The QseEF Two-Component System-GlmY Small RNA Regulatory Pathway Controls Swarming in Uropathogenic Proteus mirabilis

Bacterial sensing of environmental signals through the two-component system (TCS) plays a key role in modulating virulence. In the search for the host hormone-sensing TCS, we identified a conserved qseEGF locus following glmY, a small RNA (sRNA) gene in uropathogenic Proteus mirabilis. Genes of glmY-qseE-qseG-qseF constitute an operon, and QseF binding sites were found in the glmY promoter region. Deletion of glmY or qseF resulted in reduced swarming motility and swarming-related phenotypes relative to the wild-type and the respective complemented strains. The qseF mutant had decreased glmYqseEGF promoter activity. Both glmY and qseF mutants exhibited decreased flhDC promoter activity and mRNA level, while increased rcsB mRNA level was observed in both mutants. Prediction by TargetRNA2 revealed cheA as the target of GlmY. Then, construction of the translational fusions containing various lengths of cheA 5′UTR for reporter assay and site-directed mutagenesis were performed to investigate the cheA-GlmY interaction in cheA activation. Notably, loss of glmY reduced the cheA mRNA level, and urea could inhibit swarming in a QseF-dependent manner. Altogether, this is the first report elucidating the underlying mechanisms for modulation of swarming motility by a QseEF-regulated sRNA GlmY, involving expression of cheA, rcsB and flhDC in uropathogenic P. mirabilis.

Anti-Pathogenic Properties of the Combination of a T3SS Inhibitory Halogenated Pyrrolidone with C-30 Furanone

Antimicrobial resistance is one of the current public health challenges to be solved. The World Health Organization (WHO) has urgently called for the development of strategies to expand the increasingly limited antimicrobial arsenal. The development of anti-virulence therapies is a viable option to counteract bacterial infections with the possibility of reducing the generation of resistance. Here we report on the chemical structures of pyrrolidones DEXT 1–4 (previously identified as furan derivatives) and their anti-virulence activity on Pseudomonas aeruginosa strains. DEXT 1–4 were shown to inhibit biofilm formation, swarming motility, and secretion of ExoU and ExoT effector proteins. Also, the anti-pathogenic property of DEXT-3 alone or in combination with furanone C-30 (quorum sensing inhibitor) or MBX-1641 (type III secretion system inhibitor) was analyzed in a model of necrosis induced by P. aeruginosa PA14. All treatments reduced necrosis; however, only the combination of C-30 50 µM with DEXT-3 100 µM showed significant inhibition of bacterial growth in the inoculation area and systemic dispersion. In conclusion, pyrrolidones DEXT 1–4 are chemical structures capable of reducing the pathogenicity of P. aeruginosa and with the potential for the development of anti-virulence combination therapies.

Surveying a Swarm: Experimental Techniques to Establish and Examine Bacterial Collective Motion

The survival and successful spread of many bacterial species hinges on their mode of motility. One of the most distinct of these is swarming, a collective form of motility where a dense consortium of bacteria employ flagella to propel themselves across a solid surface. Surface environments pose unique challenges, derived from higher surface friction/tension and insufficient hydration. Bacteria have adapted by deploying an array of mechanisms to overcome these challenges. Beyond allowing bacteria to colonize new terrain in the absence of bulk liquid, swarming also bestows faster speeds and enhanced antibiotic resistance to the collective. These crucial attributes contribute to the dissemination, and in some cases pathogenicity, of an array of bacteria. This mini-review highlights; 1) aspects of swarming motility that differentiates it from other methods of bacterial locomotion. 2) Facilitatory mechanisms deployed by diverse bacteria to overcome different surface challenges. 3) The (often difficult) approaches required to cultivate genuine swarmers. 4) The methods available to observe and assess the various facets of this collective motion, as well as the features exhibited by the population as a whole.

Halogen-Furan-2(5H)-One Derivatives Decrease Biofilm Production in Pseudomonas aeruginosa.

