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Background: Periprosthetic joint infections (PJI) of the shoulder are a devastating complication of shoulder arthroplasty and are commonly caused by Staphylococcus and Cutibacterium acnes. Absorbable calcium sulfate (CS) beads are sometimes used for delivering antibiotics in PJI. This study evaluates the in vitro effect of different combinations of gentamicin, vancomycin, and ertapenem in beads made from CS cement on the growth of C acnes and coagulase-negative Staphylococcus (CNS) strains., Methods: Three strains of C acnes and 5 strains of CNS from clinically proven shoulder PJI were cultured and plated with CS beads containing combinations of vancomycin, gentamicin, and ertapenem. Plates with C acnes were incubated anaerobically while plates with Staphylococcus were incubated aerobically at 37 °C. Zones of inhibition were measured at intervals of 3 and 7 days using a modified Kirby Bauer technique, and beads were moved to plates containing freshly streaked bacteria every seventh day. This process was run in triplicate over the course of 56 days. Statistical analysis was conducted using SPSS v. 28 with repeated measures analysis of variance (ANOVA) and pairwise comparisons with Tukey correction., Results: In experiments with C acnes, beads containing ertapenem + vancomycin and vancomycin alone formed the largest zones of inhibition over time (P < .001). In experiments with Staphylococcus, beads containing vancomycin alone formed the largest zones of inhibition over time for all 5 strains (P < .001). Zones of inhibition were 1.4x larger for C acnes than for Staphylococcus with beads containing vancomycin alone. For both C acnes and Staphylococcus, beads containing ertapenem had the strongest initial effect, preventing all bacterial growth in C acnes and almost all growth for Staphylococcus during the first week but dropping substantially by the second week. Beads containing gentamicin alone consistently created smaller zones of inhibition than beads containing vancomycin alone, with vancomycin producing zones 5.3x larger than gentamicin in C acnes and 1.3x larger in Staphylococcus (P < .001)., Discussion: These data suggest that for both C acnes and Staphylococcal species, CS beads impregnated with vancomycin were most effective at producing a robust antibiotic effect. Additionally, ertapenem may be a viable supplement in order to create a more potent initial antibiotic effect but is not as effective as vancomycin when used alone. Gentamicin alone was not effective in maintaining consistent and long-term antibiotic effects. These results indicate that amongst the antibiotics currently commercially available to be used with CS, vancomycin is consistently superior to gentamicin in the setting of C. acnes and CNS., (Copyright © 2024 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

Our laboratory recently reported that induction of the SOS response, triggered by SOS-inducing drugs, was accompanied by a large release of DNA from enteric bacteria. The SOS response release had not previously been reported to include release of extracellular DNA from bacterial cells. We followed up on those observations in this current study and found that not just double-stranded DNA was being released, but also single-stranded DNA, RNA, and protein. SOS-inducing drugs also triggered formation of biofilm at the air-fluid interface on glass, and the biofilms contained DNA. We extended our study to test whether inhibitors of the SOS response would block DNA release and found that SOS inhibitors, including zinc salts, nitric oxide donors, and dequalinium, inhibited SOS-induced DNA release. The understanding that SOS-induced DNA release is associated with formation of biofilms increases our appreciation of the role of the SOS response in pathogenesis, as well as in emergence of new antibiotic resistance. Our findings with SOS inhibitors also suggest that regimens might be devised that could block the deleterious effects of the SOS response, at least temporarily, when this is desired.

The SOS response is a conserved stress response pathway that is triggered by DNA damage in the bacterial cell. Activation of this pathway can, in turn, cause the rapid appearance of new mutations, sometimes called hypermutation. We compared the ability of various SOS-inducing drugs to trigger the expression of RecA, cause hypermutation, and produce elongation of bacteria. During this study, we discovered that these SOS phenotypes were accompanied by the release of large amounts of DNA into the extracellular medium. The release of DNA was accompanied by a form of bacterial aggregation in which the bacteria became tightly enmeshed in DNA. We hypothesize that DNA release triggered by SOS-inducing drugs could promote the horizontal transfer of antibiotic resistance genes by transformation or by conjugation.

Background: Previous reports have differed as to whether nitric oxide inhibits or stimulates the SOS response, a bacterial stress response that is often triggered by DNA damage. The SOS response is an important regulator of production of Shiga toxins (Stx) in Shiga-toxigenic E. coli (STEC). In addition, the SOS response is accompanied by hypermutation, which can lead to de novo emergence of antibiotic resistance. We studied these effects in vitro as well as in vivo ., Results: Nitric oxide donors inhibited induction of the SOS response by classical inducers such as mitomycin C, ciprofloxacin, and zidovudine, as measured by assays for E. coli RecA. Nitric oxide donors also inhibited Stx toxin protein production as well as stx2 RNA in vitro and in vivo . In vivo experiments were performed with ligated ileal segments in the rabbit using a 20 h infection. The NO donor S-nitroso-acetylpenicillamine (SNAP) reduced hypermutation in vitro and in vivo , as measured by emergence of rifampin resistance. SNAP blocked the ability of the RecA protein to bind to single-stranded DNA in an electrophoretic mobility shift assay (EMSA) in vitro , an early event in the SOS response. The inhibitory effects of SNAP were additive with those of zinc acetate., Conclusions: Nitric oxide donors blocked the initiation step of the SOS response. Downstream effects of this blockade included inhibition of Stx production and of hypermutation. Infection of rabbit loops with STEC resulted in a downregulation, rather than stimulation, of nitric oxide host defenses at 20 h of infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Crane, Burke and Alvarado.)

Colonic epithelium-commensal interactions play a very important role in human health and disease development. Colonic mucus serves as an ecologic niche for a myriad of commensals and provides a physical barrier between the epithelium and luminal content, suggesting that communication between the host and microbes occurs mainly by soluble factors. However, the composition of epithelia-derived metabolites and how the commensal flora influences them is less characterized. Here, we used mucus-producing human adult stem cell-derived colonoid monolayers exposed apically to probiotic E. coli strain Nissle 1917 to characterize the host-microbial communication via small molecules. We measured the metabolites in the media from host and bacterial monocultures and from bacteria-colonoid co-cultures. We found that colonoids secrete amino acids, organic acids, nucleosides, and polyamines, apically and basolaterally. The metabolites from host-bacteria co-cultures markedly differ from those of host cells grown alone or bacteria grown alone. Nissle 1917 affects the composition of apical and basolateral metabolites. Importantly, spermine, secreted apically by colonoids, shows antibacterial properties, and inhibits the growth of several bacterial strains. Our data demonstrate the existence of a cross-talk between luminal bacteria and human intestinal epithelium via metabolites, which might affect the numbers of physiologic processes including the composition of commensal flora via bactericidal effects.

