Bacterial biofilm prevalence in dental unit waterlines: a systematic review and meta-analysis.

BACKGROUNDS: Numerous studies have shown that dental unit water lines (DUWLs) are often contaminated by a wide range of micro-organisms (bacteria, fungi, protozoa) and various prevalence have been reported for it in previous studies. Therefore, this review study aims to describe the prevalence of bacterial biofilm contamination of DUWLs. METHODS: This is a systematic review and meta-analysis in which the related keywords in different international databases, including Medline (via PubMed) and Scopus were searched. The retrieved studies were screened and the required data were extracted from the included studies. Three standard methods including American Dental Association (ADA), The Center for Disease Control and Prevention (CDC) and contaminated > 100 CFU/ml(C-100) standards were used to assess the bacterial biofilm contamination of DUWLs. All studies that calculated the prevalence of bacterial biofilm contamination of DUWLs, and English full-text studies were included in the meta-analysis. Studies that did not have relevant data or used unusual laboratory methods were excluded. Methodological risk of bias was assessed by a related checklist and finally, the data were pooled by fixed or random-effect models. RESULTS: Seven hundred and thirty-six studies were identified and screened and 26 related studies were included in the meta-analysis. The oldest included study was published in 1976 and the most recent study was published in 2020. According to the ADA, CDC and C-100 standards, the prevalence of bacterial contamination was estimated to be 85.0% (95% confidence interval (CI): 66.0-94.0%), 77.0% (95%CI: 66.0-85.0%) and 69.0% (95%CI: 67.0-71.0%), respectively. The prevalence of Legionella Pneumophila and Pseudomonas Aeruginosa in DUWLs was estimated to be 12.0% (95%CI: 10.0-14.0%) and 8.0% (95%CI: 2.0-24.0%), respectively. CONCLUSION: The results of this review study suggested a high prevalence of bacterial biofilm in DUWLs; therefore, the use of appropriate disinfecting protocol is recommended to reduce the prevalence of contamination and reduce the probable cross-infection.

A Case of Coccidioidal Meningitis With Biofilm Obstructing VP Shunt Due to Cutibacterium acnes.

Herein described is a case of biofilm obstructing ventriculoperitoneal shunt due to Cutibacteirum acnes infection in a patient with coccidioidal meningitis. Cutibacterium acnes infects and obstructs cerebral shunts by the production of biofilm; however, diagnosis is usually missed by routine aerobic cultures. Obtaining anaerobic cultures routinely in patients with foreign body implants leading to central nervous system infections could prevent a missed diagnosis of this pathogen. Penicillin G is the first-line treatment.

Biofilm Formation From Listeria monocytogenes Isolated From Pangasius Fish-processing Plants.

Biofilm formation of Listeria monocytogenes in food processing environments cause potential source of cross-contamination to foodstuffs; hence, the control of biofilm is currently addressed to find effective solutions for preventing biofilm formation or eliminating the established one. Forty-five strains of Listeria monocytogenes isolated from Pangasius fish-processing plants were studied for their capability to form a biofilm on 96-well microtiter plate by using the conventional crystal violet staining. Additionally, the inhibitory effect of biofilm formation by food additives including monascus pigment and ε-polylysine was examined. The average OD value showing biofilm mass of all 45 strains L. monocytogenes increased with an increasing temperature and time (p < 0.05). Monascus pigment and ε-polylysine significantly decreased biofilm formation by 80 ± 5.5% and 20 ± 5.9%, respectively, at the tested concentration (p < 0.05) Further, the effects of lysozyme (0.1 mg/mL) alone or in combination with slightly acidic hypochlorous water (SAHW) with 40 mg/L available chlorine or sodium hypochlorite (NaOCl) with 100 mg/L available chlorine against 7-d established biofilm of L. monocytogenes were investigated. The results indicated that slightly acidic hypochlorous water alone exhibited significant antibacterial activity (p < 0.05), decreasing the viable count by 5.2 ± 0.5 log CFU/mL. It seems that sequential treatment of lysozyme and SAHW showed an additional efficacy against biofilm of L. monocytogenes on polystyrene plate surface, reducing 70% of biomass of biofilm and 7.6 ± 0.3 log of biofilm viable cells (p < 0.05). Additionally, SAHW exhibited greater bactericidal activity against viable biofilm cells than NaOCl did. This result reveals that SAHW is a promising disinfectant agent against L. monocytogenes and the potential alternative to NaOCl in practice.

