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Pseudomonas aeruginosa is one of the most common pathogens encountered in clinical wound infections. Clinical studies have shown that P. aeruginosa infection results in a larger wound area, inhibiting healing, and a high prevalence of antimicrobial resistance. Hydroxypyridinone-derived iron chelator Deferiprone (Def) and heme analogue Gallium-Protoporphyrin (GaPP) in a chitosan-dextran hydrogel (Chitogel) have previously been demonstrated to be effective against PAO1 and clinical isolates of P. aeruginosa in vitro. Moreover, this combination of these two agents has been shown to improve sinus surgery outcomes by quickly reducing bleeding and preventing adhesions. In this study, the efficacy of Def-GaPP Chitogel was investigated in a P. aeruginosa biofilm-infected wound murine model over 6 days. Two concentrations of Def-GaPP Chitogel were investigated: Def-GaPP high dose (10 mM Def + 500 µg/mL GaPP) and Def-GaPP low dose (5 mM Def + 200 µg/mL GaPP). The high-dose Def-GaPP treatment reduced bacterial burden in vivo from day 2, without delaying wound closure. Additionally, Def-GaPP treatment decreased wound inflammation, as demonstrated by reduced neutrophil infiltration and increased anti-inflammatory M2 macrophage presence within the wound bed to drive wound healing progression. Def-GaPP Chitogel treatment shows promising potential in reducing P. aeruginosa cutaneous infection with positive effects observed in the progression of wound healing. [ABSTRACT FROM AUTHOR]

One of the most causative agents for many opportunistic diseases is the Pseudomonas aeruginosa which has a high percentage of multidrug resistance disease through construction of biofilm. The current study aimed for evaluating the correlation between quorum sensing genes (which is lasI gene) and biofilm formation. The biofilm construction and antibiotics susceptibility test were achieved for all the isolates under the study. The PCR and sequencing techniques were also carried out to detect the type of variation in lasI gene for each scheme of biofilm formation (weak, strong, and moderate). High antibiotic resistance was recorded among biofilm producing isolates. The genic pattern for the weak biofilm producer, which is isolate No 19, carries lysine instead of arginine in position 92 and aspartic acid instead of asparagine in position 115. Phylogenetic analysis indicated that the horizontal genetic transfer between isolates was very high. Finally, we conclude that the lasI gene does participate in biofilm construction of P. aeruginosa. [ABSTRACT FROM AUTHOR]

Chronic pulmonary infection caused by Pseudomonas aeruginosa (P. aeruginosa) is a common lung disease with high mortality, posing severe threats to public health. Highly resistant biofilm and intrinsic resistance make P. aeruginosa hard to eradicate, while powerful virulence system of P. aeruginosa may give rise to the recurrence of infection and eventual failure of antibiotic therapy. To address these issues, infection-microenvironment responsive nanoparticles functioning on biofilm eradication and virulence inhibition were simply prepared by electrostatic complexation between dimethylmaleic anhydride (DA) modified negatively charged coating and epsilon-poly(l -lysine) derived cationic nanoparticles loaded with azithromycin (AZI) (DA-AZI NPs). Charge reversal responsive to acidic condition enabled DA-AZI NPs to successively penetrate through both mucus and biofilms, followed by targeting to P. aeruginosa and permeabilizing its outer/inner membrane. Then in situ released AZI, which was induced by the lipase-triggered NPs dissociation, could easily enter into bacteria to take effects. DA-AZI NPs exhibited enhanced eradication activity against P. aeruginosa biofilms with a decrease of >99.999% of bacterial colonies, as well as remarkable inhibitory effects on the production of virulence factors and bacteria re-adhesion & biofilm re-formation. In a chronic pulmonary infection model, nebulization of DA-AZI NPs into infected mice resulted in prolonged retention and increased accumulation of the NPs in the infected sites of the lungs. Moreover, they significantly reduced the burden of P. aeruginosa , effectively alleviating lung tissue damages and inflammation. Overall, the proposed DA-AZI NPs highlight an innovative strategy for treating chronic pulmonary infection. [Display omitted] [ABSTRACT FROM AUTHOR]

Multidrug-resistance (MDR) strains of Pseudomonas aeruginosa has resulted in the medical therapy complicated. Antimicrobial resistance and biofilm production are virulence factors responsible for the persistence of prolonged infections. In this study, antimicrobial susceptibility pattern and phenotypic characteristics of biofilm of P. aeruginosa were investigated. The aim and objective of this study is to evaluate the antibacterial susceptibility testing and the presence of Biofilm Formation Potential among clinical isolates of Pseudomonas aeruginosa at a Tertiary care centre. This was a cross sectional study carried out in the Department of Microbiology, Rama Medical College Hospital and Research Centre, Mandhana, Kanpur for a period of 1 year i.e, between March 2022 to March 2023. A Total of 355 clinical isolates from different clinical samples including urine, sputum, blood, Et secreation, pus, throat swab were identified by Standard microbiological techniques and the isolates were further tested for the detection of biofilm along with the MBL by Imipenem-EDTA combined disc test and MBL Etest (Imipenem) according to the CLSI guidelines 2021. A total of 355 clinical samples were studied out of which 132 P. aeruginosa isolates were obtained with the prevalence rate of 37.1% with the males 75(56.8%) being affected the most as compared to the females 57 (43.1%). The age group of 21-30 years of age followed by 31-40 was affected the most. In the age group of 0-10 years and above 71 years was the least affected with the Pseudomonas aeruginosa infection. It was also observed that the maximum number of isolates observed were from the Urine sample and least was from the blood and throat swab. The Sensitivities of Colistin was (96.9%), Piperacillin tazobactam (73.4%), Amikacin (77.2 %) and cefepime (73.4%) were found to be the most effective Antibiotics. The resistance to ciprofloxacin was (42.5%), Levofloxacin (50%), Gentamicin(63.6%), Imipenem (63.6%), Tobramycin( 69.6%), Ceftazidime (69.6%). Of which 20(15.1%%) were MBL positive by Imipenem-EDTA combined disc test, and 19(20.45%) by MBL E- test. It was observed that 50.75% of the Pseudomonas aeruginosa isolates were Biofilm producers with 1.5% strong and 3.7% were moderate biofilm producers. In conclusion, the present study showed combination therapy including an anti-pseudomonal beta-lactamand an aminoglycoside along with anti-biofilm agents can also be selected to eradicate biofilm-associated Pseudomonal infections, with strict infection control practices. MBL E-Test was found to be the best phenotypic method for detection of MBL is recommended into the daily routine of clinical laboratory. [ABSTRACT FROM AUTHOR]

The use of non‐antibiotic strategies to combat refractory drug‐resistant bacterial infections, especially biofilms and accompanying inflammation, has recently aroused widespread interest. Herein, a photo‐therapeutic nanocomposite with bio‐responsive oxygen (O2) self‐supplying is introduced by integrating manganese dioxide (MnO2) nanozymes onto photosensitizer (indocyanine green, ICG)‐loaded mesoporous polydopamine nanoparticles (MPDA), namely MI‐MPDA NPs. MI‐MPDA can activate O2 generation in the infection microenvironment, thereby effectively alleviating biofilm hypoxia. Under near‐infrared light (NIR) irradiation, continuous O2 supplying further boosts the level of singlet oxygen (1O2), enabling robust biofilm elimination through O2‐potentiated photodynamic/photothermal therapy. Interestingly, MI‐MPDA down‐regulates the factor expression of inflammatory signaling pathways through MnO2‐mediated reactive oxygen species scavenging, which ameliorates the inflammatory condition. Meanwhile, O2 supplying prevents the M1‐phenotype switch of macrophages from the overexpression of hypoxia‐inducible factor‐1α (HIF‐1α), thereby prompting macrophage reprogramming toward pro‐regenerative M2‐phenotype. In the mouse models of subcutaneous implant‐associated infection caused by methicillin‐resistant Staphylococcus aureus (MRSA) biofilms and burn infection caused by Pseudomonas aeruginosa biofilms, NIR‐irradiated MI‐MPDA not only effectively eliminates the formed biofilms, but also alleviates the oxidative stress and accompanying inflammation, and drives the cascade reaction of immunomodulation‐wound healing. Overall, this O2‐potentiated photo‐therapeutic strategy provides a reliable tool for combating biofilm infections and inflammation from drug‐resistant bacteria. [ABSTRACT FROM AUTHOR]