Several classes of non-antibiotic drugs, including psychoactive drugs, proton-pump inhibitors (PPIs), non-steroidal anti-inflammatory drugs (NSAIDs), and others, appear to have strong antimicrobial properties. We considered whether psychoactive drugs induce the SOS response in E. coli bacteria and, consequently, induce Shiga toxins in Shiga-toxigenic E. coli (STEC). We measured the induction of an SOS response using a recA-lacZ &nbsp; E. coli reporter strain, as RecA is an early, reliable, and quantifiable marker for activation of the SOS stress response pathway. We also measured the production and release of Shiga toxin 2 (Stx2) from a classic E. coli O157:H7 strain, derived from a food-borne outbreak due to spinach. Some, but not all, serotonin selective reuptake inhibitors (SSRIs) and antipsychotic drugs induced an SOS response. The use of SSRIs is widespread and increasing; thus, the use of these antidepressants could account for some cases of hemolytic-uremic syndrome due to STEC and is not attributable to antibiotic administration. SSRIs could have detrimental effects on the normal intestinal microbiome in humans. In addition, as SSRIs are resistant to environmental breakdown, they could have effects on microbial communities, including aquatic ecosystems, long after they have left the human body.

The SOS response to DNA damage is a conserved stress response in Gram-negative and Gram-positive bacteria. Although this pathway has been studied for years, its relevance is still not familiar to many working in the fields of clinical antibiotic resistance and stewardship. Under some conditions, the SOS response favors DNA repair and preserves the genetic integrity of the organism. On the other hand, the SOS response also includes induction of error-prone DNA polymerases, which can increase the rate of mutation, called the mutator phenotype or "hypermutation." As a result, mutations can occur in genes conferring antibiotic resistance, increasing the acquisition of resistance to antibiotics. Almost all of the work on the SOS response has been on bacteria exposed to stressors in vitro . In this study, we sought to quantitate the effects of SOS-inducing drugs in vivo , in comparison with the same drugs in vitro . We used a rabbit model of intestinal infection with enteropathogenic Escherichia coli strain E22. SOS-inducing drugs triggered the mutator phenotype response in vivo as well as in vitro . Exposure of E. coli strain E22 to ciprofloxacin or zidovudine, both of which induce the SOS response in vitro , resulted in increased antibiotic resistance to 3 antibiotics: rifampin, minocycline, and fosfomycin. Zinc was able to inhibit the SOS-induced emergence of antibiotic resistance in vivo , as previously observed in vitro . Our findings may have relevance in reducing the emergence of resistance to new antimicrobial drugs.

The advanced radiographic capability (ARC) laser system, part of the National Ignition Facility (NIF) at Lawrence Livermore National Laboratory, is a short-pulse laser capability integrated into the NIF. The ARC is designed to provide adjustable pulse lengths of ∼1-38 p s in four independent beamlets, each with energies up to 1 kJ (depending on pulse duration). A detailed model of the ARC lasers has been developed that predicts the time- and space-resolved focal spots on target for each shot. Measurements made to characterize static and dynamic wavefront characteristics of the ARC are important inputs to the code. Modeling has been validated with measurements of the time-integrated focal spot at the target chamber center (TCC) at low power, and the space-integrated pulse duration at high power, using currently available diagnostics. These simulations indicate that each of the four ARC beamlets achieves a peak intensity on target of up to a few 10 18 W / c m 2 .

Many surgeons continue to face the clinical dilemma of interpreting a positive aspiration or unexpected positive Cutibacterium acnes (C. acnes) culture. There are factors that complicate the interpretation of positive cultures including variations in both frequency of false positive cultures and virulence properties. As indices of virulence, hemolytic strains, from previously confirmed clinically infected shoulders, were compared with non-hemolytic isolates determined to be contaminants, by RNA-sequencing (RNA-Seq). Six C. acnes isolates from patients who underwent revision total shoulder arthroplasty (TSA) were identified based on previously described infection criteria. Three C. acnes isolates from each group underwent RNA-Seq. Differential gene expression analysis, principal component analysis (PCA), and heatmap analysis were used to determine the gene variation and patterning between the definite infection and probable contaminant isolates. Differential gene expression analysis identified genes that were differentially expressed between the isolates classified as definite infection and isolates classified as probable contaminants. PCA using a 500 gene subset of identified genes was able to find combinations of these genes that separated out the definite infection and probable contaminants isolates. The heatmap demonstrated similar gene expression in the three Definite Infections isolates, and significantly different expression when compared with the probable contaminant isolates. Clinical significance: C. acnes revision TSA isolates classified as definite infection and probable contaminant demonstrated a similar gene expression pattern to each respective group and different gene expression pattern when compared between groups. These findings indicate distinct differences in C. acnes strains associated with clinically relevant orthopedic TSA infections., (© 2020 Orthopaedic Research Society. Published by Wiley Periodicals LLC.)

Background Enthusiasm for the use of metal nanoparticles in human and veterinary medicine is high. Many articles describe the effects of metal nanoparticles on microbes in vitro, and a smaller number of articles describe effects on the immune system, which is the focus of this review. Methods Articles were retrieved by performing literature searches in Medline, of the National Institute of Medicine, as well as via Google Scholar. Results In vitro studies show that metal nanoparticles have antimicrobial effects. Some metal nanoparticles augment innate host immune defenses, such as endogenous antimicrobial peptides, and nitric oxide. Metal nanoparticles may also function as vaccine adjuvants. Metal nanoparticles can migrate to locations distant from the site of administration, however, requiring careful monitoring for toxicity. Conclusions Metal nanoparticles show a great deal of potential as immunomodulators, as well as direct antimicrobial effects. Before metal particles can be adopted as therapies; however, more studies are needed to determine how nanoparticles migrate though the body and on possible adverse effects.

