An iron-chelating sulfonamide identified from Drosophila-based screening for antipathogenic discovery.

We exploited bacterial infection assays using the fruit fly Drosophila melanogaster to identify anti-infective compounds that abrogate the pathological consequences in the infected hosts. Here, we demonstrated that a pyridine-3-N-sulfonylpiperidine derivative (4a) protects Drosophila from the acute infections caused by bacterial pathogens including Pseudomonas aeruginosa. 4a did not inhibit the growth of P. aeruginosa in vitro, but inhibited the production of secreted toxins such as pyocyanin and hydrogen cyanide, while enhancing the production of pyoverdine and pyochelin, indicative of iron deprivation. Based on its catechol moiety, 4a displayed iron-chelating activity in vitro toward both iron (II) and iron (III), more efficiently than the approved iron-chelating drugs such as deferoxamine and deferiprone, concomitant with more potent antibacterial efficacy in Drosophila infections and unique transcriptome profile. Taken together, these results delineate a Drosophila-based strategy to screen for antipathogenic compounds, which interfere with iron uptake crucial for bacterial virulence and survival in host tissues.

Crystal structures of YeiE from Cronobacter sakazakii and the role of sulfite tolerance in gram-negative bacteria.

SignificanceYeiE has been identified as a master virulence factor of Cronobacter sakazakii. In this study, we determined the crystal structures of the regulatory domain of YeiE in complex with its physiological ligand sulfite ion (SO32-). The structure provides the basis for the molecular mechanisms for sulfite sensing and the ligand-dependent conformational changes of the regulatory domain. The genes under the control of YeiE in response to sulfite were investigated to reveal the functional roles of YeiE in the sulfite tolerance of the bacteria. We propose the molecular mechanism underlying the ability of gram-negative pathogens to defend against the innate immune response involving sulfite, thus providing a strategy to control the pathogenesis of bacteria.

Artemisinin displays bactericidal activity via copper-mediated DNA damage.

Artemisinin (ARS) and its semi-synthetic derivatives are effective drugs to treat malaria and possess multiple therapeutic activities based on their endoperoxide bridge. Here, we showed that ARS displayed antibacterial efficacy in Drosophila systemic infections caused by bacterial pathogens but killed only Vibrio cholerae (VC) in vitro, involving reactive oxygen species (ROS) generation and/or DNA damage. This selective antibacterial activity of ARS was attributed to the higher intracellular copper levels in VC, in that the antibacterial activity was observed in vitro upon addition of cuprous ions even against other bacteria and was compromised by the copper-specific chelators neocuproine (NC) and triethylenetetramine (TETA) in vitro and in vivo. We suggest that copper can enhance or reinforce the therapeutic activities of ARS to be repurposed as an antibacterial drug for the treatment of bacterial infections.

An antipathogenic compound that targets the OxyR peroxide sensor in Pseudomonas aeruginosa.

Introduction. Antipathogenic or antivirulence strategy is to target a virulence pathway that is dispensable for growth, in the hope to mitigate the selection for drug resistance.Hypothesis/Gap Statment. Peroxide stress responses are one of the conserved virulence pathways in bacterial pathogens and thus good targets for antipathogenic strategy.Aim. This study aims to identify a new chemical compound that targets OxyR, the peroxide sensor required for the full virulence of the opportunistic human pathogen, Pseudomonas aeruginosa.Methodology. Computer-based virtual screening under consideration of the 'eNTRy' rules and molecular docking were conducted on the reduced form of the OxyR regulatory domain (RD). Selected hits were validated by their ability to phenocopy the oxyR null mutant and modulate the redox cycle of OxyR.Results. We first isolated three robust chemical hits that inhibit OxyR without affecting prototrophic growth or viability. One (compound 1) of those affected the redox cycle of OxyR in response to H2O2 treatment, in a way to impair its function. Compound 1 displayed selective antibacterial efficacy against P. aeruginosa in Drosophila infection model, without antibacterial activity against Staphylococcus aureus.Conclusion. These results suggest that compound 1 could be an antipathogenic hit inhibiting the P. aeruginosa OxyR. More importantly, our study provides an insight into the computer-based discovery of new-paradigm selective antibacterials to treat Gram-negative bacterial infections presumably with few concerns of drug resistance.

Phage-Derived Antibacterials: Harnessing the Simplicity, Plasticity, and Diversity of Phages.

Despite the successful use of antibacterials, the emergence of multidrug-resistant bacteria has become a serious threat to global healthcare. In this era of antibacterial crisis, bacteriophages (phages) are being explored as an antibacterial treatment option since they possess a number of advantages over conventional antibacterials, especially in terms of specificity and biosafety; phages specifically lyse target bacteria while not affecting normal and/or beneficial bacteria and display little or no toxicity in that they are mainly composed of proteins and nucleic acids, which consequently significantly reduces the time and cost involved in antibacterial development. However, these benefits also create potential issues regarding antibacterial spectra and host immunity; the antibacterial spectra being very narrow when compared to those of chemicals, with the phage materials making it possible to trigger host immune responses, which ultimately disarm antibacterial efficacy upon successive treatments. In addition, phages play a major role in horizontal gene transfer between bacterial populations, which poses serious concerns for the potential of disastrous consequences regarding antibiotic resistance. Fortunately, however, recent advancements in synthetic biology tools and the speedy development of phage genome resources have allowed for research on methods to circumvent the potentially disadvantageous aspects of phages. These novel developments empower research which goes far beyond traditional phage therapy approaches, opening up a new chapter for phage applications with new antibacterial platforms. Herein, we not only highlight the most recent synthetic phage engineering and phage product engineering studies, but also discuss a new proof-of-concept for phage-inspired antibacterial design based on the studies undertaken by our group.