Biofilms and their associated infections have posed serious threats to the society from both an economical concern and a healthcare perspective. Graphene oxide (GO) has recently prompted the emergence of applications in the area of antibacterial materials owing to its high biocompatibility and surface functionalization. However, the weak biological interactions with microbial cells limit the antibiotic activity of GO. Herein, by combining GO with currently utilized antibacterial metal ions, copper ion‐graphene oxide composite (Cu2+@GO) with exceptional bactericidal activity is achieved in this work. Cu2+@GO composite possessed more than 95% antimicrobial efficiency of both Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa) compared with the Cu2+ only group or GO only group. Additionally, the composite inhibits the formation of bacterial biofilms, making it a potential candidate for the development of alternative antimicrobials. Antibacterial mechanism study shows that Cu2+@GO has both bacteria‐binding ability and the property of producing reactive oxygen species (ROS) to cause the bacterial destruction. In vivo experiments indicate that Cu2+@GO possess an inhibition effect to P. aeruginosa‐induced keratitis. Overall, this work demonstrates that Cu2+@GO may be promising antibacterial agents for the treatment of bacterial infection, which contributes to designing more antibiotic and safe graphene‐based materials. [ABSTRACT FROM AUTHOR]

Aimen Fatima,1 Bushra Aslam,1 Maryam Zaid,1 Muska Khattak,1 Sidra Bashir,1 Safia Masood,1 Zara Rafaque,2 Javid Iqbal Dasti11Lab of Microbial Genomics and Epidemiology, Department of Microbiology, Quaid-I-Azam University, Islamabad, 45320, Pakistan; 2Department of Microbiology Hazara University Mansehra, Mansehra 21120, PakistanCorrespondence: Javid Iqbal Dasti, Lab of Microbial Genomics and Epidemiology, Department of Microbiology, Quaid-I-Azam University, Islamabad, 45320, Pakistan, Tel +92-5190644175, Email [email protected] Background:Pseudomonas aeruginosa is an opportunistic pathogen involved in number of hospital-acquired infections such as catheter-associated urinary tract infections, bacteremia, septicemia, skin infections, and ventilator-associated pneumoniae. Biofilm formation is an important trait implicated in chronic infections, such as cystic fibrosis and chronic pulmonary obstruction. We evaluated effects of gentamicin, cefepime, and ciprofloxacin on biofilm of P. aeruginosa.Materials and Methods: A total of 266 isolates were collected from the Armed Forces Institute of Pathology (AFIP). Antibiotic susceptibility was assessed by double disk synergy testing. ESBL and carbapenemase detection was performed by phenotypic testing. Molecular screening of the genes was done by PCR. Micro-dilution broth method was used to determine minimum inhibitory concentrations of antibiotics. Biofilm formation was done by micro-titer plate assay.Results: Overall, 20% of the P. aeruginosa isolates were extensively drug-resistant (XDR-PA), and 25% were multi-drug-resistant (MDR-PA). Likewise, 43% of the isolates were ESBL producers, and carbapenemase production was detected in 40% of the isolates. Molecular analysis confirmed occurrence of different resistant factors in ESBL-positive isolates; 67% carried blaTEM, 62% blaCTXM-15, 41% blaSHV, 34% blaCTXM-14, and 33% blaOXA-1. In addition, 68% of the carbapenem-resistant isolates were positive for blaNDM-1, 25% for blaOXA-48, and 22% for blaKPC-2. Biofilm formation was assessed for 234 isolates, out of which 28% were strong biofilm formers. Moderate and weak biofilm formers constituted 46% and 23%, respectively. Overall, ciprofloxacin, levofloxacin, and cefepime showed inhibitory effects on P. aeruginosa biofilms. Antibiotics in combination showed strong synergistic effects (ciprofloxacin and cefepime), while gentamicin and cefepime resulted in complete eradication of P. aeruginosa biofilm.Conclusion: We confirm strong synergistic effects of gentamicin and cefepime that completely eradicated P. aeruginosa biofilm. We further confirm inhibitory effects of ciprofloxacin, levofloxacin, and cefepime on P. aeruginosa biofilms. Hence, combination therapy can be more effective against biofilm-associated infections. [ABSTRACT FROM AUTHOR]

Bacterial biofilms generally contribute to chronic infections, including wound infections. Due to the antibiotic resistance mechanisms protecting bacteria living in the biofilm, they are a serious problem in the wound healing process. To accelerate the wound healing process and avoid bacterial infection, it is necessary to select the appropriate dressing material. In this study, the promising therapeutic properties of alginate lyase (AlgL) immobilised on BC membranes for protecting wounds from Pseudomonas aeruginosa infection were investigated. The AlgL was immobilised on never dried BC pellicles via physical adsorption. The maximum adsorption capacity of AlgL was 6.0 mg/g of dry BC, and the equilibrium was reached after 2 h. The adsorption kinetics was studied, and it has been proven that the adsorption was consistent with Langmuir isotherm. In addition, the impact of enzyme immobilisation on bacterial biofilm stability and the effect of simultaneous immobilisation of AlgL and gentamicin on the viability of bacterial cells was investigated. The obtained results showed that the AlgL immobilisation significantly reduced the amount of polysaccharides component of the P. aeruginosa biofilm. Moreover, the biofilm disruption by AlgL immobilised on BC membranes exhibited synergism with the gentamicin, resulting in 86.5% more dead P. aeruginosa PAO-1 cells. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium implicated in acute and chronic nosocomial infections and a leading cause of patient mortality. Pseudomonas aeruginosa infections are frequently associated with the development of biofilms, which give the bacteria additional drug resistance and increase their virulence. The goal of this study was to find strong compounds that block the Anthranilate-CoA ligase enzyme made by the pqsA gene. This would stop the P. aeruginosa quorum signaling system. This enzyme plays a crucial role in the pathogenicity of P. aeruginosa by producing autoinducers for cell-to-cell communication that lead to the production of biofilms. Pharmacophore-based virtual screening was carried out utilizing a library of commercially accessible enzyme inhibitors. The most promising hits obtained during virtual screening were put through molecular docking with the help of MOE. The virtual screening yielded 7/160 and 10/249 hits (ZINC and Chembridge). Finally, 2/7 ZINC hits and 2/10 ChemBridge hits were selected as potent lead compounds employing diverse scaffolds due to their high pqsA enzyme binding affinity. The results of the pharmacophore-based virtual screening were subsequently verified using a molecular dynamic simulation-based study (MDS). Using MDS and post-MDS, the stability of the complexes was evaluated. The most promising lead compounds exhibited a high binding affinity towards protein-binding pocket and interacted with the catalytic dyad. At least one of the scaffolds selected will possibly prove useful for future research. However, further scientific confirmation in the form of preclinical and clinical research is required before implementation. [ABSTRACT FROM AUTHOR]

Background and Aim: Multidrug-resistant (MDR) Pseudomonas aeruginosa is a global threat to public health. This study aimed to determine biofilms and efflux pump regulatory gene (mexR) in MDR P. aeruginosa isolates. Materials and Methods: A total of 42 fecal samples of aquatic migratory birds collected during hunting season in Egypt were evaluated for the detection of P. aeruginosa according to standard culture-based methods. The antibiotic susceptibility of P. aeruginosa strains was evaluated using disk diffusion methods. The biofilm formation ability of the isolates was phenotypically determined using a colorimetric microtitration plate assay. Polymerase chain reaction amplification was performed to detect biofilm genes (PelA and PslA) and mexR. Results: In total, 19 isolates (45.2%) were recovered from the 42 fecal samples of migratory birds. All isolates were identified as MDR P. aeruginosa, and 78.9% of the strains produced biofilms at different degrees. Molecular detection of biofilm extracellular polymeric substances revealed that PelA was the most predominant gene in the biofilm-producing isolates, followed by PslA. mexR was detected in 63.2% of MDR P. aeruginosa isolates, and its prevalence was higher in non-biofilm-producing strains (75%) than in biofilm-producing strains (60%). Conclusion: Antibiotic resistance in P. aeruginosa isolates recovered from migratory birds through various mechanisms is a major public and animal health problem. It is important to consider the significance of migratory birds in disease transmission. [ABSTRACT FROM AUTHOR]

