Discovery and structure-activity relationships of a novel oxazolidinone class of bacterial type II topoisomerase inhibitors.

There is an increasingly urgent and unmet medical need for novel antibiotic drugs that tackle infections caused by multidrug-resistant (MDR) pathogens. Novel bacterial type II topoisomerase inhibitors (NBTIs) are of high interest due to limited cross-resistance with fluoroquinolones, however analogues with Gram-negative activity often suffer from hERG channel inhibition. A novel series of bicyclic-oxazolidinone inhibitors of bacterial type II topoisomerase were identified which display potent broad-spectrum anti-bacterial activity, including against MDR strains, along with an encouraging in vitro safety profile. In vivo proof of concept was achieved in a A. baumannii mouse thigh infection model.

Antibiofilm Efficacy of Polihexanide, Octenidine and Sodium Hypochlorite/Hypochlorous Acid Based Wound Irrigation Solutions against Staphylococcus aureus, Pseudomonas aeruginosa and a Multispecies Biofilm.

Infection and the formation of biofilms have been shown to have a significant role in increased inflammation and delayed wound healing. Wound irrigation solutions are used to debride wounds, removing cell debris and infecting microorganisms, therefore preventing infection. The aim of this study was to evaluate a Polihexanide (PHMB) based wound irrigation solution, Octenidine HCl based wound irrigation solution and electrolysed water based wound care solution for antibiofilm efficacy against Staphylococcus aureus, Pseudomonas aeruginosa and a multispecies biofilm in several models to gain a broad understanding of ability. The PHMB based wound irrigation solution demonstrated broad range antibiofilm efficacy against P. aeruginosa, S. aureus and the multispecies biofilm. The Octenidine HCl based wound irrigation solution and the electrolysed water based wound care solution demonstrated potent antibiofilm efficacy against S. aureus and to a lesser extent P. aeruginosa. Overall, less efficacy was observed in the drip flow bioreactor model for all 3 test solutions, which may be attributed to the continuous flow of nutrients during treatment, which may have diluted or washed away the solution. The data presented also highlights the importance of testing antibiofilm activity in a range of biofilm models and against different bacterial strains to get an overall representation of efficacy.

In Vitro Evaluation of Resistance Development to Silver Sulfadiazine and Subsequent Cross-Resistance to Antibiotics.

Treatment of chronic wounds that are at risk of infection, or that are infected, require the use of antimicrobial dressings, most often those that contain silver. Silver exerts its antimicrobial effects by binding to multiple cellular components and, as such, bacterial resistance to it is low; however, molecular silver resistance has been documented and is attributed to the presence of the sil operon or changes in genes encoding porin and efflux pump expression. The aim of this study was to evaluate spontaneous silver resistance development in common opportunistic pathogens, Staphylococcus, Pseudomonas and Enterococcus cloacae, as well as resistance development when exposed to subtherapeutic concentrations over a prolonged period. Furthermore, following silver resistance development, cross-resistance to several classes of antibiotics was evaluated. Following exposure of the strains to silver sulfadiazine (SSD) at two times and four times minimum inhibitory concentration (MIC), the mutation rate was <1010 colony forming unit (CFU)/mL. Serial passage of S. aureus and P. aeruginosa in subinhibitory concentrations of SSD selected for no resistant mutants. The SSD MIC of E. cloacae increased past the solubility limit of SSD at serial passage 17. MIC testing of this isolate showed a >2048-fold increase in MIC to silver in comparison to the parent strain. MIC testing of the serial passage isolates demonstrated no cross-resistance to antibiotics from six different classes. Overall, the results of this study show resistance development to silver is low and, if it does occur, it does not confer resistance to several antibiotic classes. However, as this study was carried out with a small number of strains, a study with a larger panel of strains and sequencing of the strains to determine the exact mechanism of resistance would be needed to investigate the threat of silver resistance further.

Controlled-release iodine foam dressings demonstrate broad-spectrum biofilm management in several in vitro models.