Clinical evidence has shown that bacterial infections are more difficult to eradicate when form-ing a biofilm aggregate than when are produced by bacteria in planktonic form. Therefore, com-pounds that inhibit biofilm formation could be used against severe infections. It has been re-ported that bromo 2-(5H) furanones inhibited biofilm formation by their anti-quorum sensing properties. To determine if the 2-(5H) furanone moiety is essential to induce inhibition of biofilm formation, we evaluated ten halogen 2-(5H) furanones derivates previously synthesized. Besides evaluating the inhibition of biofilm formation, we assessed pyocyanin production, swarming motility, and transcription of essential QS genes: rsaL, rhlA, pqsA and phz1 genes. Our results showed that although three bromo-furan-2(5H)-one-type derivatives (A1-A3) and two bromo-4-(phenylamino)-furan-2(5H)-one-type compounds (B2 and B6) inhibited the biofilm formation in both P. aeruginosa PA14 (reference) and PA64 (drug-resistant) strains only the furanones A1-A3 were efficient to inhibit QSS.

Social motility of biofilm-like microcolonies in a gliding bacterium

Bacterial biofilms are aggregates of surface-associated cells embedded in an extracellular polysaccharide (EPS) matrix, and are typically stationary. Studies of bacterial collective movement have largely focused on swarming motility mediated by flagella or pili, in the absence of a biofilm. Here, we describe a unique mode of collective movement by a self-propelled, surface-associated biofilm-like multicellular structure. Flavobacterium johnsoniae cells, which move by gliding motility, self-assemble into spherical microcolonies with EPS cores when observed by an under-oil open microfluidic system. Small microcolonies merge, creating larger ones. Microscopic analysis and computer simulation indicate that microcolonies move by cells at the base of the structure, attached to the surface by one pole of the cell. Biochemical and mutant analyses show that an active process drives microcolony self-assembly and motility, which depend on the bacterial gliding apparatus. We hypothesize that this mode of collective bacterial movement on solid surfaces may play potential roles in biofilm dynamics, bacterial cargo transport, or microbial adaptation. However, whether this collective motility occurs on plant roots or soil particles, the native environment for F. johnsoniae, is unknown.

Inhibition of Quorum Sensing and Biofilm Formation of Esculetin on Aeromonas Hydrophila

Quorum sensing (QS) and biofilm formation inhibition activity of esculetin on Aeromonas hydrophila SHAe 115 were evaluated. Exposure to esculetin at 25, 50, and 100μg/ml significantly inhibited the production of protease and hemolysin, the formation of biofilms and attenuated the swarming motility of A. hydrophila SHAe 115. Biofilm forming inhibition was also observed through confocal laser scanning microscopy and scanning electron microscope. Quantitative real-time PCR analysis indicated that genes positively related to QS and biofilm formation were downregulated to varying degrees, while gene (litR) negatively related to biofilm formation was significantly upregulated. The phenotypic results were in good agreement with gene expression levels. These results indicated that esculetin would be a potential QS inhibitor for A. hydrophila.

Chemical Composition, Anti-Quorum Sensing, Enzyme Inhibitory, and Antioxidant Properties of Phenolic Extracts of Clinopodium nepeta L. Kuntze

Phenolic extracts of Clinopodium nepeta were prepared and their preliminary phenolic profiles determined using HPLC-DAD with 26 phenolic standards. Apigenin (21.75 ± 0.41 µg/g), myricetin (72.58 ± 0.57 µg/g), and rosmarinic acid (88.51 ± 0.55 µg/g) were the most abundant compounds in DCM (dichloromethane), AcOEt (ethyl acetate), and BuOH (butanol) extracts, respectively. The DCM and AcOEt extracts inhibited quorum-sensing mediated violacein production by C. violaceum CV12472. Anti-quorum-sensing zones on C. violaceum CV026 at MIC (minimal inhibitory concentration) were 10.3 ± 0.8 mm for DCM extract and 12.0 ± 0.5 mm for AcOEt extract. Extracts showed concentration-dependent inhibition of swarming motility on flagellated P. aeruginosa PA01 and at the highest test concentration of 100 μg/mL, AcOEt (35.42 ± 1.00%) extract displayed the best activity. FRAP assay indicated that the BuOH extract (A0.50 = 17.42 ± 0.25 µg/mL) was more active than standard α-tocopherol (A0.50 = 34.93 ± 2.38 µg/mL). BuOH extract was more active than other extracts except in the ABTS●+, where the DCM extract was most active. This antioxidant activity could be attributed to the phenolic compounds detected. C. nepeta extracts showed moderate inhibition on acetylcholinesterase (AChE), butyrylcholinesterase (BChE), tyrosinase, and α-amylase. The results indicate that C. nepeta is a potent source of natural antioxidants that could be used in managing microbial resistance and Alzheimer′s disease.

