ResearchPad - bacteriology https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Confocal Raman microscopy to identify bacteria in oral subgingival biofilm models]]> https://www.researchpad.co/article/elastic_article_7675 The study of oral disease progression, in relation to the accumulation of subgingival biofilm in gingivitis and periodontitis is limited, due to either the ability to monitor plaque in vitro. When compared, optical spectroscopic techniques offer advantages over traditional destructive or biofilm staining approaches, making it a suitable alternative for the analysis and continued development of three-dimensional structures. In this work, we have developed a confocal Raman spectroscopy analysis approach towards in vitro subgingival plaque models. The main objective of this study was to develop a method for differentiating multiple oral subgingival bacterial species in planktonic and biofilm conditions, using confocal Raman microscopy. Five common subgingival bacteria (Fusobacterium nucleatum, Streptococcus mutans, Veillonella dispar, Actinomyces naeslundii and Prevotella nigrescens) were used and differentiated using a 2-way orthogonal Partial Least Square with Discriminant Analysis (O2PLS-DA) for the collected spectral data. In addition to planktonic growth, mono-species biofilms cultured using the ‘Zürich Model’ were also analyzed. The developed method was successfully used to predict planktonic and mono-species biofilm species in a cross validation setup. The results show differences in the presence and absence of chemical bands within the Raman spectra. The O2PLS-DA model was able to successfully predict 100% of all tested planktonic samples and 90% of all mono-species biofilm samples. Using this approach we have shown that Confocal Raman microscopy can analyse and predict the identity of planktonic and mono-species biofilm species, thus enabling its potential as a technique to map oral multi-species biofilm models.

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<![CDATA[Chitosan-propolis nanoparticle formulation demonstrates anti-bacterial activity against Enterococcus faecalis biofilms]]> https://www.researchpad.co/article/5989db50ab0ee8fa60bdc122

Propolis obtained from bee hives is a natural substance with antimicrobial properties. It is limited by its insolubility in aqueous solutions; hence ethanol and ethyl acetate extracts of Malaysian propolis were prepared. Both the extracts displayed antimicrobial and anti-biofilm properties against Enterococcus faecalis, a common bacterium associated with hospital-acquired infections. High performance liquid chromatography (HPLC) analysis of propolis revealed the presence of flavonoids like kaempferol and pinocembrin. This study investigated the role of propolis developed into nanoparticles with chitosan for its antimicrobial and anti-biofilm properties against E. faecalis. Bacteria that grow in a slimy layer of biofilm are resistant to penetration by antibacterial agents. The use of nanoparticles in medicine has received attention recently due to better bioavailability, enhanced penetrative capacity and improved efficacy. A chitosan-propolis nanoformulation was chosen based on ideal physicochemical properties such as particle size, zeta potential, polydispersity index, encapsulation efficiency and the rate of release of the active ingredients. This formulation inhibited E. faecalis biofilm formation and reduced the number of bacteria in the biofilm by ~90% at 200 μg/ml concentration. When tested on pre-formed biofilms, the formulation reduced bacterial number in the biofilm by ~40% and ~75% at 200 and 300 μg/ml, respectively. The formulation not only reduced bacterial numbers, but also physically disrupted the biofilm structure as observed by scanning electron microscopy. Treatment of biofilms with chitosan-propolis nanoparticles altered the expression of biofilm-associated genes in E. faecalis. The results of this study revealed that chitosan-propolis nanoformulation can be deemed as a potential anti-biofilm agent in resisting infections involving biofilm formation like chronic wounds and surgical site infections.