Multiple in vitro models were utilised to evaluate the biofilm management capabilities of seven commercially-available wound dressings, varying in composition and antibacterial ingredients, to reduce common aerobic, anaerobic, and multispecies biofilms. The Center for Disease Control bioreactor was used to evaluate single species Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) 24 and 48 hours biofilms, as well as a multispecies biofilm consisting of these two organisms in addition to Enterococcus faecalis (E. faecalis). As wound biofilms often exist in hypoxic wound environments, a direct contact anaerobic model system was used to evaluate efficacy on Bacteroides fragilis (B. fragilis). Biofilm control was evaluated against P. aeruginosa in the drip flow bioreactor model, where a constant flow of proteinaceous media is used to create a more challenging and wound-like model. The results demonstrated that biofilm management capabilities varied amongst wound dressings. Two dressings, a controlled-release iodine foam dressing and a silver nanocrystalline dressing, showed potent >4 log reductions in recovered organisms compared with untreated controls in all biofilm models evaluated. The effectiveness of other dressings to manage bioburden varied between dressing, test organism, and model system. A silver foam dressing showed moderate biofilm control in some models. However, biofilm exposure to methylene blue and gentian violet-containing foam dressings showed negligible log reductions in all in vitro biofilm methods examined. The data outlined in this in vitro study support the use of the iodine foam dressing for wounds with infection and biofilm.

The Ability of a Concentrated Surfactant Gel to Reduce an Aerobic, Anaerobic and Multispecies Bacterial Biofilm In Vitro.

Biofilm formation in wounds can lead to increased inflammation, infection and delayed wound healing. Additionally, biofilms show increased recalcitrance to antimicrobials compared to their planktonic counterparts making them difficult to manage and treat. Biofilms are frequently polymicrobial, consisting of aerobic and anaerobic bacteria, as well as fungi and yeasts. The aim of this study was to evaluate the effects of a concentrated surfactant gel with antibacterial preservative agents (CSG) against wound relevant opportunistic pathogens, including an aerobic biofilm, anaerobic biofilm and multispecies biofilm. The CSG was added to a 48 h anaerobic biofilm of Bacteroides fragilis, a 24 h multispecies biofilm of Acinetobacter baumannii, Staphylococcus aureus and Staphylococcus epidermidis and a 24 h biofilm of Pseudomonas aeruginosa grown in an in vitro wound relevant environment. Following a contact time of 24 h with the CSG, the bacterial cell density of the biofilms was reduced by 2-4 log in comparison to an untreated control. The results demonstrate the ability of the CSG to disrupt wound relevant biofilms and support the use of the CSG in the clinic to treat wounds caused by biofilm related infections.

Structural and mechanistic analysis of ATPase inhibitors targeting mycobacterial DNA gyrase.

OBJECTIVES: To evaluate the efficacy of two novel compounds against mycobacteria and determine the molecular basis of their action on DNA gyrase using structural and mechanistic approaches. METHODS: Redx03863 and Redx04739 were tested in antibacterial assays, and also against their target, DNA gyrase, using DNA supercoiling and ATPase assays. X-ray crystallography was used to determine the structure of the gyrase B protein ATPase sub-domain from Mycobacterium smegmatis complexed with the aminocoumarin drug novobiocin, and structures of the same domain from Mycobacterium thermoresistibile complexed with novobiocin, and also with Redx03863. RESULTS: Both compounds, Redx03863 and Redx04739, were active against selected Gram-positive and Gram-negative species, with Redx03863 being the more potent, and Redx04739 showing selectivity against M. smegmatis. Both compounds were potent inhibitors of the supercoiling and ATPase reactions of DNA gyrase, but did not appreciably affect the ATP-independent relaxation reaction. The structure of Redx03863 bound to the gyrase B protein ATPase sub-domain from M. thermoresistibile shows that it binds at a site adjacent to the ATP- and novobiocin-binding sites. We found that most of the mutations that we made in the Redx03863-binding pocket, based on the structure, rendered gyrase inactive. CONCLUSIONS: Redx03863 and Redx04739 inhibit gyrase by preventing the binding of ATP. The fact that the Redx03863-binding pocket is distinct from that of novobiocin, coupled with the lack of activity of resistant mutants, suggests that such compounds could have potential to be further exploited as antibiotics.

The Effects of a Concentrated Surfactant Gel on Biofilm EPS.