The increasing risk of antibiotic failure in the treatment of Pseudomonas aeruginosa infections is largely related to the production of a wide range of virulence factors. The use of non-thermal plasma (NTP) is a promising alternative to antimicrobial treatment. Nevertheless, there is still a lack of knowledge about the effects of NTP on the virulence factors production. We evaluated the ability of four NTP-affected P. aeruginosa strains to re-form biofilm and produce Las-B elastase, proteases, lipases, haemolysins, gelatinase or pyocyanin. Highly strains-dependent inhibitory activity of NTP against extracellular virulence factors production was observed. Las-B elastase activity was reduced up to 82% after 15-min NTP treatment, protease activity and pyocyanin production by biofilm cells was completely inhibited after 60 min, in contrast to lipases and gelatinase production, which remained unchanged. However, for all strains tested, a notable reduction in biofilm redevelopment ability was depicted using spinning disc confocal microscopy. In addition, NTP exposure of mature biofilms caused disruption of biofilm cells and their dispersion into the environment, as shown by transmission electron microscopy. This appears to be a key step that could help overcome the high resistance of P. aeruginosa and its eventual elimination, for example in combination with antibiotics still highly effective against planktonic cells. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa contributes to many chronic infections and has been found to be resistant to multiple antibiotics. Pseudomonas use a quorum sensing system (QS) to control biofilm establishment and virulence factors, and, thus, quorum sensing inhibitors (QSIs), such as meta-bromo-thiolactone (mBTL), are promising anti-infective agents. Accordingly, this study intended to investigate the antibacterial and anti-virulence activity of mBTL-loaded calcium alginate nanoparticles (CANPs) against Pseudomonas aeruginosa and different QS mutants. The results show that the mBTL-CANPs had higher antibacterial activity, which was made evident by decreases in all tested strains except the ∆lasR/∆rhlR double mutant, with MIC50 (0.5 mg/mL) of mBTL-CANPs compared with free mBTL at MIC50 (˃1 mg/mL). The biofilm formation of P. aeruginosa and some QS-deficient mutants were reduced in response to 0.5–0.125 mg/mL of mBTL-encapsulating CANPs. The pyocyanin production of the tested strains except ∆lasA and ∆rhlR decreased when challenged with 0.5 mg/mL of mBTL-loaded NPs. The subsequent characterization of the cytotoxic effect of these NPs on human lung epithelial cells (A549) and cystic fibrosis fibroblast cells (LL 29) demonstrated that synthesized NPs were cytocompatible at MIC50 in both cell lines and markedly reduced the cytotoxic effect observed with mBTL alone on these cells. The resulting formulation reduced the P. aeruginosa strains' adhesion to A549 comparably with mBTL, suggesting their potential anti-adhesive effect. Given the virulence suppressing action, cytocompatibility, and enhanced anti-biofilm effect of mBTL-CANPs, and the advantage of alginate-based NPs as an antimicrobial delivery system these nanoparticles have great potential in the prophylaxis and treatment of infection caused by Pseudomonas aeruginosa. [ABSTRACT FROM AUTHOR]

Biofilm infections are tolerant to the host responses and recalcitrance to antibiotic drugs and disinfectants. The induced host-specific innate and adaptive immune responses by established biofilms are significantly implicated and contributes to the course of the infections. Essentially, the host response may be the single one factor impacting the outcome most, especially in cases where the biofilm is caused by low virulent opportunistic bacterial species. Due to the chronicity of biofilm infections, activation of the adaptive immune response mechanisms is frequently experienced, and instead of clearing the infection, the adaptive response adds to the pathogenesis. To a high degree, this has been reported for chronic Pseudomonas aeruginosa lung infections, where both a pronounced antibody response and a skewed Th1/Th2 balance has been related to a poorer outcome. In addition, detection of an adaptive immune response can be used as a significant indicator of a chronic P. aeruginosa lung infection and is included in the clinical definitions as such. Those issues are presented in the present review, along with a characterization of the airway structure in relation to immune responses towards P. aeruginosa pulmonary infections. [ABSTRACT FROM AUTHOR]

The physiology of an organism in the environment reflects its interactions with the diverse physical, chemical, and biological properties of the surface. These principles come into consideration during model selection to study biofilm–host interactions. Biofilms are communities formed by beneficial and pathogenic bacteria, where cells are held together by a structured extracellular matrix. When biofilms are associated with a host, chemical gradients and their origins become highly relevant. Conventional biofilm laboratory models such as multiwall biofilm models and agar plate models poorly mimic these gradients. In contrast, ex vivo models possess the partial capacity to mimic the conditions of tissue-associated biofilm and a biofilm associated with a mineralized surface enriched in inorganic components, such as the human dentin. This review will highlight the progress achieved using these settings for two models of persistent infections: the infection of the lung tissue by Pseudomonas aeruginosa and the infection of the root canal by Enterococcus faecalis. For both models, we conclude that the limitations of the conventional in vitro systems necessitate a complimentary experimentation with clinically relevant ex vivo models during therapeutics development. [ABSTRACT FROM AUTHOR]

The opportunistic pathogen Pseudomonas aeruginosa is often involved in airway infections of cystic fibrosis (CF) patients. It persists in the hostile CF lung environment, inducing chronic infections due to the production of several virulence factors. In this regard, the ability to form a biofilm plays a pivotal role in CF airway colonization by P. aeruginosa. Bacterial virulence mitigation and bacterial cell adhesion hampering and/or biofilm reduced formation could represent a major target for the development of new therapeutic treatments for infection control. Essential oils (EOs) are being considered as a potential alternative in clinical settings for the prevention, treatment, and control of infections sustained by microbial biofilms. EOs are complex mixtures of different classes of organic compounds, usually used for the treatment of upper respiratory tract infections in traditional medicine. Recently, a wide series of EOs were investigated for their ability to modulate biofilm production by different pathogens comprising S. aureus, S. epidermidis, and P. aeruginosa strains. Machine learning (ML) algorithms were applied to develop classification models in order to suggest a possible antibiofilm action for each chemical component of the studied EOs. In the present study, we assessed the biofilm growth modulation exerted by 61 commercial EOs on a selected number of P. aeruginosa strains isolated from CF patients. Furthermore, ML has been used to shed light on the EO chemical components likely responsible for the positive or negative modulation of bacterial biofilm formation. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa and Aspergillus fumigatus frequently coexist in the airways of immunocompromised patients or individuals with cystic fibrosis. Ciprofloxacin (CIP) is a synthetic quinolone antibiotic commonly used to treat bacterial infections, such as those produced by Pseudomonas aeruginosa. CIP binds iron, and it is unclear what effect this complex would have on the mycobiome. The effects of CIP on Aspergillus were dependent on the iron levels present, and on the presence of Aspergillus siderophores. We found that CIP alone stimulated wildtype planktonic growth, but not biofilm metabolism. At high concentrations, CIP antagonized a profungal effect of iron on wildtype Aspergillus metabolism, presumably owing to iron chelation. CIP interfered with the metabolism and growth of an Aspergillus siderophore mutant, with the effect on metabolism being antagonized by iron. CIP acted synergistically with iron on the growth of the mutant, and, to a lesser extent, the wildtype. In summary, CIP can increase fungal growth or affect fungal metabolism, depending on the local iron concentration and available siderophores. Therefore, high local CIP concentrations during treatment of Pseudomonas–Aspergillus co-infections may increase the fungal burden. [ABSTRACT FROM AUTHOR]

During the different waves of the coronavirus (COVID-19) pandemic, there has been an increased incidence of diabetes mellitus and diabetic foot infections. Among gram-negative bacteria, Pseudomonas aeruginosa is the predominant causative agent for diabetic foot ulcer infections in low-resource countries. P. aeruginosa possesses a variety of virulence factors, including biofilm formation. Biofilm formation is an important benchmark characteristic in the pathophysiology of diabetic foot ulceration. The main objective of the current study was to identify the most commonly isolated organisms and their antibiotic susceptibility patterns in diabetic foot patients during the COVID-19 pandemic. We also determined the genes associated with bacterial persistence and biofilm formation in the predominantly isolated organism. Accordingly, 100 wound swab samples were collected from diabetic foot patients from different hospitals in Alexandria, Egypt. Through phenotypic detection of biofilm formation, 93% (40) of the 43 P. aeruginosa isolates examined were categorized as biofilm producers. Molecular detection of the biofilm-encoding genes among the 43 P. aeruginosa isolates was as follows: algD (100%), pelF (88%) and pslD (49.7%), and this highlights a need for biofilm formation inhibitors to prevent the persistence of bacterial pathogens, and thus achieve better clinical outcomes in diabetic foot ulcer infections. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is the predominant cause of chronic biofilm infections that form in the lungs of people with cystic fibrosis (CF). These infections are highly resistant to antibiotics and persist for years in the respiratory tract. One of the main research challenges is that current laboratory models do not accurately replicate key aspects of a P. aeruginosa biofilm infection, highlighted by previous RNA-sequencing studies. We compared the P. aeruginosa PA14 transcriptome in an ex vivo pig lung (EVPL) model of CF and a well-studied synthetic cystic fibrosis sputum medium (SCFM). P. aeruginosa was grown in the EVPL model for 1, 2, and 7 days, and in vitro in SCFM for 1 and 2 days. The RNA was extracted and sequenced at each time point. Our findings demonstrate that expression of antimicrobial resistance genes was cued by growth in the EVPL model, highlighting the importance of growth environment in determining accurate resistance profiles. The EVPL model created two distinct growth environments: tissue-associated biofilm and the SCFM surrounding tissue, each cuing a transcriptome distinct from that seen in SCFM in vitro. The expression of quorum sensing associated genes in the EVPL tissue-associated biofilm at 48 h relative to in vitro SCFM was similar to CF sputum versus in vitro conditions. Hence, the EVPL model can replicate key aspects of in vivo biofilm infection that are missing from other current models. It provides a more accurate P. aeruginosa growth environment for determining antimicrobial resistance that quickly drives P. aeruginosa into a chronic-like infection phenotype. Pseudomonas aeruginosa lung infections that affect people with cystic fibrosis are resistant to most available antimicrobial treatments. The lack of a laboratory model that captures all key aspects of these infections hinders not only research progression but also clinical diagnostics. We used transcriptome analysis to demonstrate how a model using pig lungs can more accurately replicate key characteristics of P. aeruginosa lung infection, including mechanisms of antibiotic resistance and infection establishment. Therefore, this model may be used in the future to further understand infection dynamics to develop novel treatments and more accurate treatment plans. This could improve clinical outcomes as well as quality of life for individuals affected by these infections. [ABSTRACT FROM AUTHOR]