Competitive advantage of oral streptococci for colonization of the middle ear mucosa

The intermittent aeration of the middle ear seeds its mucosa with saliva aerosols and selects for a distinct community of commensals adapted to the otic microenvironment. We gained insights into the selective forces that enrich for specific groups of oral migrants in the middle ear mucosa by investigating the phylogeny and physiology of 19 strains enriched (Streptococcus) or transiently present (Staphylococcus, Neisseria and actinobacterial Micrococcus and Corynebacterium) in otic secretions. Phylogenetic analyses of full length 16S rRNA sequences resolved close relationships between the streptococcal strains and oral commensals as well as between the transient migrants and known nasal and oral species. Physiological functions that facilitate mucosal colonization (swarming motility, surfactant production) and nutrition (mucin and protein degradation) were widespread in all the otic cultivars, as was the ability of most of the isolates to grow both aerobically and anaerobically. However, streptococci stood out for their enhanced biofilm-forming abilities under oxic and anoxic conditions and for their efficient fermentation of mucosal substrates into lactate, a key metabolic intermediate in the otic trophic webs. Additionally, the otic streptococci inhibited the growth of common otopathogens, an antagonistic interaction that could exclude competitors and protect the middle ear mucosa from infections by transient pathobionts. These adaptive traits allow streptococcal migrants to colonize the otic mucosa and grow microcolonies with syntrophic anaerobic partners, establishing trophic webs with other commensals similar to those formed by the oral ancestors in buccal biofilms.

In vitro antibiofilm activity of resveratrol against avian pathogenic Escherichia coli

Background Avian pathogenic Escherichia coli (APEC) strains cause infectious diseases in poultry. Resveratrol is extracted from Polygonum cuspidatum, Cassia tora Linn and Vitis vinifera, and displays good antimicrobial activity. The present study aimed to investigate the antibiofilm effect of resveratrol on APEC in vitro. The minimum inhibitory concentration (MIC) of resveratrol and the antibiotic florfenicol toward APEC were detected using the broth microdilution method. Then, the effect of resveratrol on swimming and swarming motility was investigated using a semisolid medium culture method. Subsequently, the minimum biofilm inhibitory concentration (MBIC) and the biofilm eradication rate were evaluated using crystal violet staining. Finally, the antibiofilm activity of resveratrol was observed using scanning electron microscopy (SEM). Meanwhile, the effects of florfenicol combined with resveratrol against biofilm formation by APEC were evaluated using optical microscopy (OM) and a confocal laser scanning microscopy (CLSM). Results The MICs of resveratrol and florfenicol toward APEC were 128 μg/mL and 64 μg/mL, respectively. The swimming and swarming motility abilities of APEC were inhibited in a resveratrol dose-dependent manner. Furthermore, resveratrol showed a significant inhibitory activity against APEC biofilm formation at concentrations above 1 μg/mL (p < 0.01). Meanwhile, the inhibitory effect of resveratrol at 32 μg/mL on biofilm formation was observed using SEM. The APEC biofilm was eradicated at 32 μg/mL of resveratrol combined with 64 μg/mL of florfenicol, which was observed using CLSM and OM. Florfenicol had a slight eradication effect of biofilm formation, whereas resveratrol had a strong biofilm eradication effect toward APEC. Conclusion Resveratrol displayed good antibiofilm activity against APEC in vitro, including inhibition of swimming and swarming motility, biofilm formation, and could eradicate the biofilm.