INTRODUCTION: The aim of this study was to evaluate if a poloxamer-based concentrated surfactant gel (CSG), containing antibacterial preservative agents, had the ability to reduce the levels of biofilm extracellular polymeric substances (EPS), specifically proteins and extracellular DNA (eDNA), as these are found to be the most immunogenic, within an in vitro biofilm. MATERIALS AND METHODS: A 24-hour biofilm of P. aeruginosa ATCC 15442 was grown in a 12-well plate and treated for 24 hours with a CSG coated onto Multisorb® (BSN Medical Limited, Hull, United Kingdom). EPS were extracted from each sample using 1M sodium chloride. Protein and DNA in EPS extractions was determined quantitatively using the Pierce™ Coomassie (Bradford) protein assay kit and a microplate SYTO 9™ (ThermoFisher Scientific, Paisley, United Kingdom) fluorescent assay, respectively. Protein and DNA was also determined qualitatively using confocal laser scanning microscopy (CLSM). RESULTS: Following 24-hour growth of P. aeruginosa ATCC 15442 biofilm, 7.38mg/mL protein was isolated from the extracted EPS in the untreated control. In comparison, the protein concentration found in the extracted EPS from biofilms treated with a CSG was 6.39mg/mL, showing a 13.4% reduction. Following 24-hour growth of P. aeruginosa ATCC 15442 biofilm, 11.71mg/mL eDNA was isolated from the extracted EPS in the untreated control. In comparison, the eDNA concentration found in the extracted EPS from biofilms treated with a CSG was 0.65mg/mL, showing a 94.5% reduction. Following statistical analysis of the data, the decrease in protein isolated following CSG treatment was within error; however, the decrease in eDNA isolated was statistically significant, showing the ability of the CSG to break up biofilm EPS in vitro. Using confocal laser microscopy and staining techniques, a large quantity of protein and eDNA could be observed in samples from the untreated control. In comparison, a reduction in EPS protein and eDNA was observed in samples that had been treated with a CSG. CONCLUSION: The data presented here potentially shows the ability of a CSG to reduce components of the P. aeruginosa biofilm EPS. The reduction in eDNA following CSG treatment may contribute to the dispersal of the biofilm, potentially increasing the susceptibility of it to antimicrobials, and should be explored further.

A comparative study on the cellular viability and debridement efficiency of antimicrobial-based wound dressings.

A concentrated surfactant gel containing polyhexamethylene biguanide (CSG-PHMB) (CSG: Plurogel) was evaluated for in vitro cell cytotoxicity using the direct contact, extraction, and cell insert assays, along with its ability to breakdown artificial wound eschar and slough, compared with other clinically available wound gels: a wound gel loaded with 0.13% benzalkonium chloride (BXG) and a highly viscous gel loaded with 0.1% polyhexamethylene biguanide (PXG). Following treatment with CSG-PHMB, BXG, and PXG at day 1, the viability of L929 and HDFa cells sharply decreased to lower than 20% of the culture media control in the direct contact assay; however, cell viability of L929 was 128.65 ± 1.41%, 99.90 ± 2.84%*, and 64.08 ± 5.99%* respectively; HDFa was 84.58 ± 10.41%, 19.54 ± 3.06%**, and 96.28 ± 33.67%, respectively, in the extraction assay. In the cell insert model, cell viability of L929 cells were 95.25 ± 0.96%, 47.49 ± 5.37%**, and 48.63 ± 7.00%**, respectively; HDFa cell viability were 92.80 ± 1.29%, 38.86 ± 4.28%**, and 49.90 ± 2.55%** (*: P < .01; **P < .001 compared with CSG-PHMB; cell viability of culture medium without treatment at day 1 was 100%). The cell extraction model on day 1 indicated that CSG-PHMB had higher viability of L929 cells compared with BXG. In addition, the cellular viability results indicated that CSG-PHMB gel exhibited lower cytotoxicity when compared with BXG and PXG in the cell insert model assay. Within the in vitro debridement model, CSG-PHMB exhibited an ability to potentially increase the loosening of the collagen matrix. The reason for this may be because of the concentrated surfactant found within the CSG-PHMB, which has the ability to lower the surface tension, aiding in the movements of fragments and debris in the fluorescent artificial wound eschar model (fAWE).

Silver, biofilms and wounds: resistance revisited.