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<![CDATA[Comparing in vitro and in vivo virulence phenotypes of Burkholderia pseudomallei type G strains]]> https://www.researchpad.co/article/5989db52ab0ee8fa60bdc780

Burkholderia pseudomallei (Bpm) is a saprophytic rod-shaped gram-negative bacterium and the causative agent of melioidosis. This disease has previously been described as endemic in areas such as northern Australia and Southeast Asia, but, more recently, a better understanding of the epidemiology of melioidosis indicated that the disease is distributed worldwide, including regions of the Americas and Africa. A 16S-23S rDNA internal transcribed spacer (ITS) typing system has been developed for Bpm and has revealed that ITS types C, E, and hybrid CE are mainly associated with Australia and Southeast Asia while type G strains are more associated with cases of melioidosis in the Western Hemisphere. The purpose of the current study was to determine the in vitro and in vivo virulence profiles of the understudied Bpm type G strains Ca2009, Ca2013a, Mx2013, and 724644 and compared such phenotypes to the commonly studied Bpm type C strain K96243. We evaluated virulence by measuring invasion/uptake and survival of these Bpm strains in murine respiratory epithelial LA-4 cells and alveolar macrophage MH-S cells using different multiplicity of infections (MOIs of 1 and 10). We also calculated the lethal dose 50 values (LD50) in BALB/c mice that were inoculated intranasally with either Ca2009, Ca2013a, or Mx2013. Overall, the virulence and lethality phenotypes of Bpm type G strains were similar to the Bpm type C strain K96243. Additional comparative analyses between the Bpm ITS types may lead to a better understanding of the contribution of the ITS type to the epidemiology and ecology of Bpm strains.

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<![CDATA[Investigation of synovial fluid induced Staphylococcus aureus aggregate development and its impact on surface attachment and biofilm formation]]> https://www.researchpad.co/article/Nf13f73b5-5132-41b9-b894-3d4dd0a113b1

Periprosthetic joint infections (PJIs) are a devastating complication that occurs in 2% of patients following joint replacement. These infections are costly and difficult to treat, often requiring multiple corrective surgeries and prolonged antimicrobial treatments. The Gram-positive bacterium Staphylococcus aureus is one of the most common causes of PJIs, and it is often resistant to a number of commonly used antimicrobials. This tolerance can be partially attributed to the ability of S. aureus to form biofilms. Biofilms associated with the surface of indwelling medical devices have been observed on components removed during chronic infection, however, the development and localization of biofilms during PJIs remains unclear. Prior studies have demonstrated that synovial fluid, in the joint cavity, promotes the development of bacterial aggregates with many biofilm-like properties, including antibiotic resistance. We anticipate these aggregates have an important role in biofilm formation and antibiotic tolerance during PJIs. Therefore, we sought to determine specifically how synovial fluid promotes aggregate formation and the impact of this process on surface attachment. Using flow cytometry and microscopy, we quantified the aggregation of various clinical S. aureus strains following exposure to purified synovial fluid components. We determined that fibrinogen and fibronectin promoted bacterial aggregation, while cell free DNA, serum albumin, and hyaluronic acid had minimal effect. To determine how synovial fluid mediated aggregation affects surface attachment, we utilized microscopy to measure bacterial attachment. Surprisingly, we found that synovial fluid significantly impeded bacterial surface attachment to a variety of materials. We conclude from this study that fibrinogen and fibronectin in synovial fluid have a crucial role in promoting bacterial aggregation and inhibiting surface adhesion during PJI. Collectively, we propose that synovial fluid may have conflicting protective roles for the host by preventing adhesion to surfaces, but by promoting bacterial aggregation is also contributing to the development of antibiotic tolerance.

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<![CDATA[Toxin-neutralizing antibodies elicited by naturally acquired cutaneous anthrax are elevated following severe disease and appear to target conformational epitopes]]> https://www.researchpad.co/article/N0733fdcc-4c39-44e4-82cd-032e69d54dbc