This research is aimed to eradicate the biofilm formed by bacteria causing wound infection through using proteinase K enzyme. For this purpose six different concentrations of proteinase K were used for the degradation of biofilm produced by Staphylococcus aureus and Pseudomonas areuginosa. These two species are the most common abundant bacteria causing infection by biofilm. Each of the concentrations was kept in contact with the pathogenic bacteria for 1, 2 and three hours. After 3 hours of incubation period the concentration (4 µg/ml) resulted in the highest eradication ability against S. aureus biofilm, while the same concentration was no significant in the eradication of Pseudomonas aeruginosa biofilm. [ABSTRACT FROM AUTHOR]

98 samples were collected from various clinical sources included (Burns, wounds, urines, sputums, blood) From the city of Baghdad, After performing the biochemical and microscopic examination, 52 isolates were obtained for Pseudomonas aeruginosa, 17 (32.7 %) isolates from burn infection, 12 (23%) isolates from Wound infection 11 (21.2 %) isolates from urine infection, 7 (13.5 %) isolates of sputum and 5 (9.6 %) isolates from blood. Bacteria susceptibility to form biofilm has been detectedby microtiter plate method, The results showed that 80 % of the bacterial isolates were produced the biofilm with different proportions, alg D gene (alginate production) has been detected by polymerase chain reaction (PCR) Which plays an essential role in the formation of the biofilm, The PCR results showed that the percentage of gene presence was(95.4 %). [ABSTRACT FROM AUTHOR]

The study was designed to explore the distribution and association of the biofilm genotype pattern(algD -/pslD -/pelF -) with multidrug-resistant in clinical local Pseudomonas aeruginosa isolates. A total of one hundred isolates of Pseudomonas aeruginosa were included in this study, which has been collected from different specimens, from July to September 2020. The isolates included were 34 from burns, 19 from wounds, 23 from ear infections, 22 from urinary tract infections (UTI), and 2 from cystic fibrosis (CF). Identification of the isolates was carried out using microscopical, cultural characterization on MacConkey agar, Cetrimide agar, then Pseudomonas agar. Biochemical tests were performed, and further identification was carried out by the VITEK_2_compact system. Genotypic identification has been completed by16SrRNA. To assess the frequency of multidrug-resistant of Pseudomonas aeruginosa (MDR), the antibiotic susceptibility test was done. It was carried out by using different groups of antibiotics (10 antibiotics) using the Kirby-Bauer disk diffusion method. The results showed that the resistance were Ceftazidime(62%), Gentamicin(26%, (Piperacillin-tazobactam(25%), Ticarcillin(24%), Meropenem(20%), Cefepime (18%), Amikacin(17%) Levofloxacin(16%), Colistin(15%) Imipenem(10%). Biofilm production was assessed using a microplate examination method. The results showed that 93% of isolates were positive for biofilm production, while (7%) were non-biofilm producers. There were differences in the rates of biofilm-production distributed into 21 (21%) were strong biofilm producer (OD was more than 2.156), 25 (25%) intermediate biofilm producer, and 47 (47%) were weak biofilm producer (OD was less than 1.078), and the non -biofilm producer was 7(7%). Three virulence factors genes (algD, pslD, and pelf) were chosen, which responsible for the phenotypic pattern of biofilm formation and identified as genotypic algD -/pslD -/pelF - pattern. The differences in genotypic pattern prevalence among the MDR-positive isolates of different origins were statistically significant. Chi-square analysis showed a highly significant association between strong biofilm capacity and genotype pattern (p<0.0001), also the analysis showed a highly significant association between moderate biofilm capacity and genotype pattern (p<0.002). Chi-square analysis showed a highly significant association between weak biofilm capacity and genotype pattern (p<0.001). In the current study the percentage of resistance among P. aeruginosa local isolates for multiple antibiotics (MDR) was relatively low, maybe due to the combination strategies based on appropriate anti-pseudoantibiotic agents that may be used to improve treatment from the related infections, according to these results, P. aeruginosa local isolates that produced biofilm were mostly (70%) indicated as non-MDR. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is the most prevalent bacterial species that contribute to cystic fibrosis (CF) respiratory failure. The impaired function of CF transmembrane conductance regulator leads to abnormal epithelial Cl–/HCO3– transport and acidification of airway surface liquid. However, it remains unclear why the CF lung is most commonly infected by Pseudomonas aeruginosa versus other pathogens. We carried out studies to investigate if lower pH helps Pseudomonas aeruginosa adapt and thrive in the CF-like acidic lung environment. Our results revealed that Pseudomonas aeruginosa generally forms more biofilm, induces antibiotic resistance faster in acidic conditions, and can be reversed by returning the acidic environment to physiologically neutral conditions. Pseudomonas aeruginosa appears to be highly adaptive to the CF-like acidic pH environment. By studying the effects of an acidic environment on bacterial response, we may provide a new therapeutic option in preventing chronic Pseudomonas aeruginosa infection and colonization. [ABSTRACT FROM AUTHOR]

Through experience and surveying undergraduate microbiology curriculum, we found that most schools discuss biofilms in lecture classes but not laboratory courses. Biofilms are a concern for both industry and the medical field and should be studied in student laboratories. To study this at an institution, there would need to be an affordable method to assay them. As a sample specimen, Pseudomonas aeruginosa was used as it proliferates into biofilms when it is starved for nutrients, which can be easily simulated in a laboratory environment. Known assays for studying biofilms are expensive, and most departments do not have the materials. In Trypticase Soy Broth (TSB), P. aeruginosa can also be induced to form a biofilm, but the technique is not seen very often because it has not been sufficiently standardized for undergraduate microbiology education laboratories. To account for the absence of specialized reagents, we incubated bacteria for longer periods of time as a means to diminish nutrients or starve the specimen. Using a two-tailed t-test, we were able to show that glass tubes inoculated with P. aeruginosa in TSB for 48 hours were forming distinct biofilms on the glass surface, which is appropriate for undergraduate microbiology laboratory studies. [ABSTRACT FROM AUTHOR]

Bacterial biofilms can lead to chronic infections, increase tolerance to antibiotics and disinfectants, resistance to phagocytosis, and other components of the body's immune system. Biofilm formation is implicated in the persistence of staphylococcal infections and chronic Pseudomonas aeruginosa lung infections in cystic fibrosis (CF) patients (which can result from biofilm-growing mucoid strains). Conventional treatments utilize aggressive antibiotic prophylaxis/therapy to prevent/eliminate biofilms, followed by chronic suppressive therapy. Recently, the use of enzymes to dissolve the biofilm matrix was investigated, in addition to quorum sensing inhibitors to increase biofilm susceptibility to antibiotics. Here, we propose a novel strategy, utilizing ultrasound-induced inertial cavitation, to increase antibiotic efficacy. The wall shear stress at the biofilm interface is calculated, and viscoplastic constitutive equations are used to examine the biofilm response to the mechanical stress. Our simulations suggest that the maximum biofilm detachment occurs at high pressure/low frequency, and the mechanical disruption can affect the biochemical processes inside the biofilm resulting in vulnerability to antibiotics. [ABSTRACT FROM AUTHOR]