Silver is added to an array of commercially available healthcare products including wound dressings. However, overuse of silver is being raised as a potential health concern due to the possible selection of tolerant or resistant bacteria and as a factor that may induce cross resistance to antibiotics. To date, there are only a limited number of studies that have documented evidence of silver resistance in bacteria isolated from medical situations. These studies have indicated low levels of silver resistance in bacteria. However, in comparison to antibiotics, only a small number of studies have been undertaken to investigate silver resistance. It is clear that more studies are required to confirm the most effective therapeutic levels of silver that are needed to kill microbes. In addition, it is probable that sub-therapeutic levels of silver may potentially select for enhanced microbial tolerance. Nevertheless, to date, there still remains very little evidence that silver resistance is a growing health concern in wound care; more studies are clearly needed to substantiate this concern, which has not been observed clinically to any major degree. The issue of biofilm tolerance to silver is more complicated and data on the effect of silver on biofilms is sparse at present.

The Efficacy of an Electrolysed Water Formulation on Biofilms.

Electrolysed water is a basic process whereby an electric current is passed through deionised water containing a low concentration of sodium chloride in an electrolysis chamber, which results in a more complex chemistry resulting in the production of a strong bactericidal and fungicidal solution at the anode. This microbicidal solution contains hypochlorous acid that is fast-acting and environmentally safe, as upon bacterial killing, the equilibrium shifts from hypochlorous acid back to salt and water. Other antimicrobial agents produced in this process include sodium hypochlorite and chlorine. The use of electrolysed water formulations in wound care to control wound bioburden is underway. However, there is limited evidence of the efficacy of electrolysed water on the control of biofilms, which are renowned for their tolerance to a variety of antimicrobials. Therefore this study aimed to assess a new electrolysed water formulation on in vitro Staphylococcus aureus and Pseudomonas aeruginosa biofilms. Results showed that the electrolysed water formulation effectively reduced biofilm in all models following a 15 min contact time. Microbial cell counts confirmed the reduction biofilm bacteria. Additional cytotoxicity using L929 fibroblasts confirmed that a 50% and 25% dilution of the electrolysed water formulation was non-cytotoxic to cells. In conclusion, this study has confirmed that the application of a new electrolysed water product effectively removed biofilm after a short exposure time. The use of this technology as a wound cleanser may help to control existing biofilms in complicated, non-healing wounds.

In vitro cellular viability studies on a concentrated surfactant-based wound dressing.

In this study, three cellular cytotoxic assays (direct contact assay, extraction assay, and cell insert assay) were applied to evaluate the effects of a concentrated surfactant gel preserved with antimicrobials and a concentrated surfactant gel with 1% silver sulfadiazine on both the mouse fibroblast cell line L929 and human dermal fibroblasts (HDFa). Also, the in vitro wound model was wounded by a 100 µL pipette tip and used to assess cell migration and wound closure after treatment with both gels. A needle-scratched membrane disruption model was used to preliminarily evaluate membrane stabilisation and the membrane-resealing effects of concentrated surfactant gels. It was demonstrated that the concentrated surfactant gel preserved with antimicrobials was not toxic to both L929 and HDFa. However, the concentrated surfactant gel with 1% silver sulfadiazine demonstrated a degree of cytotoxicity to both cell types. After treatment with a concentrated surfactant gel preserved with antimicrobials, cell movement to close the scratch gap was enhanced at 24 and 48 hours. The results also showed that cells treated with the concentrated surfactant gel preserved with antimicrobials decreased cell necrosis and improved cell resistance of the f-actin rearrangement after a needle scratch. The results demonstrated that a concentrated surfactant gel preserved with antimicrobials is non-cytotoxic and has ability to accelerate wound closure by enhancing cell mobility. Furthermore, the concentrated surfactant gel appeared to stabilise the plasma membrane and demonstrated a resealing ability and helped to retain the plasma membrane integrity and enhanced wound healing.

Design, synthesis and antibacterial properties of pyrimido[4,5-b]indol-8-amine inhibitors of DNA gyrase.

According to the World Health Organization (WHO), approximately 1.7 million deaths per year are caused by tuberculosis infections. Furthermore, it has been predicted that, by 2050, antibacterial resistance will be the cause of approximately 10 million deaths annually if the issue is not tackled. As a result, novel approaches to treating broad-spectrum bacterial infections are of vital importance. During the course of our wider efforts to discover unique methods of targeting multidrug-resistant (MDR) pathogens, we identified a novel series of amide-linked pyrimido[4,5-b]indol-8-amine inhibitors of bacterial type II topoisomerases. Compounds from the series were highly potent against gram-positive bacteria and mycobacteria, with excellent potency being retained against a panel of relevant Mycobacterium tuberculosis drug-resistant clinical isolates.