Understanding immune responses to native antigens in response to natural infections can lead to improved approaches to vaccination. This study sought to characterize the humoral immune response to anthrax toxin components, capsule and spore antigens in individuals (n = 46) from the Kayseri and Malatya regions of Turkey who had recovered from mild or severe forms of cutaneous anthrax infection, compared to regional healthy controls (n = 20). IgG antibodies to each toxin component, the poly-γ-D-glutamic acid capsule, the Bacillus collagen-like protein of anthracis (BclA) spore antigen, and the spore carbohydrate anthrose, were detected in the cases, with anthrax toxin neutralization and responses to Protective Antigen (PA) and Lethal Factor (LF) being higher following severe forms of the disease. Significant correlative relationships among responses to PA, LF, Edema Factor (EF) and capsule were observed among the cases. Though some regional control sera exhibited binding to a subset of the tested antigens, these samples did not neutralize anthrax toxins and lacked correlative relationships among antigen binding specificities observed in the cases. Comparison of serum binding to overlapping decapeptides covering the entire length of PA, LF and EF proteins in 26 cases compared to 8 regional controls revealed that anthrax toxin-neutralizing antibody responses elicited following natural cutaneous anthrax infection are directed to conformational epitopes. These studies support the concept of vaccination approaches that preserve conformational epitopes.

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<![CDATA[Potential combinations of endocannabinoid/endocannabinoid-like compounds and antibiotics against methicillin-resistant Staphylococcus aureus]]> https://www.researchpad.co/article/Ne8a72c2e-13c7-43d3-9f49-0ed6410d9d0b

Infections caused by antibiotic-resistant strains of Staphylococcus aureus have reached epidemic proportions globally. Our previous study showed antimicrobial effects of anandamide (AEA) and arachidonoyl serine (AraS) against methicillin (MET)-resistant S. aureus (MRSA) strains, proposing the therapeutic potential of these endocannabinoid/endocannabinoid-like (EC/EC-like) agents for the treatment of MRSA. Here, we investigated the potential synergism of combinations of AEA and AraS with different types of antibiotics against MRSA grown under planktonic growth or biofilm formation. The most effective combinations under planktonic conditions were mixtures of AEA and ampicillin (AMP), and of AraS and gentamicin (GEN). The combination with the highest synergy in the biofilm formation against all tested bacterial strains was AEA and MET. Moreover, the combination of AraS and MET synergistically caused default of biofilm formation. Slime production of MRSA was also dramatically impaired by AEA or AraS combined with MET. Our data suggest the novel potential activity of combinations of EC/EC-like agents and antibiotics in the prevention of MRSA biofilm formation.

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<![CDATA[Towards understanding the antagonistic activity of phytic acid against common foodborne bacterial pathogens using a general linear model]]> https://www.researchpad.co/article/Nee28f4e6-a119-4233-a9a2-0c085b39343b

The increasing challenge of antibiotic resistance requires not only the discovery of new antibiotics, but also the development of new alternative approaches. Therefore, in the present study, we investigated for the first time the antibacterial potential of phytic acid (myo-inositol hexakisphosphate, IP6), a natural molecule that is ‘generally recognized as safe’ (FDA classification), against the proliferation of common foodborne bacterial pathogens such as Listeria monocytogenes, Staphylococcus aureus and Salmonella Typhimurium. Interestingly, compared to citric acid, IP6 was found to exhibit significantly greater inhibitory activity (P<0.05) against these pathogenic bacteria. The minimum inhibitory concentration of IP6 varied from 0.488 to 0.97 mg/ml for the Gram-positive bacteria that were tested, and was 0.244 mg/ml for the Gram-negative bacteria. Linear and general models were used to further explore the antibacterial effects of IP6. The developed models were validated using experimental growth data for L. monocytogenes, S. aureus and S. Typhimurium. Overall, the models were able to accurately predict the growth of L. monocytogenes, S. aureus, and S. Typhimuriumin Polymyxin acriflavine lithium chloride ceftazidime aesculin mannitol (PALCAM), Chapman broth, and xylose lysine xeoxycholate (XLD) broth, respectively. Remarkably, the early logarithmic growth phase of S. Typhimurium showed a rapid and severe decrease in a period of less than one hour, illustrating the bactericidal effect of IP6. These results suggest that IP6 is an efficient antibacterial agent and can be used to control the proliferation of foodborne pathogens. It has promising potential for environmentally friendly applications in the food industry, such as for food preservation, food safety, and for prolonging shelf life.