Background & Objective: The ability of Pseudomonas aeruginosa to form biofilm has an important role in establishment of chronic phase of infections. Biofilm formation can be affected by antibiotics sub-MIC concentrations. The principal aim of the present study was to evaluate the effect of gentamicin at sub-MIC concentrations on biofilm formation in 100 Pseudomonas aeruginosa clinical isolates. Methods: Determination of minimal inhibitory concentration of gentamicin for clinical isolates was done using micro broth dilution method. The amount of biofilm formation in the treated and untreated isolates with gentamicin sub-MIC (1/2&1/4MIC) concentrations was evaluated using microtitre plate assay. pelA and pslA genes were detected in clinical isolates by PCR method. Results: 99% of clinical isolates were biofilm producer. Different changes in amount of biofilm formation were observed in the treated clinical isolates with sub-MIC concentrations of gentamicin. Two dominant changes were observed in 80% of clinical isolates. These concentrations had inhibitory effect on biofilm formation in 46.4% of isolates and caused a significant decrease in its amount. While in 31.3% of the isolates, the biofilm formation was significantly increased. The frequency of pelA and pslA genes among clinical isolates was 100%. Conclusion: gentamicin sub-MIC concentrations cause different changes on biofilm formation of Pseudomonas aeruginosa clinical isolates. Therefore, further studies are needed for discovering new treatment strategies and using sub-MIC concentrations of the antibiotic in prevention and treatment of Pseudomonas aeruginosa infections. [ABSTRACT FROM AUTHOR]

We compared the effects of disinfectants on biofilms of 10 Pseudomonas aeruginosa strains isolated from different loci of patients with purulent-septic infections. Identification was carried out by standard bacteriological methods. To substantiate the prospects of using in hospitals and to assess the effect of disinfectants on P. aeruginosa biofilms, the following disinfectants were used in various concentrations: Sekusept Aktiv, A-DEZ, and Monitor Oxy. All clinical strains of P. aeruginosa showed the ability to form biofilms. Both oxygen-containing and quaternary ammonium compounds effectively inhibited the formation of biofilms. In more than 50% cases, disinfectants with different active ingredients did not destroy pre-formed of P. aeruginosa biofilms. [ABSTRACT FROM AUTHOR]

Purpose: To investigate molecular characteristics, antimicrobial resistance, and biofilm formation ability of Pseudomonas aeruginosa strains isolated from patients with aural infections. Methods: Isolates (n = 199) were collected from ear discharges of patients with aural infections from January 2019 to December 2020. Antimicrobial susceptibility testing was performed according to the Clinical and Laboratory Standards Institute guidelines. All isolates were subjected to multilocus sequence typing (MLST) with amplification and sequencing of seven housekeeping genes. Biofilm formation and eradication were quantitatively assessed in microtiter plates. Genes associated with biofilm formation and the quinolone-resistance-determining region (QRDR) of genes gyrA and parC were investigated using polymerase chain reaction amplification and sequencing. Results: Of the 199 P. aeruginosa strains isolated, 109 (54.77%) were from females and 90 (45.23%) were from males. The isolates exhibited very low rates of resistance to most antibiotics tested, including piperacillin (1.51%), ceftazidime (0.50%), and imipenem (3.52%); however, the quinolones ciprofloxacin (80.40%) and levofloxacin (82.91%) were notable exceptions. The QRDR sequence results of the quinolone-resistant P. aeruginosa isolates showed Thr83Ile (n = 155) was the most common amino acid mutation in gyrA (n = 165), while Ser87Leu (n = 157) was widely detected in parC (n = 165). MLST analysis identified 34 sequence types (STs) with most isolates belonging to ST316 (73.87%). Almost all of the P. aeruginosa isolates (96.98%) produced biofilms and biofilm-forming genes algD (98.49%), pslD (96.98%), and pelF (96.48%) were highly prevalent. Conclusion: The P. aeruginosa strains isolated from aural discharges in this study exhibited very low rates of resistance to most antibiotics tested, except for the resistance rates to quinolones, which were relatively high. The isolates also exhibited a strong biofilm formation ability and low susceptibility to eradication, indicating that more effective drugs and treatment methods are needed to combat these infections. [ABSTRACT FROM AUTHOR]

Bacteria are increasingly relying on biofilms to develop resistance to antibiotics thereby resulting in their failure in treating many infections. In spite of continuous research on many synthetic and natural compounds, ideal anti-biofilm molecule is still not found thereby warranting search for new class of molecules. The current study focuses on exploring anti-biofilm potential of selenocystine against respiratory tract infection (RTI)-causing bacteria. Anti-bacterial and anti-biofilm assays demonstrated that selenocystine inhibits the growth of bacteria in their planktonic state, and formation of biofilms while eradicating preformed-biofilm effectively. Selenocystine at a MIC50 as low as 42 and 28 μg/mL effectively inhibited the growth of Klebsiella pneumonia and Pseudomonas aeruginosa. The antibacterial effect is further reconfirmed by agar cup diffusion assay and growth-kill assay. Selenocystine showed 30–60% inhibition of biofilm formation in K. pneumonia, and 44–70% in P. aeruginosa respectively. It also distorted the preformed-biofilms by degrading the eDNA component of the Extracellular Polymeric Substance matrix. Molecular docking studies of selenocystine with quorum sensing specific proteins clearly showed that through the carboxylic acid moiety it interacts and inhibits the protein function, thereby confirming its anti-biofilm potential. With further validation selenocystine can be explored as a potential candidate for the treatment of RTIs. [ABSTRACT FROM AUTHOR]

Background: Pulmonary infections associated with Pseudomonas aeruginosa can be life-threatening for patients suffering from chronic lung diseases such as cystic fibrosis. In this scenario, the formation of biofilms embedded in a mucus layer can limit the permeation and the activity of anti-infectives.Objectives: Native human pulmonary mucus can be isolated from endotracheal tubes, but this source is limited for large-scale testing. This study, therefore, aimed to evaluate a modified artificial sputum medium (ASMmod) with mucus-like viscoelastic properties as a surrogate for testing anti-infectives against P. aeruginosa biofilms.Methods: Bacterial growth in conventional broth cultures was compared with that in ASMmod, and PAO1-GFP biofilms were imaged by confocal microscopy. Transport kinetics of three antibiotics, tobramycin, colistin, and ciprofloxacin, through native mucus and ASMmod were studied, and their activity against PAO1 biofilms grown in different media was assessed by determination of metabolic activity and cfu.Results: PAO1(-GFP) cultured in human pulmonary mucus or ASMmod showed similarities in bacterial growth and biofilm morphology. A limited permeation of antibiotics through ASMmod was observed, indicating its strong barrier properties, which are comparable to those of native human mucus. Reduced susceptibility of PAO1 biofilms was observed in ASMmod compared with LB medium for tobramycin and colistin, but less for ciprofloxacin.Conclusions: These findings underline the importance of mucus as a biological barrier to antibiotics. ASMmod appears to be a valuable surrogate for studying mucus permeation of anti-infectives and their efficacy against PAO1 biofilms. [ABSTRACT FROM AUTHOR]

BACKGROUND: Antibacterial photocatalytic therapy has been employed as a promising strategy to combat antibiotic‐resistant bacteria in the water disinfection field, especially some non‐metal inorganic nanomaterials. However, their antibacterial activities on plant phytopathogens are poorly understood. Here, the photocatalytic antibacterial mechanism of the urea‐synthesized graphitic carbon nitride nanosheets (g‐C3N4 nanosheets) against Pseudomonas syringae pv. tabaci was systematically investigated in vitro and in vivo. RESULTS: The g‐C3N4 nanosheets exhibited remarkable concentration‐dependent and irradiation‐time‐dependent antibacterial properties, and the 0.5 mg mL−1 concentration ameliorated tobacco wildfire disease in host plants. Specifically, under visible irradiation, g‐C3N4 nanosheets produced numerous reactive oxygen species (ROS), supplementing the plentiful extracellular and intracellular ROS in bacteria. After exposing light‐induced g‐C3N4 nanosheets for 1 h, 500 genes were differentially expressed, according to transcriptome analyses. Notably, the expression of genes related 'antioxidant activity' and 'membrane transport' was sharply upregulated, and those related to 'bacterial chemotaxis', 'biofilm formation', 'energy metabolism' and 'cell motility' were downregulated. After exposure for over 2 h, the longer‐time pressure on the target bacteria cause the decreased biofilm formation and flagellum motility, further injuring the cell membranes leading to cytoplasm leakage and damaged DNA, eventually resulting in the bacterial death. Concomitantly, the attachment of g‐C3N4 nanosheets was a synergistic physical antibacterial pathway. The infection capacity assessment also supported the earlier supposition. CONCLUSION: These results provide novel insights into the photocatalytic antibacterial mechanisms of g‐C3N4 nanosheets at the transcriptome level, which are expected to be useful for dissecting the response pathways in antibacterial activities and for improving g‐C3N4‐based photocatalysts practices in plant disease control. © 2021 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Summary: Pseudomonas chlororaphis PCL1606 (PcPCL1606) displays plant‐colonizing features and exhibits antagonistic traits against soil‐borne phytopathogenic fungi. Biofilm formation could be relevant for the PcPCL1606 lifestyle, and in this study the role of some putative extracellular matrix components (EMC; Fap‐like fibre, alginate and Psl‐like polysaccharides) in the biofilm architecture and biocontrol activity of this bacterium were determined. EMC such as the Fap‐like fibre and alginate polysaccharide play secondary roles in biofilm formation in PcPCL1606, because they are not fundamental to its biofilm architecture in flow cell chamber, but synergistically they have shown to favour bacterial competition during biofilm formation. Conversely, studies on Psl‐like polysaccharide have revealed that it may contain mannose, and that it is strongly involved in the PcPCL1606 biofilm architecture and niche competition. Furthermore, the Fap‐like fibre and Psl‐like exopolysaccharide play roles in early surface attachment and contribute to biocontrol activity against the white root rot disease caused by Rosellinia necatrix in avocado plants. These results constitute the first report regarding the study of the extracellular matrix of the PcPCL1606 strain and highlight the importance of a putative Fap‐like fibre and Psl‐like exopolysaccharide produced by PcPCL1606 in the biofilm formation process and interactions with the host plant root. [ABSTRACT FROM AUTHOR]