Introduction: Cutibacterium acnes is gaining recognition as a leading pathogen after orthopaedic shoulder procedures. Photodynamic therapy, a combination of light and a photosensitizer, has demonstrated antimicrobial activity against C. acnes in the treatment of acne vulgaris. We sought to evaluate the effect of photodynamic therapy using blue light and photosensitizers on C. acnes isolates from shoulder prosthetic joint infections. Methods: C. acnes strains isolated from 19 patients with shoulder PJI were exposed to blue light alone (415 nm) or in combination with photosensitizers (fluorescein, riboflavin and demeclocycline). C. acnes strains were divided into 4 categories: Highly Sensitive (HS), Sensitive (S) , Weakly Sensitive (WS), Resistant to blue light. Results: 13 of 19 C. acnes strains (68% ) were S or HS to blue light alone. Of these 19 strains tested, 11 were tested with blue light and fluorescein or blue light plus riboflavin. Fluorescein (1 µg/mL) enhanced the effect of blue light in 6 of 11 strains (55%). Blue light plus riboflavin (10 µg/mL) resulted enhanced killing in 3 of 11 strains (27%), but produced a paradoxical photoprotective effect in 4 of 11 strains (36%), resulting in a net decrease compared to blue light alone. Demeclocycline, however, enhanced the effect of blue light in 16 of 17 strains (94 %). Conclusions: Blue light with the addition of photosensitizers killed C. acnes from periprosthetic shoulder infections in vitro , with demeclocycline having the most pronounced effect., Competing Interests: Competing Interests: The authors, their immediate family, and any research foundation with which they are affiliated have not received any financial payments or other benefits from any commercial entity related to the subject of this article. We have no competing interests to declare., (© The author(s).)

We report on the design, performance, and qualification of the injection laser system designed to deliver joule-level chirped pulse beamlets arranged in dual rectangular beam formats into two main laser amplifier beamlines of the National Ignition Facility. The system is designed to meet the requirements of the Advanced Radiographic Capability upgrade with features that deliver performance, adjustability, and long-term reliability.

Introduction: The purpose of this study was to further evaluate the pathogenicity of hemolytic and nonhemolytic phenotypes of Propionibacterium acnes (P acnes) isolates from shoulders of orthopaedic patients., Methods: Thirty-one patient records were reviewed, which had a positive P acnes shoulder culture from joint aspiration fluid and/or intraoperative tissues for demographics, clinical course, culture, and laboratory data. Patients were categorized as definite infection, probable infection, or probable contaminant. Antibiotic resistance patterns and hemolysis characteristics were subsequently analyzed., Results: Hemolysis demonstrated 100% specificity with a positive predictive value of 100% and 80% sensitivity with a negative predictive value of 73% for determining definite and probable infections. Hundred percent of the patients in the hemolytic group and only 27% of patients in the nonhemolytic group were classified as infected. Presenting inflammatory markers were markedly higher in the hemolytic group. Clindamycin resistance was found in 31% of the hemolytic strains, whereas no antibiotic resistance was observed in the nonhemolytic group., Conclusion: Hemolytic strains of P acnes exhibit enhanced pathogenicity to their host by eliciting a more prominent systemic inflammatory response, increased antibiotic resistance, and a more challenging clinical course. Hemolysis may serve as a specific marker for assisting in diagnosing true infection with P acnes., Level of Evidence: Level III retrospective comparative study.

The SOS response is a conserved response to DNA damage that is found in Gram-negative and Gram-positive bacteria. When DNA damage is sustained and severe, activation of error-prone DNA polymerases can induce a higher mutation rate than is normally observed, which is called the SOS mutator phenotype or hypermutation. We previously showed that zinc blocked the hypermutation response induced by quinolone antibiotics and mitomycin C in Escherichia coli and Klebsiella pneumoniae . In this study, we demonstrate that zinc blocks the SOS-induced development of chloramphenicol resistance in Enterobacter cloacae . Zinc also blocked the transfer of an extended spectrum beta-lactamase (ESBL) gene from Enterobacter to a susceptible E. coli strain. A zinc ionophore, zinc pyrithione, was ~100-fold more potent than zinc salts in inhibition of ciprofloxacin-induced hypermutation in E. cloacae . Other divalent metals, such as iron and manganese, failed to inhibit these responses. Electrophoretic mobility shift assays (EMSAs) revealed that zinc, but not iron or manganese, blocked the ability of the E. coli RecA protein to bind to single-stranded DNA, an important early step in the recognition of DNA damage in enteric bacteria. This suggests a mechanism for zinc's inhibitory effects on bacterial SOS responses, including hypermutation.

Infections of the central nervous system due to Candida albicans are uncommon and are usually only observed in special circumstances, such as following neurosurgery or penetrating head trauma, in immunosuppressed patients, premature infants or in patients with ventriculoperitoneal shunts. The author reports a case of an immunocompetent man who presented with a thoracic intraspinal abscess due to C. albicans Despite surgical drainage and 6 weeks of high-dose fluconazole therapy, the abscess extended and recurred in the cervical spine, requiring a second operation to arrest the infection., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Zinc inhibits the virulence of diarrheagenic E. coli by inducing the envelope stress response and inhibiting the SOS response. The SOS response is triggered by damage to bacterial DNA. In Shiga-toxigenic E. coli, the SOS response strongly induces the production of Shiga toxins (Stx) and of the bacteriophages that encode the Stx genes. In E. coli, induction of the SOS response is accompanied by a higher mutation rate, called the mutator response, caused by a shift to error-prone DNA polymerases when DNA damage is too severe to be repaired by canonical DNA polymerases. Since zinc inhibited the other aspects of the SOS response, we hypothesized that zinc would also inhibit the mutator response, also known as hypermutation. We explored various different experimental paradigms to induce hypermutation triggered by the SOS response, and found that hypermutation was induced not just by classical inducers such as mitomycin C and the quinolone antibiotics, but also by antiviral drugs such as zidovudine and anti-cancer drugs such as 5-fluorouracil, 6-mercaptopurine, and azacytidine. Zinc salts inhibited the SOS response and the hypermutator phenomenon in E. coli as well as in Klebsiella pneumoniae, and was more effective in inhibiting the SOS response than other metals. We then attempted to determine the mechanism by which zinc, applied externally in the medium, inhibits hypermutation. Our results show that zinc interferes with the actions of RecA, and protects LexA from RecA-mediated cleavage, an early step in initiation of the SOS response. The SOS response may play a role in the development of antibiotic resistance and the effect of zinc suggests ways to prevent it.