Inactivation of Staphylococcus aureus by Levulinic Acid Plus Sodium Dodecyl Sulfate and their Antibacterial Mechanisms on S. aureus Biofilms by Transcriptomic Analysis.

The combination of levulinic acid (LVA) and sodium dodecyl sulfate (SDS) in recent years has shown a considerable potential to use as an antimicrobial intervention. The objectives of this study were to evaluate the antimicrobial efficacy of the combination against Staphylococcus aureus in both planktonic and biofilm states and to investigate the transcriptional changes in S. aureus biofilms coincubated with sublethal concentrations of LVA and/or SDS. The minimum inhibitory concentrations (MICs) of LVA and SDS determined by the microdilution method were 3.125 and 0.039 mg/mL, respectively. An additive bacteriostatic interaction (fractional inhibitory concentration index = 1) between the two compounds was observed by the checkerboard assay, whereas a synergistic bactericidal activity was displayed by the time-kill assay. The biomass and viable cells in the biofilms were reduced by both antimicrobials either alone or in combination in a dose-dependent manner. Transcriptomics indicated that more differentially expressed (DE) genes were observed in the biofilm treated with SDS (103 up- and 205 downregulated DE genes) and LVA + SDS (187 up and 162 down) than that coincubated with LVA (34 up and 32 down). The SDS and LVA + SDS treatments mainly affected the expression of genes responsible for cell surface proteins, virulence factors, adhesins, and capsular polysaccharides. Both the antibiofilm assay and the transcriptomics indicated that SDS, not LVA, was the major chemical contributing to the antibacterial efficacy of the combination. This study reveals the behavioral responses and protective mechanisms of S. aureus to LVA and SDS applied individually or in combination.

Single layer centrifugation (SLC) for bacterial removal with Porcicoll positively modifies chromatin structure in boar spermatozoa.

The storage of boar semen samples at 17 °C for artificial insemination (AI) doses enables the proliferation of the bacteria, making antibiotics necessary. This can contribute to the development of antimicrobial resistance (AMR). This study tested bacterial presence and sperm chromatin structure after using a low-density colloid (Porcicoll) as an antibiotic alternative to eliminate bacteria. Ejaculates (8 boars, 3 ejaculates each) were split as control and low-density colloid centrifugation (single layer centrifugation, SLC, 20%, and 30% Porcicoll) into 500 ml tubes. Analyses were carried out at days 0, 3, and 7 (17 °C) for microbial presence and sperm chromatin structure analysis: %DFI (DNA fragmentation) and %HDS (chromatin immaturity), monobromobimane (mBBr; free thiols and disulfide bridges), and chromomycin A3 (CMA3; chromatin compaction). Besides comparing bacterial presence (7 species identified) and chromatin variables between treatments, the associations between these sets of variables were described by canonical correlation analysis (CCA). Results showed a significant decrease of some bacteria or a complete removal after SLC (especially for P30). SLC also caused a decrease of %HDS and an increase of disulfide bridges and low and medium mBBr populations, suggesting the removal of immature sperm (poor chromatin compaction). CCA showed an association pattern compatible with the degradation of sperm chromatin parameters with bacterial contamination, especially Enterobacteria, P. aeuriginosa, and K. variicola. In conclusion, bacterial contamination affects sperm chromatin beyond DNA fragmentation; SLC with low-density colloid not only removes bacteria from boar semen, but also chromatin structure is enhanced after selection.