Antimicrobial therapy is facing a worrisome and underappreciated challenge, the phenomenon of heteroresistance (HR). HR has been gradually documented in clinically relevant pathogens (e.g. Pseudomonas aeruginosa, Staphylococcus aureus, Burkholderia spp., Acinetobacter baumannii, Klebsiella pneumoniae, Candida spp.) towards several drugs and is believed to complicate the clinical picture of chronic infections. This type of infections are typically mediated by polymicrobial biofilms, wherein microorganisms inherently display a wide range of physiological states, distinct metabolic pathways, diverging refractory levels of stress responses, and a complex network of chemical signals exchange. This review aims to provide an overview on the relevance, prevalence, and implications of HR in clinical settings. Firstly, related terminologies (e.g. resistance, tolerance, persistence), sometimes misunderstood and overlapped, were clarified. Factors generating misleading HR definitions were also uncovered. Secondly, the recent HR incidences reported in clinically relevant pathogens towards different antimicrobials were annotated. The potential mechanisms underlying such occurrences were further elucidated. Finally, the link between HR and biofilms was discussed. The focus was to recognize the presence of heterogeneous levels of resistance within most biofilms, as well as the relevance of polymicrobial biofilms in chronic infectious diseases and their role in resistance spreading. These topics were subject of a critical appraisal, gaining insights into the ascending clinical implications of HR in antimicrobial resistance spreading, which could ultimately help designing effective therapeutic options. [ABSTRACT FROM AUTHOR]

Biofilms are complex microbial architectures that encase microbial cells in a matrix comprising self-produced extracellular polymeric substances. Microorganisms living in biofilms are much more resistant to hostile environments than their planktonic counterparts and exhibit enhanced resistance against the microbicides. From the human perspective, biofilms can be classified into beneficial, neutral, and harmful. Harmful biofilms impact food safety, cause plant and animal diseases, and threaten medical fields, making it urgent to develop effective and robust strategies to control harmful biofilms. In this review, we discuss various strategies to control biofilm formation on infected tissues, implants, and medical devices. We classify the current strategies into three main categories: (i) changing the properties of susceptible surfaces to prevent biofilm formation; (ii) regulating signalling pathways to inhibit biofilm formation; (iii) applying external forces to eradicate the biofilm. We hope this review would motivate the development of innovative and effective strategies for controlling harmful biofilms. [ABSTRACT FROM AUTHOR]

Vibrio cholerae is the causative agent of the diarrheal disease cholera, for which biofilm communities are considered to be environmental reservoirs. In endemic regions, and after algal blooms, which may result from phosphate enrichment following agricultural runoff, the bacterium is released from biofilms resulting in seasonal disease outbreaks. However, the molecular mechanism by which V. cholerae senses its environment and switches lifestyles from the biofilm‐bound state to the planktonic state is largely unknown. Here, we report that the major biofilm scaffolding protein RbmA undergoes autocatalytic proteolysis via a phosphate‐dependent induced proximity activation mechanism. Furthermore, we show that RbmA mutants that are defective in autoproteolysis cause V. cholerae biofilms to grow larger and mechanically stronger, correlating well with the observation that RbmA stability directly affects microbial community homeostasis and rheological properties. In conclusion, our biophysical study characterizes a novel phosphate‐dependent breakdown pathway of RbmA, while microbiological data suggest a new, sensory role of this biofilm scaffolding element. [ABSTRACT FROM AUTHOR]

The prevalence of antibacterial resistance has brought about a growing surge to develop novel approaches to control infectious diseases. Targeting Quorum-sensing (QS) - controlled virulence factors in bacteria has indicated a promising strategy for antipathogenic drugs. Extracts of Philippine Ilongot-Egongot ethnobotanicals Stachytarpeta jamaicensis, Adenanthera intermedia, Mikania micrantha, Hyptis suaveolens, Premna odorata, Cymbopogan winterianus, Phyllanthus urinaria, Dillenia philippinensis, Hydrocotyle vulgaris, Senna alata, Urena lobata, Ceiba pentandra, Ficus sp., Eleusine indica, Diplazium esculentum and Talahib (no known scientific name) were screened using microtiter plate biofilm formation assay for their QS inhibition activity against biofilm formation in Pseudomonas aeruginosa clinical isolate and Pseudomonas aeruginosa PNCM 1335. Extracts of M. micrantha, H. suaveolens flowers, H. vulgaris, A. intermedia, E. indica leaves, D. esculentum and Talahib have inhibitory effect on P. aeruginosa clinical isolate biofilm formation. Decrease in biofilm formation was shown in extracts of S. jamaicensis., M. micrantha, H. suaveolens, H. vulgaris, U. lobata, C. pentandra, A. intermedia, E. indica, D. esculentum and Talahib. against P. aeruginosa PNCM 1335. RNA extracts of M. micrantha, Talahib and A. intermedia exhibited downregulation of lasR in both test bacteria using qRT-PCR analysis through absolute quantification. The prospects of these ethnobotanicals to inhibit bacterial virulence avoiding antibiotic resistance is presented in this paper. [ABSTRACT FROM AUTHOR]

Bacterial biofilms are microbial lifestyles found in all environments. Up to 80% of human infections and 60–70% of hospital-acquired infections have a biofilm origin, with Staphylococcus aureus one of the leading causes of these infections. Microorganisms in biofilms exhibit significant antimicrobial resistance which poses important treatment challenges, hence the urgent need to identify novel antibiofilm strategies. Microbes form biofilms in response to various factors, and once these 3-dimentional structures form they are highly recalcitrant to removal. The switch from planktonic lifestyle to the biofilm protected mode of growth results in a phenotypic shift in the behavior of the microorganisms in terms of growth rate and gene expression. Given these changes, investigation of microbial gene expression and their modulation at different stages of biofilm maturation is needed to provide vital insight into the behavior of biofilm cells. In this study, we analyzed publicly available transcriptomic dataset of S. aureus biofilms at different stages of maturation to identify consistently upregulated genes irrespective of the biofilm maturation stage. Our reanalysis identified a total of 6 differentially expressed genes upregulated in both 48 and 144-h old S. aureus biofilms. Functional analysis revealed that these genes encode for proteins which play a role in key microbial metabolic pathways. However, these genes, as yet, are unrelated or fully studied in the context of biofilm. Moreover, the findings of this in silico work, suggest that these genes may represent potential novel targets for the development of more effective antibiofilm strategies against S. aureus biofilm-associated infections. [ABSTRACT FROM AUTHOR]

Individuals with cystic fibrosis (CF) are given antimicrobials as prophylaxis against bacterial lung infection, which contributes to the growing emergence of multidrug resistant (MDR) pathogens isolated. Pathogens such as Pseudomonas aeruginosa that are commonly isolated from individuals with CF are armed with an arsenal of protective and virulence mechanisms, complicating eradication and treatment strategies. While translation of phage therapy into standard care for CF has been explored, challenges such as the lack of an appropriate animal model demonstrating safety in vivo exist. In this review, we have discussed and provided some insights in the use of primary airway epithelial cells to represent the mucoenvironment of the CF lungs to demonstrate safety and efficacy of phage therapy. The combination of phage therapy and antimicrobials is gaining attention and has the potential to delay the onset of MDR infections. It is evident that efforts to translate phage therapy into standard clinical practice have gained traction in the past 5 years. Ultimately, collaboration, transparency in data publications and standardized policies are needed for clinical translation. [ABSTRACT FROM AUTHOR]