Tolerance of Biofilms to Antimicrobials and Significance to Antibiotic Resistance in Wounds.

A biofilm is a community of microorganisms that adhere to each other and to surfaces and secrete extracellular polymeric substances (EPS) encasing themselves in a matrix. Biofilms are a major healthcare concern, as they can form on medical devices leading to infection. Additionally, there is growing evidence to show their ability to form in chronic wounds, which leads to delayed wound healing and inflammation. Due to a number of reasons, such as formation of the EPS resulting in sub-inhibitory concentrations of antimicrobials reaching the bacterial cells, slow growth rate of bacterial cells rendering some antibiotics ineffective, and the presence of persister cells, biofilms show increased tolerance to many antimicrobials and antibiotics. Additionally, studies have started to emerge showing a link between resistance to antimicrobials and antibiotics. Cross-resistance can be attributed to a number of factors, for example, increased expression of multidrug efflux pumps that efflux a wide range of substrates and horizontal gene transfer of genetic material encoding multiple resistance genes between different species within the polymicrobial biofilm. Antimicrobial resistance is an increasing threat caused by multiple factors including cross-resistance, and it is a global health concern. This review focuses on current research on antimicrobial and antibiotic resistance and cross-resistance found between antimicrobials and antibiotics commonly used in woundcare to evaluate the significance of this acquired antibiotic resistance. Furthermore, the review discusses the significance of antimicrobial tolerance and the role biofilms play in enhancing antibiotic resistance.

Efficacy of a Surfactant-Based Wound Dressing in the Prevention of Biofilms.

OBJECTIVE: To assess the biofilm prevention action of two wound dressings, a concentrated surfactant gel preserved with antimicrobials and a concentrated surfactant gel with 1% silver sulfadiazine. METHODS: The microorganisms Staphylococcus aureus, methicillin-resistant S aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Enterococcus faecalis were used. Several biofilm models were used whereby the surfaces of each model were coated with either the concentrated surfactant gel preserved with antimicrobials or the concentrated surfactant gel with SSD before biofilm growth. MAIN RESULTS: Results showed the concentrated surfactant gel with SSD prevented biofilm growth in the modified minimum biofilm eradication concentration and Centers for Disease Control and Prevention biofilm models. The concentrated surfactant gel preserved with antimicrobials prevented microbial penetration for up to 48 hours, whereas the concentrated surfactant with SSD prevented microbial penetration for at least 72 hours. Using confocal laser scanning microscopy, researchers showed that a surface coated with the concentrated surfactant gel preserved with antimicrobials enhanced microbial sequestration of planktonic microorganisms. CONCLUSIONS: These results demonstrated that a concentrated surfactant gel preserved with antimicrobials can sequester and cause the immobilization of planktonic bacteria. Further, the concentrated surfactant gel with SSD can effectively kill planktonic and sessile microorganisms.

Mode of action of poloxamer-based surfactants in wound care and efficacy on biofilms.

Surfactants are widely used as detergents, emulsifiers, wetting agents, foaming agents, and dispersants in both the food and oil industry. Their use in a clinical setting is also common, particularly in wound care. Complicated or chronic wounds show clinical signs of delayed healing, persistent inflammation, and the production of non-viable tissue. These types of wounds also present challenges such as infection and potentially house antimicrobial-tolerant biofilms. The use of wound cleansers to aid cleaning and debridement of the wound is essential. A large proportion of skin and wound cleansers contain surfactants but there is only a small amount of data that shows the effectiveness of them in the enhancement of wound closure. This review paper aims to explore the available literature surrounding the use and mode of action of surfactants in wound healing, in particular Poloxamer 188 (Pluronic F-68) and Poloxamer 407 (Pluronic F-127), and also uncover the potential mechanisms behind the enhancement of wound healing and comparison to other surfactants used in wound care. Furthermore, the presence of a microbial biofilm in the wound is a significant factor in delayed wound healing. Therefore, the effect of clinically used surfactants on biofilms will be discussed, with emphasis on poloxamer-based surfactants.

Role of anaerobes in polymicrobial communities and biofilms complicating diabetic foot ulcers.