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<![CDATA[Generating Vesicular Stomatitis Virus Pseudotype Bearing the Severe Acute Respiratory Syndrome Coronavirus Spike Envelope Glycoprotein for Rapid and Safe Neutralization Test or Cell-Entry Assay]]> https://www.researchpad.co/article/N7ad6498c-cfd9-496d-b21e-30e11fcbdc29

abstract:  We generated a recombinant vesicular stomatitis virus (VSV) pseudotype (VSV Δ G*SG) by replacing the envelope G gene with the GFP gene and complementing with spike glycoprotein (S) of SARS‐CoV in trans. The neutralization and infection blocking tests showed that the VSV Δ G*SG and SARS‐CoV reacted similarly to SARS‐CoV specific antiserum, suggesting the VSVΔ G*SG can be a safe replacement of the live SARS‐CoV for neutralization test and cell‐entry assay.

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<![CDATA[Optimizing DNA Extraction Methods for Nanopore Sequencing of Neisseria gonorrhoeae Directly from Urine Samples]]> https://www.researchpad.co/article/N8364d333-de5f-42de-96e3-fdf514dca0d7

Empirical gonorrhea treatment at initial diagnosis reduces onward transmission. However, increasing resistance to multiple antibiotics may necessitate waiting for culture-based diagnostics to select an effective treatment. There is a need for same-day culture-free diagnostics that identify infection and detect antimicrobial resistance. We investigated if Nanopore sequencing can detect sufficient Neisseria gonorrhoeae DNA to reconstruct whole genomes directly from urine samples.

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<![CDATA[Evaluation of liposomal ciprofloxacin formulations in a murine model of anthrax]]> https://www.researchpad.co/article/Ne17111d7-5152-4c88-81b1-0e84a1b58e42

The in vivo efficacy of liposomal encapsulated ciprofloxacin in two formulations, lipoquin and apulmiq, were evaluated against the causative agent of anthrax, Bacillus anthracis. Liposomal encapsulated ciprofloxacin is attractive as a therapy since it allows for once daily dosing and achieves higher concentrations of the antibiotic at the site of initial mucosal entry but lower systemic drug concentrations. The in vivo efficacy of lipoquin and apulmiq delivered by intranasal instillation was studied at different doses and schedules in both a post exposure prophylaxis (PEP) therapy model and in a delayed treatment model of murine inhalational anthrax. In the mouse model of infection, the survival curves for all treatment cohorts differed significantly from the vehicle control. Ciprofloxacin, lipoquin and apulmiq provided a high level of protection (87–90%) after 7 days of therapy when administered within 24 hours of exposure. Reducing therapy to only three days still provided protection of 60–87%, if therapy was provided within 24 hours of exposure. If treatment was initiated 48 hours after exposure the survival rate was reduced to 46–65%. These studies suggest that lipoquin and apulmiq may be attractive therapies as PEP and as part of a treatment cocktail for B. anthracis.

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<![CDATA[Reconstruction of Dispersal Patterns of Hypervirulent Meningococcal Strains of Serogroup C:cc11 by Phylogenomic Time Trees]]> https://www.researchpad.co/article/N4bc71ea8-df5a-4b07-bcf4-07ef5adfda55

Neisseria meningitidis is one of the few commensal bacteria that can even cause large epidemics of invasive meningococcal disease (IMD). N. meningitis serogroup C belonging to the hypervirulent clonal complex 11 (cc11) represents an important public health threat worldwide. We reconstructed the dispersal patterns of hypervirulent meningococcal strains of serogroup C:cc11 by phylogenomic time trees.

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<![CDATA[Evaluation of WASPLab Software To Automatically Read chromID CPS Elite Agar for Reporting of Urine Cultures]]> https://www.researchpad.co/article/N298d1a35-5b49-4390-8a6e-f5ec3422562e

Urine cultures are among the most common specimens received by clinical laboratories and generate a major share of the laboratory workload. Chromogenic agar can expedite culture results, but technologist review is still needed. In this study, we evaluated the ability of the WASPLab software to interpret urine specimens plated onto chromID CPS Elite (CPSE) agar. Urine specimens submitted for bacterial culture were plated onto CPSE agar with a 1-μl loop using the WASP.