Aims: Relatively, few anti‐biofilm polysaccharides against Pseudomonas aeruginosa were done to investigate the underlying molecular mechanism. Exopolysaccharide EPS273 can clearly reduce biofilm formation and infection of P. aeruginosa. This study aims to investigate its anti‐biofilm and anti‐infection mechanism on transcriptional level. Methods and Results: Herein, we used an RNA‐Seq transcriptomic approach to investigate the underlying anti‐biofilm and anti‐infection mechanism of EPS273. The expression levels of a large number of genes were changed after P. aeruginosa PAO1 was treated with EPS273. Especially, the genes related to biofilm formation, such as gene involved in production of extracellular matrix and virulence factor, genes involved in flagella and cell motility and genes involved in iron acquisition. Notably, the expression levels of genes involved in regulatory and signal transduction were markedly downregulated, such as two‐component system PhoP‐PhoQ and quorum sensing (QS) system LasI/LasR and RhlI/RhlR. Furthermore, when genes phoP and phoQ were disrupted, respectively, the reduction of biofilm formation and cell motility in mutant △phoP or △phoQ was also detected. Conclusion: EPS273 may exert its anti‐biofilm and anti‐infection function by downregulating gene expression of two‐component system PhoP‐PhoQ and QS systems LasI/LasR and RhlI/RhlR of P. aeruginosa, which further regulated expression of genes involved in biofilm formation. Significance and Impact of the Study: Our data will expand understanding of anti‐biofilm mechanisms of polysaccharides on transcriptomic level. [ABSTRACT FROM AUTHOR]

Stenotrophomonas maltophilia is one of the most frequently isolated multidrug-resistant nosocomial opportunistic pathogens. It contributes to disease progression in cystic fibrosis (CF) patients and is frequently isolated from wounds, infected tissues, and catheter surfaces. On these diverse surfaces S. maltophilia lives in single-species or multispecies biofilms. Since very little is known about common processes in biofilms of different S. maltophilia isolates, we analyzed the biofilm profiles of 300 clinical and environmental isolates from Europe of the recently identified main lineages Sgn3, Sgn4, and Sm2 to Sm18. The analysis of the biofilm architecture of 40 clinical isolates revealed the presence of multicellular structures and high phenotypic variability at a strain-specific level. Further, transcriptome analyses of biofilm cells of seven clinical isolates identified a set of 106 shared strongly expressed genes and 33 strain-specifically expressed genes. Surprisingly, the transcriptome profiles of biofilm versus planktonic cells revealed that just 9.43% 1.36% of all genes were differentially regulated. This implies that just a small set of shared and commonly regulated genes is involved in the biofilm lifestyle. Strikingly, iron uptake appears to be a key factor involved in this metabolic shift. Further, metabolic analyses implied that S. maltophilia employs a mostly fermentative growth mode under biofilm conditions. The transcriptome data of this study together with the phenotypic and metabolic analyses represent so far the largest data set on S. maltophilia biofilm versus planktonic cells. This study will lay the foundation for the identification of strategies for fighting S. maltophilia biofilms in clinical and industrial settings. [ABSTRACT FROM AUTHOR]

Co-cultures allow for the study of cell–cell interactions between different eukaryotic species or with bacteria. Such an approach has enabled researchers to more closely mimic complex tissue structures. This review is focused on co-culture systems modelling the oral cavity, which have been used to evaluate this unique cellular environment and understand disease progression. Over time, these systems have developed significantly from simple 2D eukaryotic cultures and planktonic bacteria to more complex 3D tissue engineered structures and biofilms. Careful selection and design of the co-culture along with critical parameters, such as seeding density and choice of analysis method, have resulted in several advances. This review provides a comparison of existing co-culture systems for the oral environment, with emphasis on progression of 3D models and the opportunity to harness techniques from other fields to improve current methods. While filling a gap in navigating this literature, this review ultimately supports the development of this vital technique in the field of oral biology. [ABSTRACT FROM AUTHOR]

Bacteria and Candida albicans are prominent gut microbiota, and the translocation of these organisms into blood circulation might induce mixed-organism biofilms, which warrants the exploration of mixed- versus single-organism biofilms in vitro and in vivo. In single-organism biofilms, Acinetobacter baumannii and Pseudomonas aeruginosa (PA) produced the least and the most prominent biofilms, respectively. C. albicans with P. aeruginosa (PA+CA) induced the highest biofilms among mixed-organism groups as determined by crystal violet straining. The sessile form of PA+CA induced higher macrophage responses than sessile PA, which supports enhanced immune activation toward mixed-organism biofilms. In addition, Candida incubated in pre-formed Pseudomonas biofilms (PA>CA) produced even higher biofilms than PA+CA (simultaneous incubation of both organisms) as determined by fluorescent staining on biofilm matrix (AF647 color). Despite the initially lower bacteria during preparation, bacterial burdens by culture in mixed-organism biofilms (PA+CA and PA>CA) were not different from biofilms of PA alone, supporting Candida -enhanced Pseudomonas growth. Moreover, proteomic analysis in PA>CA biofilms demonstrated high AlgU and mucA with low mucB when compared with PA alone or PA+CA, implying an alginate-related mucoid phenotype in PA>CA biofilms. Furthermore, mice with PA>CA biofilms demonstrated higher bacteremia with more severe sepsis compared with mice with PA+CA biofilms. This is possibly due to the different structures. Interestingly, l-cysteine, a biofilm matrix inhibitor, attenuated mixed-organism biofilms both in vitro and in mice. In conclusion, Candida enhanced Pseudomonas alginate–related biofilm production, and Candida presentation in pre-formed Pseudomonas biofilms might alter biofilm structures that affect clinical manifestations but was attenuated by l-cysteine. [ABSTRACT FROM AUTHOR]

Proteus mirabilis is the third most common etiological factor of urinary tract infection. It produces urease, which contributes to the formation of a crystalline biofilm, considered to be one of the most important virulence factors of P. mirabilis strains, along with their ability to swarm on a solid surface. The aim of this study was to analyze the pathogenic properties of two selected groups of clinical P. mirabilis isolates, antimicrobial susceptible and multidrug resistant (MDR), collected from hospitals in different regions in Poland. The strains were examined based on virulence gene profiles, urease and hemolysin production, biofilm formation, and swarming properties. Additionally, the strains were characterized based on the Dienes test and antibiotic susceptibility patterns. It turned out that the MDR strains exhibited kinship more often than the susceptible ones. The strains which were able to form a stronger biofilm had broader antimicrobial resistance profiles. It was also found that the strongest swarming motility correlated with susceptibility to most antibiotics. The correlations described in this work encourage further investigation of the mechanisms of pathogenicity of P. mirabilis. [ABSTRACT FROM AUTHOR]

Piscirickettsia salmonis is a facultative intracellular bacterium that generates piscirickettsiosis affecting salmonids in Chile. The bacterium has the adaptability to survive in the marine environment under multiple stressful conditions. In this sense, this work focused on the analysis of a gene battery associated with biofilm formation under different culture conditions and on the adaptability of this biofilm to different media. The results indicated that the strains LF-89, IBM-034 and IBM-040 were strong biofilm producers, evidencing adaptability to the media by increasing the amount of biofilm through successive growths. Transcript levels of six genes described in various bacteria and P. salmonis , considered to have metabolic functions, and playing a relevant role in biofilm formation, were analyzed to evaluate bacterial functionality in the biofilm. The genes mazE-mazF , implicated in biofilm and stress, were markedly overexpressed in the biofilm condition in the three strains. For its part, gene gltA , an indicator of metabolic activity and related to virulence inhibition in Salmonella typhimurium , also seems to restrain the pathogenesis process in P. salmonis by inhibiting the expression of the virulence-associated genes liso and tcf. Finally, the expression of the glnA gene suggests the use of glutamine as an essential element for the growth of the biofilm. [ABSTRACT FROM AUTHOR]