Infected tissues in the feet of people with diabetes in the form of a diabetic foot ulcer (DFU) present a complex pathology for clinicians to manage. This is partly attributed to the multi-factorial nature of the disease, which may include; altered foot architecture leading to excessive plantar pressures and frictional forces peripheral arterial disease and loss of protective sensation. In addition, to the above co-morbid variables, it is understood that a delayed wound healing state may be perpetuated by the presence of microorganisms residing in the wound tissue. The microbiology of chronic DFUs has often been reported as being polymicrobial. Of growing interest is the presence and potential role of anaerobic microorganisms in the pathology of DFUs and how they may contribute to the infective process or delayed healing. The presence of anaerobes in DFUs has been greatly underestimated, largely due to the limitations of conventional culture methods in identifying them from samples. Advancements in molecular and microscopy techniques have extended our view of the wound microbiome in addition to observing the growth and behaviour (planktonic or biofilm) of microorganisms in situ. This review paper will reflect on the evidence for the role and significance of anaerobes in DFUs and infection. A focus of this review will be to explore recent advancements in molecular genomics and microscopy techniques in order to better assess the roles of anaerobic bacteria in chronic DFUs and in biofilm-based wound care.

Absence of the Polar Organizing Protein PopZ Results in Reduced and Asymmetric Cell Division in Agrobacterium tumefaciens.

Agrobacterium tumefaciens is a rod-shaped bacterium that grows by polar insertion of new peptidoglycan during cell elongation. As the cell cycle progresses, peptidoglycan synthesis at the pole ceases prior to insertion of new peptidoglycan at midcell to enable cell division. The A. tumefaciens homolog of the Caulobacter crescentus polar organelle development protein PopZ has been identified as a growth pole marker and a candidate polar growth-promoting factor. Here, we characterize the function of PopZ in cell growth and division of A. tumefaciens Consistent with previous observations, we observe that PopZ localizes specifically to the growth pole in wild-type cells. Despite the striking localization pattern of PopZ, we find the absence of the protein does not impair polar elongation or cause major changes in the peptidoglycan composition. Instead, we observe an atypical cell length distribution, including minicells, elongated cells, and cells with ectopic poles. Most minicells lack DNA, suggesting a defect in chromosome segregation. Furthermore, the canonical cell division proteins FtsZ and FtsA are misplaced, leading to asymmetric sites of cell constriction. Together, these data suggest that PopZ plays an important role in the regulation of chromosome segregation and cell division.IMPORTANCEA. tumefaciens is a bacterial plant pathogen and a natural genetic engineer. However, very little is known about the spatial and temporal regulation of cell wall biogenesis that leads to polar growth in this bacterium. Understanding the molecular basis of A. tumefaciens growth may allow for the development of innovations to prevent disease or to promote growth during biotechnology applications. Finally, since many closely related plant and animal pathogens exhibit polar growth, discoveries in A. tumefaciens may be broadly applicable for devising antimicrobial strategies.

Polar Organizing Protein PopZ Is Required for Chromosome Segregation in Agrobacterium tumefaciens.

Despite being perceived as relatively simple organisms, many bacteria exhibit an impressive degree of subcellular organization. In Caulobacter crescentus, the evolutionarily conserved polar organizing protein PopZ facilitates cytoplasmic organization by recruiting chromosome centromeres and regulatory proteins to the cell poles. Here, we characterize the localization and function of PopZ in Agrobacterium tumefaciens, a genetically related species with distinct anatomy. In this species, we find that PopZ molecules are relocated from the old pole to the new pole in the minutes following cell division. PopZ is not required for the localization of the histidine kinases DivJ and PdhS1, which become localized to the old pole after PopZ relocation is complete. The histidine kinase PdhS2 is temporally and spatially related to PopZ in that it localizes to transitional poles just before they begin to shed PopZ and disappears from the old pole after PopZ relocalization. At the new pole, PopZ is required for tethering the centromere of at least one of multiple replicons (chromosome I), and the loss of popZ results in a severe chromosome segregation defect, aberrant cell division, and cell mortality. After cell division, the daughter that inherits polar PopZ is shorter in length and delayed in chromosome I segregation compared to its sibling. In this cell type, PopZ completes polar relocation well before the onset of chromosome segregation. While A. tumefaciens PopZ resembles its C. crescentus homolog in chromosome tethering activity, other aspects of its localization and function indicate distinct properties related to differences in cell organization.IMPORTANCE Members of the Alphaproteobacteria exhibit a wide range of phenotypic diversity despite sharing many conserved genes. In recent years, the extent to which this diversity is reflected at the level of subcellular organization has become increasingly apparent. However, which factors control such organization and how they have changed to suit different body plans are poorly understood. This study focuses on PopZ, which is essential for many aspects of polar organization in Caulobacter crescentus, but its role in other species is unclear. We explore the similarities and differences in PopZ functions between Agrobacterium tumefaciens and Caulobacter crescentus and conclude that PopZ lies at a point of diversification in the mechanisms that control cytoplasmic organization and cell cycle regulation in Alphaproteobacteria.