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<![CDATA[Hash-Based Core Genome Multilocus Sequence Typing for Clostridium difficile]]> https://www.researchpad.co/article/Nfd09c02b-d7d1-4e63-b309-b8c343ea14f6

Pathogen whole-genome sequencing has huge potential as a tool to better understand infection transmission. However, rapidly identifying closely related genomes among a background of thousands of other genomes is challenging. Here, we describe a refinement to core genome multilocus sequence typing (cgMLST) in which alleles at each gene are reproducibly converted to a unique hash, or short string of letters (hash-cgMLST).

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<![CDATA[Kestose supplementation exerts bifidogenic effect within fecal microbiota and increases fecal butyrate concentration in dogs]]> https://www.researchpad.co/article/Nba33ae03-7b68-4646-b56a-aa630e828338

Kestose, a fructooligosaccharide (FOS) with one fructose monomer linked to sucrose, is a key component of the prebiotic activity of FOS. This study aimed to evaluate the prebiotic potential of Kestose in terms of the impact on population change in the intestinal microbiota and fecal short-chain fatty acid (SCFA) concentration in dogs. Kestose 2 g per dog was administered daily with conventional diet to 6 healthy, adult beagle dogs for 8 weeks followed by 4 weeks of follow-up period without Kestose supplementation. Fresh fecal samples were obtained before and every 4 weeks until the end of the follow-up period. Genomic DNA extracted from the fecal samples was subjected to 16S rRNA gene analysis using next generation sequencer and to quantitative polymerase chain reaction (qPCR). Fecal acetate, propionate, butyrate, lactate and ethanol concentrations were measured by high-performance liquid chromatography. 16S rRNA gene analysis and qPCR showed increasing trend of genus Bifidobacterium after Kestose supplementation while genera Bacteroides and Sutterella decreased. Clostridium perfringens decreased below the detection limit within first 4 weeks after starting Kestose supplementation. Fecal butyrate concentration was significantly increased at week 8 and returned to the base level after 4 weeks of the washing period. To the best of our knowledge, this is the first study to reveal effect of Kestose on the populational changes in fecal microbiota and fecal butyrate concentration in dogs.

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<![CDATA[Changes in antimicrobial resistance phenotypes and genotypes in Streptococcus suis strains isolated from pigs in the Tokai area of Japan]]> https://www.researchpad.co/article/N0ffba502-a722-4941-89ad-e8bb0fa9c58d

Streptococcus suis strains isolated from porcine endocarditis and tonsils in the Tokai area of Japan during 2004–2007 and 2014–2016 (n=114) were tested for antimicrobial susceptibility and distribution of selected resistance genes. No strains showed resistance to penicillin, ampicillin, cefotaxime, meropenem, vancomycin, and levofloxacin. High resistance to tetracycline (80.7%), clindamycin (65.8%), erythromycin (56.1%), and clarithromycin (56.1%) was observed. In chloramphenicol and sulfamethoxazole-trimethoprim, there was a trend towards increased resistance between the first (2004–2007) and second (2014–2016) periods. tet(O) and erm(B) genes were the most frequently detected, and tet(M) and mef(A/E) genes were only detected in strains isolated during 2014–2016. These results indicate that chloramphenicol and sulfamethoxazole-trimethoprim resistance, and tet(M) and mef(A/E) genes emerged in S. suis of this area after 2014.

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<![CDATA[Evolutionary behaviour of bacterial prion-like proteins]]> https://www.researchpad.co/article/5c8823f7d5eed0c484639437