Background: Bacterial biofilms pose a challenge in treating implant-associated infections. Biofilms provide bacteria with protection against antimicrobial agents and the immune response and often are invisible to the naked eye. As a biofilm-disclosing agent, methylene blue (MB) has shown promise, but lacks rigorous in vitro evaluation. The purposes of the present study were to assess MB as a biofilm-disclosing agent in vitro for common biofilm-forming organisms and to determine performance characteristics across implant materials and healthy tissue types.Methods: Staphylococcus aureus (ATCC 6538) and Pseudomonas aeruginosa (ATCC 27853) biofilms were grown on culture for 2 days in CDC biofilm reactors on titanium, cobalt chromium, polyethylene, and polyether ether ketone (PEEK) coupons. Biofilms were stained with MB solutions of either 0.005% or 0.01% and then were washed with normal saline solution. Digital photographs were obtained to compare the visual sensitivity of the blue dye at these dilutions. Scanning electron microscopy (SEM) was performed to confirm the absence or presence of biofilm on MB-stained areas. Uninoculated controls were also assessed. Healthy adult sheep tissues were also stained to determine the staining characteristics of the host tissue. ImageJ was used to determine the relative blue intensity of stained implants and tissues compared with standard curves.Results: S. aureus and P. aeruginosa biofilms stained avidly on titanium, cobalt chromium, polyethylene, and PEEK coupons. There was visible dose-dependent staining based on dye concentration. MB was visible only where biofilms were present as confirmed by SEM. MB did not stain uninoculated controls. Articular cartilage and meniscus demonstrated appreciable staining; bone, tendon, muscle, nerve, and fat did not. Bacterial biofilms demonstrated both dose-dependent and species-specific staining.Conclusions: MB is an effective disclosing agent for S. aureus and P. aeruginosa biofilms in vitro. MB did not stain implant materials, nor did it stain most healthy tissues in vitro. MB may allow surgeons to see biofilms and may allow for enhanced debridement once visualized. [ABSTRACT FROM AUTHOR]

The current study was designed to explore the association between the pigments production and biofilm construction in local Pseudomonas aeruginosa isolates. Out of 143 patients suffering from burns, urinary tract infections (UTI), respiratory tract infections and cystic fibrosis obtained from previous study by Mahmood (2015), twenty two isolates (15.38%) were identified from (11) hospitals in Iraq, splitted into three provinces, Baghdad, Al-Anbar and Karbala for the duration of June 2017 to April 2018. Characterization was carried out by using microscopical, morphological and biochemical methods which showed that all these isolates belong to P. aeruginosa. Screening of biofilm production isolates was carried out by using nutrient broth supplemented with glucose (0.25%) production medium which encourage this biofilm production. The percentage of pigmented isolates were collected from a total of 143 samples, 2.8% of the isolates from burns, 2.1% isolates from cystic fibrosis and 0.7% isolates from UTI. Quantitative assays for biofilm formation were conducted using ELIZA technique. The results showed that all (22) isolates produced biofilm except one (B1 isolate). Biofilm quantities were varied from strong to medium production in comparison with control (0.0663). Statistical analysis results using Fischer's Exact test (p<0.05) were nonsignificant, therefore the pigment production has no association with biofilm formation for all of them. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa biofilm-related infections are difficult to treat with antibiotics. Along the different layers of the biofilm, the P. aeruginosa population is heterogeneous, exhibiting an extreme ability to adapt his metabolic activity to the local microenvironment. At the deepest layers of the biofilm is a subset of dormant cells, called persister cells. Though antimicrobial failure might be multifactorial, it is now demonstrated that these persister cells, genetically identical to a fully susceptible strain, but phenotypically divergent, are highly tolerant to antibiotics, and contribute to antimicrobial failure. By eradicating susceptible, metabolically active cells, antibiotics bring out pre-existing persister cells. The biofilm mode of growth creates microenvironment conditions that activate stringent response mechanisms, SOS response and toxin-antitoxin systems that render the bacterial population highly tolerant to antibiotics. Using diverse, not standardized, models of biofilm infection, a large panel of antibiotic regimen has been evaluated. They demonstrated that biofilm growth had an unequal impact of antibiotic activity, colistin and meropenem being the less impacted antibiotics. Different combination and sequential antimicrobial therapies were also evaluated, and could be partially efficient, but none succeeded in eradicating persister cells, so that non-antibiotic alternative strategies are currently under development. This article reviews the molecular mechanisms involved in antibiotic tolerance and persistence in P. aeruginosa biofilm infections. A review of the antimicrobial regimen evaluated for the treatment of P. aeruginosa biofilm infection is also presented. While tremendous progress has been made in the understanding of biofilm-related infections, alternative non-antibiotic strategies are now urgently needed. [ABSTRACT FROM AUTHOR]

Population-wide tolerance and persisters enable susceptible bacterial cells to endure hostile environments, including antimicrobial exposure. The SOS response can play a significant role in the generation of persister cells, population-wide tolerance, and shielding. The SOS pathway is an inducible DNA damage repair system that is also pivotal for bacterial adaptation, pathogenesis, and diversification. In addition to the two key SOS regulators, LexA and RecA, some other stressors and stress responses can control SOS factors. Bacteria are exposed to DNA-damaging agents and other environmental and intracellular factors, including cigarette smoke, that trigger the SOS response at a number of sites within the host. The Escherichia coli TisB/IstR module is as yet the only known SOS-regulated toxin–antitoxin module involved in persister formation. Nevertheless, the SOS response plays a key role in the formation of biofilms that are highly recalcitrant to antimicrobials and can be abundant in persisters. Furthermore, the dynamic biofilm environment generates DNA-damaging factors that trigger the SOS response within the biofilm, fueling bacterial adaptation and diversification. This review highlights the SOS response in relation to antimicrobial recalcitrance to antimicrobials in four clinically significant species, Escherichia coli , Pseudomonas aeruginosa , Staphylococcus aureus , and Mycobacterium tuberculosis. [ABSTRACT FROM AUTHOR]

Objectives: Patients with immunodeficiency or cystic fibrosis frequently suffer from respiratory fungal infections. In particular, biofilm-associated fungi cause refractory infection manifestations, linked to increased resistance to anti-infective agents. One emerging filamentous fungus is Lomentospora prolificans. Here, the biofilm-formation capabilities of L. prolificans isolates were investigated and the susceptibility of biofilms to various antifungal agents was analysed.Methods: Biofilm formation of L. prolificans (n = 11) was estimated by crystal violet stain and antibiofilm activity was additionally determined via detection of metabolically active biofilm using an XTT assay. Amphotericin B, micafungin, voriconazole and olorofim were compared with regard to their antibiofilm effects when added prior to adhesion, after adhesion and on mature and preformed fungal biofilms. Imaging via confocal laser scanning microscopy was carried out to demonstrate the effect of drug treatment on the fungal biofilm.Results: Antibiofilm activities of the tested antifungal agents were shown to be most effective on adherent cells whilst mature biofilm was the most resistant. The most promising antibiofilm effects were detected with voriconazole and olorofim. Olorofim showed an average minimum biofilm eradication concentration (MBEC) of 0.06 mg/L, when added prior to and after adhesion. The MBECs of voriconazole were ≤4 mg/L. On mature biofilm the MBECs of olorofim and voriconazole were higher than the previously determined MICs against planktonic cultures. In contrast, amphotericin B and especially micafungin did not exhibit sufficient antibiofilm activity against L. prolificans.Conclusions: To our knowledge, this is the first study demonstrating the antibiofilm potential of olorofim against the human pathogenic fungus L. prolificans. [ABSTRACT FROM AUTHOR]

Biofilms are microbial communities established in the self‐produced extracellular substances that include up to 80% of associated microbial infections. During biofilm formation, bacterial cells shift from the planktonic forms to aggregated forms surrounded by an extracellular polymeric substance. The bacterial biofilm shows resistance against immune reactions as well as antibiotics and is potentially able to cause disorders by both device‐related and nondevice‐related infections. The nondevice‐related bacterial biofilm infections include dental plaque, urinary tract infections, cystic fibrosis, otitis media, infective endocarditis, tonsillitis, periodontitis, necrotizing fasciitis, osteomyelitis, infectious kidney stones, and chronic inflammatory diseases. In this review, we will summarize and examine the literature about bacterial biofilm infections unrelated to indwelling devices. [ABSTRACT FROM AUTHOR]

In clinical environments, many serious antibiotic-resistant infections are caused by biofilm-forming species. This presents issues when attempting to determine antimicrobial dosing as traditional antibiotic susceptibility tests (ASTs) are typically designed around planktonic bacteria and thus offer information that is not relevant to the biofilm phenotype present in the patient. Even the popular Calgary biofilm device may provide inaccurate minimum biofilm inhibitory concentrations (MBICs) and can be time- and material-intensive. In this work, we present a method utilizing oxygen-sensitive nanosensor technology to monitor the oxygen consumption dynamics of living biofilms as they are exposed to antibiotics. We incorporated our nanosensors into biofilms grown from P. aeruginosa strains of varying sensitivity to traditional classes of antibiotics. Through measuring nanosensor response under antibiotic administration we determined the concentrations able to cease biofilm metabolism. This method provides information on the MBIC as well as kinetic response information in a manner that requires fewer materials and is more reflective of biofilm behavior than a traditional AST. [ABSTRACT FROM AUTHOR]