Background: Many studies have noted an increase in the number of recognized cases of invasive infections due to Propionibacterium acnes , especially after shoulder replacement surgery. The increase in the number of recognized cases of P. acnes , a nonspore-forming, anaerobic, Gram-positive organism, appears due to both an increase in the number of shoulder operations being performed and more specimens being sent for anaerobic cultures. Nevertheless, the optimal surgical and antibiotic management of P. acnes remains controversial., Methods: We tested the susceptibility of 106 P. acnes strains from sterile body sites collected at the Erie County Medical Center between 2012 and 2015, using Etest gradient antibiotic strips., Results: P. acnes is very susceptible to the penicillins and the first-generation cephalosporins. We noted an association between hemolytic phenotype on Brucella Blood Agar and clindamycin resistance., Conclusions: Antimicrobial susceptibility testing of P. acnes should no longer just be confined to the research laboratory but expanded and incorporated into routine microbiological evaluation of P. acnes . This would improve patient care as well as help clarify the relationship between hemolysis and clindamycin resistance., Competing Interests: TRD discloses a research grant and consulting fees from Zimmer-Biomet, outside the work presented here. Other authors disclose no potential conflicts of interest.

In previous work, we identified xanthine oxidase (XO) as an important enzyme in the interaction between the host and enteropathogenic Escherichia coli(EPEC) and Shiga-toxigenic E. coli(STEC). Many of the biological effects of XO were due to the hydrogen peroxide produced by the enzyme. We wondered, however, if uric acid generated by XO also had biological effects in the gastrointestinal tract. Uric acid triggered inflammatory responses in the gut, including increased submucosal edema and release of extracellular DNA from host cells. While uric acid alone was unable to trigger a chloride secretory response in intestinal monolayers, it did potentiate the secretory response to cyclic AMP agonists. Uric acid crystals were formed in vivo in the lumen of the gut in response to EPEC and STEC infections. While trying to visualize uric acid crystals formed during EPEC and STEC infections, we noticed that uric acid crystals became enmeshed in the neutrophilic extracellular traps (NETs) produced from host cells in response to bacteria in cultured cell systems and in the intestine in vivo Uric acid levels in the gut lumen increased in response to exogenous DNA, and these increases were enhanced by the actions of DNase I. Interestingly, addition of DNase I reduced the numbers of EPEC bacteria recovered after a 20-h infection and protected against EPEC-induced histologic damage., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Passive immunotherapy for established infections, as opposed to active immunization to prevent disease, remains a tiny niche in the world of antimicrobial therapies. Many of the passive immunotherapies currently available are directed against bacterial toxins, such as botulism, or are intended for agents of bioterrorism such as anthrax, which fortunately has remained rare. The emergence of Ebola virus and multi-drug resistant pathogens, however, may breathe new life into the immunotherapy field as researchers seek non-antibiotic interventions for infectious diseases.

Elevated minimum inhibitory concentrations (MICs) of vancomycin against meticillin-resistant Staphylococcus aureus (MRSA) and the emergence of heteroresistant S. aureus strains have led to increased use of anti-MRSA antibiotics other than vancomycin. Ceftaroline fosamil is a novel cephalosporin with activity against MRSA, but there are limited clinical data on its use for MRSA bacteraemia (MRSAB) and against strains exhibiting high vancomycin MICs (2-4 μg/mL). This multicentre, retrospective, case-control study compared the microbiological and clinical effectiveness of ceftaroline used after vancomycin failure with that of vancomycin-treated controls for the treatment of MRSA with vancomycin MICs ≥ 2 μg/mL. In total, 32 patients were matched 1:1 with respect to vancomycin MIC, age and origin of bacteraemia. In the ceftaroline group, patients received prior MRSA therapy for a median of 5 days [interquartile range (IQR), 3-15.8 days] prior to switching to ceftaroline. Median time to eradication of MRSA was significantly less after treatment with ceftaroline compared with vancomycin [4 days (IQR, 3-7.5 days) vs. 8 days (IQR, 5.8-19.5 days); P=0.02]. Both clinical success at the end of treatment and recurrence of MRSA at Day 7 were trending towards being inferior in the vancomycin group, although the results did not attain statistical significance [81% vs. 44% (P=0.06) and 6% vs. 38% (P=0.08), respectively]. Ceftaroline added at the point of vancomycin failure resolves MRSAB more rapidly and with a higher rate of clinical success, therefore ceftaroline should be considered as an alternative for these difficult-to-treat infections., (Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

Background: Zinc supplements can treat or prevent enteric infections and diarrheal disease. Many articles on zinc in bacteria, however, highlight the essential nature of this metal for bacterial growth and virulence, suggesting that zinc should make infections worse, not better. To address this paradox, we tested whether zinc might have protective effects on intestinal epithelium as well as on the pathogen., Results: Using polarized monolayers of T84 cells we found that zinc protected against damage induced by hydrogen peroxide, as measured by trans-epithelial electrical resistance. Zinc also reduced peroxide-induced translocation of Shiga toxin (Stx) across T84 monolayers from the apical to basolateral side. Zinc was superior to other divalent metals to (iron, manganese, and nickel) in protecting against peroxide-induced epithelial damage, while copper also showed a protective effect.The SOS bacterial stress response pathway is a powerful regulator of Stx production in STEC. We examined whether zinc's known inhibitory effects on Stx might be mediated by blocking the SOS response. Zinc reduced expression of recA, a reliable marker of the SOS. Zinc was more potent and more efficacious than other metals tested in inhibiting recA expression induced by hydrogen peroxide, xanthine oxidase, or the antibiotic ciprofloxacin. The close correlation between zinc's effects on recA/SOS and on Stx suggested that inhibition of the SOS response is one mechanism by which zinc protects against STEC infection., Conclusions: Zinc's ability to protect against enteric bacterial pathogens may be the result of its combined effects on host tissues as well as inhibition of virulence in some pathogens. Research focused solely on the effects of zinc on pathogenic microbes may give an incomplete picture by failing to account for protective effects of zinc on host epithelia.