One-Pot Preparation of Cetylpyridinium Chloride-Containing Nanoparticles for Biofilm Eradication.

Quaternary ammonium compounds (QACs) have been widely used due to their excellent antimicrobial activity. However, using the technology where nanomaterials are employed as drug carriers to deliver QAC drugs has not been fully explored. In this study, mesoporous silica nanoparticles (MSNs) with short rod morphology were synthesized in a one-pot reaction using an antiseptic drug cetylpyridinium chloride (CPC). CPC-MSN were characterized via various methods and tested against three bacterial species (Streptococcus mutans, Actinomyces naeslundii, and Enterococcus faecalis), which are associated with oral infections, caries, and endodontic pathology. The nanoparticle delivery system used in this study prolonged the release of CPC. The manufactured CPC-MSN effectively killed the tested bacteria within the biofilm, and their size allowed them to penetrate into dentinal tubules. This CPC-MSN nanoparticle delivery system demonstrates potential for applications in dental materials.

Combination d-Amino Acid and Photothermal Hydrogel for the Treatment of Prosthetic Joint Infections.

Prosthetic joint infection (PJI) is a devastating complication requiring surgical intervention and prolonged antimicrobial treatment. The prevalence of PJI is on the rise, with an average incidence of 60,000 cases per year and a projected annual cost of $1.85 billion in the US. The underlying pathogenesis of PJI involves the formation of bacterial biofilms that protect the pathogen from the host immune response and antibiotics, making it difficult to eradicate such infections. Biofilms on implants are also resistant to mechanical brushing/scrubbing methods of removal. Since the removal of biofilms is currently only achievable by the replacement of the prosthesis, therapies aimed at eradicating biofilms while enabling retention of implants will revolutionize the management of PJIs. To address severe complications associated with biofilm-related infections on implants, we have developed a combination treatment that is based on a hydrogel nanocomposite system, containing d-amino acids (d-AAs) and gold nanorods, which can be delivered and transforms from a solution to a gel state at physiological temperature for sustained release of d-AAs and light-activated thermal treatment of infected sites. Using this two-step approach to utilize a near-infrared light-activated hydrogel nanocomposite system for thermal treatment, following initial disruption with d-AAs, we were able to successfully demonstrate in vitro the total eradication of mature Staphylococcus aureus biofilms grown on three-dimensional printed Ti-6Al-4V alloy implants. Using a combination of cell assays, computer-aided scanning electron microscopy analyses, and confocal microscopy imaging of the biofilm matrix, we could show 100% eradication of the biofilms using our combination treatment. In contrast, we were only able to see 25% eradication of the biofilms using the debridement, antibiotics, and implant retention method. Moreover, our hydrogel nanocomposite-based treatment approach is adaptable in the clinical setting and capable of combating chronic infections brought about by biofilms on medical implants.

Perspective: The viscoelastic properties of biofilm infections and mechanical interactions with phagocytic immune cells.

Biofilms are viscoelastic materials that are a prominent public health problem and a cause of most chronic bacterial infections, in large part due to their resistance to clearance by the immune system. Viscoelastic materials combine both solid-like and fluid-like mechanics, and the viscoelastic properties of biofilms are an emergent property of the intercellular cohesion characterizing the biofilm state (planktonic bacteria do not have an equivalent property). However, how the mechanical properties of biofilms are related to the recalcitrant disease that they cause, specifically to their resistance to phagocytic clearance by the immune system, remains almost entirely unstudied. We believe this is an important gap that is ripe for a large range of investigations. Here we present an overview of what is known about biofilm infections and their interactions with the immune system, biofilm mechanics and their potential relationship with phagocytosis, and we give an illustrative example of one important biofilm-pathogen (Pseudomonas aeruginosa) which is the most-studied in this context. We hope to inspire investment and growth in this relatively-untapped field of research, which has the potential to reveal mechanical properties of biofilms as targets for therapeutics meant to enhance the efficacy of the immune system.