Novel Bacterial Topoisomerase Inhibitors with Potent Broad-Spectrum Activity against Drug-Resistant Bacteria.

The novel bacterial topoisomerase inhibitor class is an investigational type of antibacterial inhibitor of DNA gyrase and topoisomerase IV that does not have cross-resistance with the quinolones. Here, we report the evaluation of the in vitro properties of a new series of this type of small molecule. Exemplar compounds selectively and potently inhibited the catalytic activities of Escherichia coli DNA gyrase and topoisomerase IV but did not block the DNA breakage-reunion step. Compounds showed broad-spectrum inhibitory activity against a wide range of Gram-positive and Gram-negative pathogens, including biodefence microorganisms and Mycobacterium tuberculosis No cross-resistance with fluoroquinolone-resistant Staphylococcus aureus and E. coli isolates was observed. Measured MIC90 values were 4 and 8 µg/ml against a panel of contemporary multidrug-resistant isolates of Acinetobacter baumannii and E. coli, respectively. In addition, representative compounds exhibited greater antibacterial potency than the quinolones against obligate anaerobic species. Spontaneous mutation rates were low, with frequencies of resistance typically <10-8 against E. coli and A. baumannii at concentrations equivalent to 4-fold the MIC. Compound-resistant E. coli mutants that were isolated following serial passage were characterized by whole-genome sequencing and carried a single Arg38Leu amino acid substitution in the GyrA subunit of DNA gyrase. Preliminary in vitro safety data indicate that the series shows a promising therapeutic index and potential for low human ether-a-go-go-related gene (hERG) inhibition (50% inhibitory concentration [IC50], >100 µM). In summary, the compounds' distinct mechanism of action relative to the fluoroquinolones, whole-cell potency, low potential for resistance development, and favorable in vitro safety profile warrant their continued investigation as potential broad-spectrum antibacterial agents.

Efficacy of a surfactant-based wound dressing on biofilm control.

The aim of this study was to evaluate the efficacy of both a nonantimicrobial and antimicrobial (1% silver sulfadiazine-SSD) surfactant-based wound dressing in the control of Pseudomonas aeruginosa, Enterococcus sp, Staphylococcus epidermidis, Staphylococcus aureus, and methicillin-resistant S. aureus (MRSA) biofilms. Anti-biofilm efficacy was evaluated in numerous adapted American Standards for Testing and Materials (ASTM) standard biofilm models and other bespoke biofilm models. The ASTM standard models employed included the Minimum biofilm eradication concentration (MBEC) biofilm model (ASTM E2799) and the Centers for Disease Control (CDC) biofilm reactor model (ASTM 2871). Such bespoke biofilm models included the filter biofilm model and the chamberslide biofilm model. Results showed complete kill of microorganisms within a biofilm using the antimicrobial surfactant-based wound dressing. Interestingly, the nonantimicrobial surfactant-based dressing could disrupt existing biofilms by causing biofilm detachment. Prior to biofilm detachment, we demonstrated, using confocal laser scanning microscopy (CLSM), the dispersive effect of the nonantimicrobial surfactant-based wound dressing on the biofilm within 10 minutes of treatment. Furthermore, the non-antimicrobial surfactant-based wound dressing caused an increase in microbial flocculation/aggregation, important for microbial concentration. In conclusion, this nonantimicrobial surfactant-based wound dressing leads to the effective detachment and dispersion of in vitro biofilms. The use of surfactant-based wound dressings in a clinical setting may help to disrupt existing biofilm from wound tissue and may increase the action of antimicrobial treatment.