Determining if a Propionibacterium acnes culture is a true infection or a contaminant remains a challenge. We conducted a study to distinguish between a true infection and a contaminated culture based on the P acnes hemolytic phenotype and clinical presentation. All P acnes strains were from orthopedic patients who had undergone arthroplasty or nonarthroplasty shoulder procedures. Hemolysis was determined according to P acnes growth on brucella blood agar plates after 48 to 72 hours. Each patient record that corresponded to the obtained P acnes strains was retrospectively reviewed for clinical data. An orthopedic surgeon involved in the care of the patients, but blinded to the hemolytic status of the bacteria, classified these infections as definite, likely, or unlikely. Of the 22 P acnes strains, 13 were hemolytic, and 9 were nonhemolytic. Of the 13 hemolytic strains, 10 were definite infections; only 3 of the 9 nonhemolytic strains were definite infections. Mean (SD) C-reactive protein level was significantly higher (P = .03) in the hemolytic group, 16 (11) mg/mL, than in the nonhemolytic group, 7.9 (10) mg/mL. A hemolytic phenotype of P acnes may represent a more pathogenic strain of bacteria, and may be more likely to be found in patients with a definite infection with P acnes rather than a contaminated culture.

Uric acid can be generated in the gastrointestinal (GI) tract from the breakdown of nucleotides ingested in the diet or from purines released from host cells as a result of pathogen-induced cell damage. Xanthine oxidase (XO) is the enzyme that converts hypoxanthine or xanthine into uric acid, a reaction that also generates hydrogen peroxide. It has been assumed that the product of XO responsible for the pro-inflammatory effects of this enzyme is hydrogen peroxide. Recent literature on uric acid, however, has indicated that uric acid itself may have biological effects. We tested whether uric acid itself has detectable pro-inflammatory effects using an in vivo model using ligated rabbit intestinal segments ("loops") as well as in vitro assays using cultured cells. Addition of exogenous uric acid increased the influx of heterophils into rabbit intestinal loops, as measured by myeloperoxidase activity. In addition, white blood cells adhered avidly to uric acid crystals, forming large aggregates of cells. Uric acid acts as a leukocyte chemoattractant in the GI tract. The role of uric acid in enteric infections and in non-infectious disorders of the GI tract deserves more attention.

Xanthine oxidase (XO) has been recognized as an important host defense enzyme for decades. In our recent study in Infection and Immunity, we found that enteropathogenic and Shiga-toxigenic E. coli (EPEC and STEC) were far more resistant to killing by the XO pathway than laboratory E. coli strains used in the past. Although XO plus hypoxanthine substrate rarely generated enough H 2O 2 to kill EPEC and STEC, the pathogens were able to sense the H2O2 and react to it with an increase in expression of virulence factors, most notably Shiga toxin (Stx). H 2O 2 produced by XO also triggered a chloride secretory response in T84 cell monolayers studied in the Ussing chamber. Adding exogenous XO plus its substrate in vivo did not decrease the number of STEC bacteria recovered from ligated intestinal loops, but instead appeared to worsen the infection and increased the amount of Stx2 toxin produced. XO plus hypoxanthine also increases the ability of Stx2 to translocate across intestinal monolayers. With regard to EPEC and STEC, the role of XO appears more complex and subtle than what has been reported in the past, since XO also plays a role in host-pathogen signaling, in regulating virulence in pathogens, in Stx production and in toxin translocation. Uric acid produced by XO may also be in itself an immune modulator in the intestinal tract.

Orthopedic surgeons at our institution have noticed an increase in the number of infections due to Propionibacterium acnes, especially following operations on the shoulder. We collected P. acnes isolates from our hospital microbiology laboratory for 1 year and performed antimicrobial susceptibility testing on 28 strains from the shoulder. Antibiotics with the lowest MIC values against P. acnes (MIC50 and MIC90) included penicillin G (0.006, 0.125), cephalothin (0.047 and 0.094), and ceftriaxone (0.016, 0.045), while others also showed activity. Strains resistant to clindamycin were noted.

Xanthine oxidase (XO), also known as xanthine oxidoreductase, has long been considered an important host defense molecule in the intestine and in breastfed infants. Here, we present evidence that XO is released from and active in intestinal tissues and fluids in response to infection with enteropathogenic Escherichia coli (EPEC) and Shiga-toxigenic E. coli (STEC), also known as enterohemorrhagic E. coli (EHEC). XO is released into intestinal fluids in EPEC and STEC infection in a rabbit animal model. XO activity results in the generation of surprisingly high concentrations of uric acid in both cultured cell and animal models of infection. Hydrogen peroxide (H(2)O(2)) generated by XO activity triggered a chloride secretory response in intestinal cell monolayers within minutes but decreased transepithelial electrical resistance at 6 to 22 h. H(2)O(2) generated by XO activity was effective at killing laboratory strains of E. coli, commensal microbiotas, and anaerobes, but wild-type EPEC and STEC strains were 100 to 1,000 times more resistant to killing or growth inhibition by this pathway. Instead of killing pathogenic bacteria, physiologic concentrations of XO increased virulence by inducing the production of Shiga toxins from STEC strains. In vivo, exogenous XO plus the substrate hypoxanthine did not protect and instead worsened the outcome of STEC infection in the rabbit ligated intestinal loop model of infection. XO released during EPEC and STEC infection may serve as a virulence-inducing signal to the pathogen and not solely as a protective host defense.