Effects of BlueM® against Streptococcus mutans biofilm and its virulence gene expression.

This study evaluated the antimicrobial capacity of BlueM® mouthwash against the bacterium Streptococcus mutans and its influence on gbpA gene expression as well as its cytotoxic effect on fibroblast cells. BlueM® showed antimicrobial activity, with MIC and MBC values of 0.005% and 0.01%, respectively. The MBIC was 6.25% for S. mutans. CFU count and confocal microscopy revealed significant effect of BlueM® on S. mutans biofilm pre-formed on dentin surfaces. Interestingly, the analysis of gbpA gene expression indicated a decrease in gene expression after 15 min of treatment with BlueM® at a concentration of 25%. Moreover, BlueM® exhibited low levels of cytotoxicity. In conclusion, our results showed the antimicrobial effectiveness of BlueM® against S. mutans, its ability to modulate the expression of the gbpA gene and its low cytotoxicity. This study supports the therapeutic potential of BlueM® as an alternative agent for the control of oral biofilm.

Comparative genome identification of accessory genes associated with strong biofilm formation in Vibrio parahaemolyticus.

Vibrio parahaemolyticus biofilms on the seafood processing plant surfaces are a potential source of seafood contamination and subsequent food poisoning. Strains differ in their ability to form biofilm, but little is known about the genetic characteristics responsible for biofilm development. In this study, pangenome and comparative genome analysis of V. parahaemolyticus strains reveals genetic attributes and gene repertoire that contribute to robust biofilm formation. The study identified 136 accessory genes that were exclusively present in strong biofilm forming strains and these were functionally assigned to the Gene Ontology (GO) pathways of cellulose biosynthesis, rhamnose metabolic and catabolic processes, UDP-glucose processes and O antigen biosynthesis (p < 0.05). Strategies of CRISPR-Cas defence and MSHA pilus-led attachment were implicated via Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation. Higher levels of horizontal gene transfer (HGT) were inferred to confer more putatively novel properties on biofilm-forming V. parahaemolyticus. Furthermore, cellulose biosynthesis, a neglected potential virulence factor, was identified as being acquired from within the order Vibrionales. The cellulose synthase operons in V. parahaemolyticus were examined for their prevalence (22/138, 15.94 %) and were found to consist of the genes bcsG, bcsE, bcsQ, bcsA, bcsB, bcsZ, bcsC. This study provides insights into robust biofilm formation of V. parahaemolyticus at the genomic level and facilitates: identification of key attributes for robust biofilm formation, elucidation of biofilm formation mechanisms and development of potential targets for novel control strategies of persistent V. parahaemolyticus.

The role of nanocomposites against biofilm infections in humans.

The use of nanomaterials in several fields of science has undergone a revolution in the last few decades. It has been reported by the National Institutes of Health (NIH) that 65% and 80% of infections are accountable for at least 65% of human bacterial infections. One of their important applications in healthcare is the use of nanoparticles (NPs) to eradicate free-floating bacteria and those that form biofilms. A nanocomposite (NC) is a multiphase stable fabric with one or three dimensions that are much smaller than 100 nm, or systems with nanoscale repeat distances between the unique phases that make up the material. Using NC materials to get rid of germs is a more sophisticated and effective technique to destroy bacterial biofilms. These biofilms are refractory to standard antibiotics, mainly to chronic infections and non-healing wounds. Materials like graphene and chitosan can be utilized to make several forms of NCs, in addition to different metal oxides. The ability of NCs to address the issue of bacterial resistance is its main advantage over antibiotics. This review highlights the synthesis, characterization, and mechanism through which NCs disrupt Gram-positive and Gram-negative bacterial biofilms, and their relative benefits and drawbacks. There is an urgent need to develop materials like NCs with a larger spectrum of action due to the rising prevalence of human bacterial diseases that are multidrug-resistant and form biofilms.