Investigation of the Diagnostic Value of Ultrasonography for Radial Neuropathy Located at the Spiral Groove.

OBJECTIVE: To determine a diagnostic cut-off value for the cross-sectional area (CSA) of the radial nerve using ultrasonography for radial neuropathy located at the spiral groove (SG). METHODS: Seventeen patients with electrodiagnostic evidence of radial neuropathy at the SG and 30 healthy controls underwent ultrasonography of the radial nerve at the SG . The CSAs at the SG were compared in the patient and control groups. The CSA at the SG between the symptomatic and asymptomatic sides (ΔSx-Asx and Sx/Asx, respectively) were analyzed to obtain the optimal cut-off value. The relationship between the electrophysiological severity of radial neuropathy and CSA was also evaluated. RESULTS: Among the variables examined, there were statistically significant differences in the CSA between the patient and control groups, ΔSx-Asx, and Sx/Asx at the SG. In a receiver operating characteristics analysis, the cut-off CSA was 5.75 mm2 at the SG (sensitivity 52.9%, specificity 90%), 1.75 mm2 for ΔSx-Asx (sensitivity 58.8%, specificity 100%), and 1.22 mm2 for Sx/Asx (sensitivity 70.6%, specificity 93.3%) in diagnosing radial neuropathy at the SG. There was no significant correlation between CSA and electrophysiological severity score for either patient group. CONCLUSION: The reference value obtained for CSA of the radial nerve at the SG may facilitate investigation of radial nerve pathologies at the SG.

Reduced Diaphragm Excursion During Reflexive Citric Acid Cough Test in Subjects With Subacute Stroke.

BACKGROUND: Diaphragm excursion is limited during respiratory maneuvers after a stroke. How the diaphragm is limited during reflexive coughs and affects the effectiveness of cough in stroke patients is unclear. This study aimed to measure reflexive cough strength by cough peak flow (CPF) induced by citric acid nebulization (2.8 mol/L), record diaphragm excursions during reflexive coughs in stroke subjects at risk of silent aspiration, and compare these values with those of stroke subjects without risk of aspiration or dysphagia. METHODS: Twenty-one subjects with subacute stroke (mean stroke onset, 13.6 d) at risk of silent aspiration (penetration-aspiration scale, 8) and 21 stroke subjects without dysphagia or aspiration (controls) were included. Diaphragmatic excursions were assessed using real-time sonography in all subjects; the main outcome measure was reflexive CPF induced by citric acid nebulization. RESULTS: The median (interquartile range) values of citric acid-induced CPF values were significantly more reduced in the 21 subjects with silent aspiration (45 [0-83] L/min) than in the control subjects (97 [66-162] L/min) (P = .004). Diaphragmatic excursions during the reflexive coughs were also significantly reduced (P < .001), although both groups had a similar range in the initial National Institutes of Health Stroke Scale scores and level of disability, as measured by the modified Barthel index. Citric acid-induced CPF was significantly correlated with the number of generated coughs (rs = 0.69), voluntary cough CPF (rs = 0.85), and degree of diaphragm excursion on both sides (rs = 0.50 [hemiplegic] and rs = 0.55 [nonhemiplegic]) but not correlated with the degree of hemiparesis, National Institutes of Health Stroke Scale score, or modified Barthel index scores. The 6-month follow up revealed that 7 subjects in group A experienced aspiration pneumonia. CONCLUSIONS: Stroke subjects at risk of silent aspiration showed reduced CPF and more limited diaphragm excursion during the citric acid-induced reflexive cough test. (ClinicalTrials.gov registration NCT02080988.).

Accuracy of Ultrasound-Guided and Non-ultrasound-Guided Botulinum Toxin Injection Into Cadaver Salivary Glands.

OBJECTIVE: To compare the accuracy of ultrasound (US)-guided and non-US-guided botulinum toxin (BTX) injection into the salivary glands (parotid and submandibular glands) of cadavers. METHODS: Two rehabilitation physician injected dye into three sites in the salivary glands (two sites in the parotid gland and one site in the submandibular gland) on one side of each cadaver (one was injected on the right side, while the other was injected on the left side), using either a non-US-guided injection procedure based on superficial landmarks or a US-guided procedure. Orange dye was used for the US-guided procedure, and green dye was used for the blind procedure. Two physicians uninvolved with the injection procedures and who were blinded to the method of injection dissected the cadavers to identify whether the dye was accurately injected into each target site. RESULTS: The accuracies of the blind and US-guided injections into the parotid gland were 79.17% and 95.83%, respectively. In the submandibular gland, the accuracies of the blind and US-guided injections were 50.00% and 91.67%, respectively. The difference in accuracy between the two procedures was statistically significant only in the submandibular gland (p=0.025). There were no significant differences in the accuracy of US-guided and non-US-guided injections between the two physicians for the two sites in the parotid gland (p=0.278 and p=0.146, respectively). CONCLUSION: US-guided BTX injection into the submandibular gland offers significantly greater accuracy over blind injection. For the treatment of drooling by injecting BTX into the submandibular gland, clinicians should consider using US guidance for improved accuracy.