Purpose: To evaluate the use of FRAX® (Fracture risk assessment tool) for changes in bone health risk factors and treatment decision-making., Data Sources: A convenience sample of seventeen women, 50 years and older, English-speaking, generally healthy, with the ability to perform weight-bearing exercise, presenting for a DXA scan in a Midwestern city between August 2009 and November 2009, and not already being treated for osteoporosis or osteopenia. Self-administered diet and exercise questionnaires were completed by participants, followed by individual counseling related to FRAX® absolute risk and NOF guidelines. Questionnaires were repeated at approximately three months later, along with a short survey regarding the information's impact., Conclusions: FRAX® increased a participant's perception of future risk for osteoporosis and desire to change bone health habits. About 50% actually made changes in calcium and vitamin D consumption and weight-bearing exercise. FRAX® could not be applied to those with osteopenia of the spine only; and a variety of bone health risk factors not covered by FRAX® were identified., Implications for Practice: The FRAX® risk assessment tool can be useful to motivate clients to change bone health behavior. However, it has limitations in its use as a tool for whether or not to prescribe bisphosphonates., (©2012 The Author(s) Journal compilation ©2012 American Academy of Nurse Practitioners.)

Previously, our laboratories reported that zinc inhibited expression of several important virulence factors in enteropathogenic Escherichia coli (EPEC) and reduced EPEC-induced intestinal damage in vivo. Since EPEC is genetically related to Shiga-toxigenic E. coli (STEC), we wondered whether the beneficial effects of zinc extended to STEC as well. Treatment options for STEC infection are very limited, since antibiotics tend to exacerbate disease via enhanced toxin production, so a safe intervention for this infection would be welcome. In this study, we report that in STEC strains zinc inhibits adherence to cultured cells as well as expression of EHEC secreted protein A (EspA). In addition, zinc inhibits the expression of Shiga toxin (Stx) at both the protein and the RNA level. Zinc inhibits basal and antibiotic-induced Stx production and inhibits both Stx1 and Stx2 by ≥90% at a concentration of 0.4 mM zinc. Rabbit EPEC strains were selected for acquisition of Stx-encoding bacteriophages, and these rabbit STEC strains (designated RDEC-H19A and E22-stx2) were used to test the effects of zinc in vivo in ligated rabbit intestinal loops. In vivo, zinc reduced fluid secretion into loops, inhibited mucosal adherence, reduced the amount of toxin in the loops, and reduced STEC-induced histological damage (villus blunting). Zinc has beneficial inhibitory effects against STEC strains that parallel those observed in EPEC. In addition, zinc strongly inhibits Stx expression; since Stx is responsible for the extraintestinal effects of STEC infection, such as hemolytic-uremic syndrome (HUS), zinc might be capable of preventing severe sequelae of STEC infection.

In order to achieve the highest intensities possible with the short-pulse Advanced Radiographic Capability beamline at the National Ignition Facility (NIF), it will be necessary to phase the individual ARC apertures. This is made especially challenging because the design of ARC results in two laser beams with different dispersions sharing the same NIF aperture. The extent to which two beams with different dispersions can be phased with each other has been an open question. This paper presents results of an analysis showing that the different dispersion values that will be encountered by the shared-aperture beams will not preclude the phasing of the two beams. We also highlight a situation in which dispersion mismatch will prevent good phasing between apertures, and discuss the limits to which higher-order dispersion values may differ before the beams begin to dephase.

Enteropathogenic E. coli (EPEC) is a common cause of diarrhea in children in developing countries. After adhering to intestinal cells, EPEC secretes effector proteins into host cells, causing cell damage and eventually death. We previously showed that EPEC infection triggers the release of ATP from host cells and that ATP is broken down to ADP, AMP, and adenosine. Adenosine produced from the breakdown of extracellular ATP triggers fluid secretion in intestinal monolayers and may be an important mediator of EPEC-induced diarrhea. Here we examined whether adenosine has any effects on EPEC bacteria. Adenosine stimulated EPEC growth in several types of media in vitro. Adenosine also altered the pattern of EPEC adherence to cultured cells from a localized adherence pattern to a more diffuse pattern. Adenosine changed the expression of virulence factors in EPEC, inhibiting the expression of the bundle-forming pilus (BFP) and enhancing expression of the EPEC secreted proteins (Esps). In vivo, experimental manipulations of adenosine levels had strong effects on the outcome of EPEC infection in rabbit intestinal loops. In addition to its previously reported effects on host tissues, adenosine has strong effects on EPEC bacteria, stimulating EPEC growth, altering its adherence pattern, and changing the expression of several important virulence genes. Adenosine, like noradrenaline, is a small, host-derived molecule that is utilized as a signal by EPEC.

Enteropathogenic Escherichia coli (EPEC) infection triggers the release of ATP from host intestinal cells, and the ATP is broken down to ADP, AMP, and adenosine in the lumen of the intestine. Ecto-5'-nucleotidase (CD73) is the main enzyme responsible for the conversion of 5'-AMP to adenosine, which triggers fluid secretion from host intestinal cells and also has growth-promoting effects on EPEC bacteria. In a recent study, we examined the role of the host enzyme CD73 in EPEC infection by testing the effect of ecto-5'-nucleotidase inhibitors. Zinc was a less potent inhibitor of ecto-5'-nucleotidase in vitro than the nucleotide analog alpha,beta-methylene-ADP, but in vivo, zinc was much more efficacious in preventing EPEC-induced fluid secretion in rabbit ileal loops than alpha,beta-methylene-ADP. This discrepancy between the in vitro and in vivo potencies of the two inhibitors prompted us to search for potential targets of zinc other than ecto-5'-nucleotidase. Zinc, at concentrations that produced little or no inhibition of EPEC growth, caused a decrease in the expression of EPEC protein virulence factors, such as bundle-forming pilus (BFP), EPEC secreted protein A, and other EPEC secreted proteins, and reduced EPEC adherence to cells in tissue culture. The effects of zinc were not mimicked by other transition metals, such as manganese, iron, copper, or nickel, and the effects were not reversed by an excess of iron. Quantitative real-time PCR showed that zinc reduced the abundance of the RNAs encoded by the bfp gene, by the plasmid-encoded regulator (per) gene, by the locus for the enterocyte effacement (LEE)-encoded regulator (ler) gene, and by several of the esp genes. In vivo, zinc reduced EPEC-induced fluid secretion into ligated rabbit ileal loops, decreased the adherence of EPEC to rabbit ileum, and reduced histopathological damage such as villus blunting. Some of the beneficial effects of zinc on EPEC infection appear to be due to the action of the metal on EPEC bacteria as well as on the host.