Prions in eukaryotes have been linked to diseases, evolutionary capacitance, large-scale genetic control and long-term memory formation. In bacteria, constructed prion-forming proteins have been described, such as the prion-forming protein recently described for Clostridium botulinum transcription terminator Rho. Here, I analyzed the evolution of the Rho prion-forming domain across bacteria, and discovered that its conservation is sporadic both in the Clostridium genus and in bacteria generally. Nonetheless, it has an apparent evolutionary reach into eight or more different bacterial phyla. Motivated by these results, I investigated whether this pattern of wide-ranging evolutionary sporadicity is typical of bacterial prion-like domains. A measure of coverage of a domain (C) within its evolutionary range was derived, which is effectively a weighted fraction of the number of species in which the domain is found. I observe that occurrence across multiple phyla is not uncommon for bacterial prion-like protein domain families, but that they tend to sample of a low fraction of species within their evolutionary range, like Rho. The Rho prion-like domain family is one of the top three most widely distributed prion-like protein domain families in terms of number of phyla. There are >60 prion-like protein domain families that have at least the evolutionary coverage of Rho, and are found in multiple phyla. The implications of these findings for evolution and for experimental investigations into prion-forming proteins are discussed.

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<![CDATA[Contact with adult hen affects development of caecal microbiota in newly hatched chicks]]> https://www.researchpad.co/article/5c8977aad5eed0c4847d32a0

Chickens in commercial production are hatched in a clean hatchery environment in the absence of any contact with adult hens. However, Gallus gallus evolved to be hatched in a nest in contact with an adult hen which may act as a donor of gut microbiota. In this study, we therefore addressed the issue of microbiota development in newly hatched chickens with or without contact with an adult hen. We found that a mere 24-hour-long contact between a hen and newly hatched chickens was long enough for transfer of hen gut microbiota to chickens. Hens were efficient donors of Bacteroidetes and Actinobacteria. However, except for genus Faecalibacterium and bacterial species belonging to class Negativicutes, hens did not act as an important source of Gram-positive Firmicutes. Though common to the chicken intestinal tract, Lactobacilli and isolates from families Erysipelotrichaceae, Lachnospiraceae and Ruminococcaceae therefore originated from environmental sources instead of from the hens. These observation may have considerable consequences for the evidence-based design of the new generation of probiotics for poultry.

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<![CDATA[Cationic chitosan-propolis nanoparticles alter the zeta potential of S. epidermidis, inhibit biofilm formation by modulating gene expression and exhibit synergism with antibiotics]]> https://www.researchpad.co/article/5c818e84d5eed0c484cc2423

Staphylococcus epidermidis, is a common microflora of human body that can cause opportunistic infections associated with indwelling devices. It is resistant to multiple antibiotics necessitating the need for naturally occurring antibacterial agents. Malaysian propolis, a natural product obtained from beehives exhibits antimicrobial and antibiofilm properties. Chitosan-propolis nanoparticles (CPNP) were prepared using Malaysian propolis and tested for their effect against S. epidermidis. The cationic nanoparticles depicted a zeta potential of +40 and increased the net electric charge (zeta potential) of S. epidermidis from -17 to -11 mV in a concentration-dependent manner whereas, ethanol (Eth) and ethyl acetate (EA) extracts of propolis further decreased the zeta potential from -17 to -20 mV. Confocal laser scanning microscopy (CLSM) depicted that CPNP effectively disrupted biofilm formation by S. epidermidis and decreased viability to ~25% compared to Eth and EA with viability of ~60–70%. CPNP was more effective in reducing the viability of both planktonic as well as biofilm bacteria compared to Eth and EA. At 100 μg/mL concentration, CPNP decreased the survival of biofilm bacteria by ~70% compared to Eth or EA extracts which decreased viability by only 40%-50%. The morphology of bacterial biofilm examined by scanning electron microscopy depicted partial disruption of biofilm by Eth and EA extracts and significant disruption by CPNP reducing bacterial number in the biofilm by ~90%. Real time quantitative PCR analysis of gene expression in treated bacteria showed that genes involved in intercellular adhesion such as IcaABCD, embp and other related genes were significantly downregulated by CPNP. In addition to having a direct inhibitory effect on the survival of S. epidermidis, CPNP showed synergism with the antibiotics rifampicin, ciprofloxacin, vancomycin and doxycycline suggestive of effective treatment regimens. This would help decrease antibiotic treatment dose by at least 4-fold in combination therapies thereby opening up ways of tackling antibiotic resistance in bacteria.

