneal wall and coordinating the cellular events leading to the development of abscesses associated with experimental intraabdominal sepsis.

Pharmacodynamics and microbiology of trovafloxacin in animal models of surgical infection.

Trovafloxacin provides broad in vitro and in vivo coverage of the aerobic and anaerobic pathogens found frequently in surgical infections. In vitro susceptibility testing indicated that trovafloxacin inhibited gram-positive staphylococci and enterococci, numerous gram-negative organisms, including Escherichia coli, and anaerobic pathogens, such as Bacteroides fragilis. Trovafloxacin protected mice from lethal infections induced by gram-negative or gram-positive organisms, even when these organisms were inoculated in combination with B. fragilis. Trovafloxacin protected rats in models of intra-abdominal sepsis induced by inoculation with E. coli and B. fragilis or with multiple aerobic and anaerobic pathogens. In these experimental models, trovafloxacin protected rats from lethal infection, reduced intra-abdominal abscess formation, and inhibited bacterial growth. Drug concentrations were greater in intra-abdominal abscesses than in serum, reflecting the good tissue penetration of trovafloxacin. These results indicate that trovafloxacin may be effective in prophylaxis and treatment of mixed infections in surgical patients.