Enteropathogenic Escherichia coli (EPEC) triggers a large release of adenosine triphosphate (ATP) from host intestinal cells and the extracellular ATP is broken down to adenosine diphosphate (ADP), AMP, and adenosine. Adenosine is a potent secretagogue in the small and large intestine. We suspected that ecto-5'-nucleotidase (CD73, an intestinal enzyme) was a critical enzyme involved in the conversion of AMP to adenosine and in the pathogenesis of EPEC diarrhea. We developed a nonradioactive method for measuring ecto-5'-nucleotidase in cultured T84 cell monolayers based on the detection of phosphate release from 5'-AMP. EPEC infection triggered a release of ecto-5'-nucleotidase from the cell surface into the supernatant medium. EPEC-induced 5'-nucleotidase release was not correlated with host cell death but instead with activation of phosphatidylinositol-specific phospholipase C (PI-PLC). Ecto-5'-nucleotidase was susceptible to inhibition by zinc acetate and by alpha,beta-methylene-adenosine diphosphate (alpha,beta-methylene-ADP). In the Ussing chamber, these inhibitors could reverse the chloride secretory responses triggered by 5'-AMP. In addition, alpha,beta-methylene-ADP and zinc blocked the ability of 5'-AMP to stimulate EPEC growth under nutrient-limited conditions in vitro. Ecto-5'-nucleotidase appears to be the major enzyme responsible for generation of adenosine from adenine nucleotides in the T84 cell line, and inhibitors of ecto-5'-nucleotidase, such as alpha,beta-methylene-ADP and zinc, might be useful for treatment of the watery diarrhea produced by EPEC infection.

Enterotoxigenic Escherichia coli (ETEC) and enteropathogenic E. coli (EPEC) are common causes of diarrhea in children in developing countries. Dual infections with both pathogens have been noted fairly frequently in studies of diarrhea around the world. In previous laboratory work, we noted that cholera toxin and forskolin markedly potentiated EPEC-induced ATP release from the host cell, and this potentiated release was found to be mediated by the cystic fibrosis transmembrane conductance regulator. In this study, we examined whether the ETEC heat-labile toxin (LT) or the heat-stable toxin (STa, also known as ST) potentiated EPEC-induced ATP release. We found that crude ETEC culture filtrates, as well as purified ETEC toxins, did potentiate EPEC-induced ATP release in cultured T84 cells. Coinfection of T84 cells with live ETEC plus EPEC bacteria also resulted in enhanced ATP release compared to EPEC alone. In Ussing chamber studies of chloride secretion, adenine nucleotides released from the host by EPEC also significantly enhanced the chloride secretory responses that were triggered by crude ETEC filtrates, purified STa, and the peptide hormone guanylin. In addition, adenosine and LT had additive or synergistic effects in inducing vacuole formation in T84 cells. Therefore, ETEC toxins and EPEC-induced damage to the host cell both enhance the virulence of the other type of E. coli. Our in vitro data demonstrate a molecular basis for a microbial interaction, which could result in increased severity of disease in vivo in individuals who are coinfected with ETEC and EPEC.

We previously reported that enteropathogenic Escherichia coli (EPEC) infection triggered a large release of ATP from the host cell that was correlated with and dependent on EPEC-induced killing of the host cell. We noted, however, that under some circumstances, EPEC-induced ATP release exceeded that which could be accounted for on the basis of host cell killing. For example, EPEC-induced ATP release was potentiated by noncytotoxic agents that elevate host cell cAMP, such as forskolin and cholera toxin, and by exposure to hypotonic medium. These findings and the performance of the EPEC espF mutant led us to hypothesize that the CFTR plays a role in EPEC-induced ATP release that is independent of cell death. We report the results of experiments using specific, cell-permeable CFTR activators and inhibitors, as well as transfection of the CFTR into non-CFTR-expressing cell lines, which incriminate the CFTR as a second pathway for ATP release from host cells. Increased ATP release via CFTR is not accompanied by an increase in EPEC adherence to transfected cells. The CFTR-dependent ATP release pathway becomes activated endogenously later in EPEC infection, and this activation is mediated, at least in part, by generation of extracellular adenosine from the breakdown of released ATP.

Enteropathogenic Escherichia coli (EPEC) is a common cause of diarrhea in children in developing countries. Protein kinase C (PKC), a serine- and threonine-directed protein kinase, is rapidly activated following EPEC infection and this is accompanied by its translocation to a membrane-bound location where it is tightly bound to phosphatidylserine (PS). EPEC infection causes host cell death, one of whose features is externalization of PS. We hypothesized that externalization of PS would be accompanied by externalization of PKC as well. We report that EPEC infection triggers the externalization of PKC to the outer surface of the host cell. Ecto-PKC remains firmly tethered to the cell but can be released by incubation with peptide or protein substrates for the enzyme. Ecto-PKC is intact and biologically active and able to phosphorylate protein substrates on the surface of the host cell. Phosphorylation of whole EPEC bacteria or EPEC-secreted proteins could not be detected. Externalization of PKC could be reproduced by the combination of an apoptotic stimulus (ultraviolet (UV) irradiation) and phorbol myristate acetate (PMA), a procedure which resulted in externalization of >25% of the total cellular content of PKC-alpha. In the presence of ATP, ecto-PKC inhibited UV-induced cell shrinkage, membrane blebbing, and propidium iodide uptake but not the activation of caspases 3 and 7. This is the first report that expression of an ecto-protein kinase is altered by a microbial pathogen and the first to note that externalization of PKC can accompany apoptosis.