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<![CDATA[TyrR is involved in the transcriptional regulation of biofilm formation and D-alanine catabolism in Azospirillum brasilense Sp7.]]> https://www.researchpad.co/article/5c6f1503d5eed0c48467ac9f

Azospirillum brasilense is one of the most studied species of diverse agronomic plants worldwide. The benefits conferred to plants inoculated with Azospirillum have been primarily attributed to its capacity to fix atmospheric nitrogen and synthesize phytohormones, especially indole-3-acetic acid (IAA). The principal pathway for IAA synthesis involves the intermediate metabolite indole pyruvic acid. Successful colonization of plants by Azospirillum species is fundamental to the ability of these bacteria to promote the beneficial effects observed in plants. Biofilm formation is an essential step in this process and involves interactions with the host plant. In this study, the tyrR gene was cloned, and the translated product was observed to exhibit homology to TyrR protein, a NtrC/NifA-type activator. Structural studies of TyrR identified three putative domains, including a domain containing binding sites for aromatic amino acids in the N-terminus, a central AAA+ ATPase domain, and a helix-turn-helix DNA binding motif domain in the C-terminus, which binds DNA sequences in promoter-operator regions. In addition, a bioinformatic analysis of promoter sequences in A. brasilense Sp7 genome revealed that putative promoters encompass one to three TyrR boxes in genes predicted to be regulated by TyrR. To gain insight into the phenotypes regulated by TyrR, a tyrR-deficient strain derived from A. brasilense Sp7, named A. brasilense 2116 and a complemented 2116 strain harboring a plasmid carrying the tyrR gene were constructed. The observed phenotypes indicated that the putative transcriptional regulator TyrR is involved in biofilm production and is responsible for regulating the utilization of D-alanine as carbon source. In addition, TyrR was observed to be absolutely required for transcriptional regulation of the gene dadA encoding a D-amino acid dehydrogenase. The data suggested that TyrR may play a major role in the regulation of genes encoding a glucosyl transferase, essential signaling proteins, and amino acids transporters.

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<![CDATA[Analysis of Pseudomonas aeruginosa biofilm membrane vesicles supports multiple mechanisms of biogenesis]]> https://www.researchpad.co/article/5c6f151dd5eed0c48467ade7

Outer Membrane Vesicles (OMVs) are ubiquitous in bacterial environments and enable interactions within and between species. OMVs are observed in lab-grown and environmental biofilms, but our understanding of their function comes primarily from planktonic studies. Planktonic OMVs assist in toxin delivery, cell-cell communication, horizontal gene transfer, small RNA trafficking, and immune system evasion. Previous studies reported differences in size and proteomic cargo between planktonic and agar plate biofilm OMVs, suggesting possible differences in function between OMV types. In Pseudomonas aeruginosa interstitial biofilms, extracellular vesicles were reported to arise through cell lysis, in contrast to planktonic OMV biogenesis that involves the Pseudomonas Quinolone Signal (PQS) without appreciable autolysis. Differences in biogenesis mechanism could provide a rationale for observed differences in OMV characteristics between systems. Using nanoparticle tracking, we found that P. aeruginosa PAO1 planktonic and biofilm OMVs had similar characteristics. However, P. aeruginosa PA14 OMVs were smaller, with planktonic OMVs also being smaller than their biofilm counterparts. Large differences in Staphylococcus killing ability were measured between OMVs from different strains, and a smaller within-strain difference was recorded between PA14 planktonic and biofilm OMVs. Across all conditions, the predatory ability of OMVs negatively correlated with their size. To address biogenesis mechanism, we analyzed vesicles from wild type and pqsA mutant biofilms. This showed that PQS is required for physiological-scale production of biofilm OMVs, and time-course analysis confirmed that PQS production precedes OMV production as it does in planktonic cultures. However, a small sub-population of vesicles was detected in pqsA mutant biofilms whose size distribution more resembled sonicated cell debris than wild type OMVs. These results support the idea that, while a small and unique population of vesicles in P. aeruginosa biofilms may result from cell lysis, the PQS-induced mechanism is required to generate the majority of OMVs produced by wild type communities.

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