Enteropathogenic Escherichia coli (EPEC) causes severe, watery diarrhea in children. We investigated ATP release during EPEC-mediated killing of human cell lines and whether released adenine nucleotides function as secretory mediators. EPEC triggered a release of ATP from all human cell lines tested: HeLa, COS-7, and T84 (colon cells) as measured using a luciferase kit. Accumulation of ATP in the supernatant medium was enhanced if an inhibitor of 5'-ectonucleotidase was included and was further enhanced if an ATP-regenerating system was added. In the presence of the inhibitor/regenerator, ATP concentrations in the supernatant medium reached 1.5-2 microM 4 h after infection with wild-type EPEC strains. In the absence of the inhibitor/regenerator system, extracellular ATP was rapidly broken down to ADP, AMP, and adenosine. Conditioned medium from EPEC-infected cells triggered a brisk chloride secretory response in intestinal tissues studied in the Ussing chamber (rabbit distal colon and T84 cell monolayers), whereas conditioned medium from uninfected cells and sterile filtrates of EPEC bacteria did not. The short-circuit current response to EPEC-conditioned medium was completely reversed by adenosine receptor blockers, such as 8-(p-sulfophenyl)-theophylline and MRS1754. EPEC killing of host cells releases ATP, which is broken down to adenosine, which in turn stimulates secretion via apical adenosine A2b receptors. These findings provide new insight into how EPEC causes watery diarrhea.

Enteropathogenic Escherichia coli (EPEC) causes diarrhoea in children in developing countries. Many EPEC genes involved in virulence are contained within the locus of enterocyte effacement (LEE), a large pathogenicity island. One of the genes at the far righthand end of the LEE encodes EspF, an EPEC secreted protein of unknown function. EspF, like the other Esps, is a substrate for secretion by the type III secretory system. Previous studies found that an espF mutant behaved as wild type in assays of adherence, invasion, actin condensation and tyrosine phosphorylation. As EPEC can kill host cells, we tested esp gene mutants for host cell killing ability. The espF mutant was deficient in host cell killing despite having normal adherence. The addition of purified EspF to tissue culture medium did not cause any damage to host cells, but expression of espF in COS or HeLa cells caused cell death. The mode of cell death in cells transfected with espF appeared to be pure apoptosis. EspF appears to be an effector of host cell death in epithelial cells; its proline-rich structure suggests that it may act by binding to SH3 domains or EVH1 domains of host cell signalling proteins.

Exploiting the energetic interaction of intense femtosecond laser pulses with deuterium clusters, it is possible to create conditions in which nuclear fusion results from explosions of these clusters. We have conducted high-resolution neutron time-of-flight spectroscopy on these plasmas and show that they yield fast bursts of nearly monochromatic fusion neutrons with temporal duration as short as a few hundred picoseconds. Such a short, nearly pointlike source now opens up the unique possibility of using these bright neutron pulses, either as a pump or a probe, to conduct ultrafast studies with neutrons.

The effect of butyrate on the response to guanylin and Escherichia coli heat-stable enterotoxin, STa, was assessed in T84 cells and Caco-2 cells, cultured colon cell lines possessing the guanylyl cyclase C which is the receptor for these peptides. Butyrate treatment of these cells resulted in an apparent increase in cyclic GMP (cGMP) accumulation when the cGMP content of cells and the supernatant medium was measured. Butyrate treatment did not change the guanylyl cyclase activity or (125)I-STa binding parameters in T84 cells, but the butyrate effect was completely blocked by cycloheximide. Butyrate did not have any effect on STa-stimulated cGMP accumulation in COS cells transfected with the human or porcine GC-C. Further experiments showed that butyrate treatment caused a large increase in the cGMP released into the culture medium, and in cells grown in polarized fashion in Transwell inserts, cGMP efflux was predominantly from the basolateral surface of the cell; intracellular cGMP was actually lowered by butyrate treatment. Exposure of T84 cells to butyrate had no effect on the disposition of cyclic AMP generated in response to forskolin. The effects of butyrate on cGMP were reversible within 24 h of butyrate withdrawal. In colon cells, butyrate treatment induced a previously undescribed, cGMP-specific efflux mechanism which lowered intracellular cGMP and elevated extracellular cGMP in response to peptide agonists such as guanylin and STa., (Copyright 2000 Academic Press.)

Food poisoning syndromes caused by four different bacteria are described. For all types, food kept at a permissive temperature allows growth of the vegetative forms of the bacteria and production of a toxin or toxins. The key features of these syndromes, as well as possible new trends of concern, are summarized in Table 1.

We report a case of Mycobacterium bovis BCG vertebral osteomyelitis in a 79-year-old man 2.5 years after intravesical BCG therapy for bladder cancer. The recovered isolate resembled M. tuberculosis biochemically, but resistance to pyrazinamide (PZA) rendered that diagnosis suspect. High-pressure liquid chromatographic studies confirmed the diagnosis of M. bovis BCG infection. The patient was originally started on a four-drug antituberculous regimen of isoniazid, rifampin, ethambutol, and PZA. When susceptibility studies were reported, the regimen was changed to isoniazid and rifampin for 12 months. Subsequently, the patient was transferred to a skilled nursing facility for 3 months, where he underwent intensive physical therapy. Although extravesical adverse reactions are rare, clinicians and clinical microbiologists need to be aware of the possibility of disseminated infection by M. bovis BCG in the appropriate setting of clinical history, physical examination, and laboratory investigation.

Enteropathogenic Escherichia coli (EPEC) is a cause of prolonged watery diarrhea in children in developing countries. The ability of EPEC to kill host cells was investigated in vitro in assays using two human cultured cell lines, HeLa (cervical) and T84 (colonic). EPEC killed epithelial cells as assessed by permeability to the vital dyes trypan blue and propidium iodide. In addition, EPEC triggered changes in the host cell, suggesting apoptosis as the mode of death; such changes included early expression of phosphatidylserine on the host cell surface and internucleosomal cleavage of host cell DNA. Genistein, an inhibitor of tyrosine kinases, and wortmannin, an inhibitor of host phosphatidylinositol 3-kinase, markedly increased EPEC-induced cell death and enhanced the features of apoptosis. EPEC-induced cell death was contact dependent and required adherence of live bacteria to the host cell. A quantitative assay for EPEC-induced cell death was developed by using the propidium iodide uptake method adapted to a fluorescence plate reader. With EPEC, the rate and extent of host cell death were less that what has been reported for Salmonella, Shigella, and Yersinia, three other genera of enteric bacteria known to cause apoptosis. However, rapid apoptosis of the host cell may not favor the pathogenic strategy of EPEC, a mucosa-adhering, noninvasive pathogen.