Intermittent antibiotic treatment of bacterial biofilms favors the rapid evolution of resistance.

Bacterial antibiotic resistance is a global health concern of increasing importance and intensive study. Although biofilms are a common source of infections in clinical settings, little is known about the development of antibiotic resistance within biofilms. Here, we use experimental evolution to compare selection of resistance mutations in planktonic and biofilm Escherichia coli populations exposed to clinically relevant cycles of lethal treatment with the aminoglycoside amikacin. Consistently, mutations in sbmA, encoding an inner membrane peptide transporter, and fusA, encoding the essential elongation factor G, are rapidly selected in biofilms, but not in planktonic cells. This is due to a combination of enhanced mutation rate, increased adhesion capacity and protective biofilm-associated tolerance. These results show that the biofilm environment favors rapid evolution of resistance and provide new insights into the dynamic evolution of antibiotic resistance in biofilms.

Probiotic disruption of quorum sensing reduces virulence and increases cefoxitin sensitivity in methicillin-resistant Staphylococcus aureus.

Therapies which target quorum sensing (QS) systems that regulate virulence in methicillin-resistant Staphylococcus aureus (MRSA) are a promising alternative to antibiotics. QS systems play a crucial in the regulation of MRSA antibiotic resistance, exotoxin production, antioxidant protection and immune cell evasion, and are therefore attractive therapeutic targets to reduce the virulence of a pathogen. In the present work the the effects of bioactive peptides isolated from two strains of lactic acid bacteria were tested against antibiotic resistance, carotenoid production, resistance to oxidative killing and biofilm structure in two clinical MRSA isolates. The results obtained from fractional-inhibitory concentration assays with bulk and semi-purified bioactive molecules showed a significant synergistic effect increasing cefoxitin mediated killing of MRSA. This was coupled to a six-fold decrease of the major membrane pigment staphyloxanthin, and a 99% increase in susceptibility to oxidative stress mediated killing. Real-time quantitative PCR analysis of the QS-genes agrA and luxS, showed differential expression between MRSA strains, and a significant downregulation of the hemolysin gene hla. Light microscopy and scanning electron microscopy revealed alteration in biofilm formation and clustering behavior. These results demonstrate that bioactive metabolites may be effectively applied in tandem with beta-lactam antibiotics to sensitize MRSA to cefoxitin. Moreover, these results shown that several key QS-controlled virulence mechanisms are diminished by probiotic metabolites.

Synergistic antibacterial and anti-biofilm mechanisms of ultrasound combined with citral nanoemulsion against Staphylococcus aureus 29213.

This study investigated the antibacterial and antibiofilm mechanism of ultrasound (US) combined with citral nanoemulsion (CLNE) against Staphylococcus aureus and mature biofilm. Combined treatments resulted in greater reductions in bacterial numbers compared to ultrasound or CLNE treatments alone. Confocal laser scanning microscopy (CLSM), flow cytometry (FCM), protein nucleic acid leakage, and N-phenyl-l-naphthylamine (NPN) uptake analysis showed that the combined treatment disrupted cell membrane integrity and permeability. Reactive oxygen species (ROS) and malondialdehyde (MDA) assays indicated that US+CLNE exacerbated cellular oxidative stress and membrane lipid peroxidation. Field emission scanning electron microscopy (FESEM) revealed that the synergistic processing of ultrasound and CLNE resulted in cell rupture and collapse. In addition, US+CLNE showed a more pronounced removal effect than both alone in the biofilm on the stainless steel sheet. US+CLNE reduced biomass, the number of viable cells in the biofilm, cell viability and EPS polysaccharide contents. The results of CLSM also showed that US+CLNE disrupted the structure of the biofilm. This research elucidates the synergistic antibacterial and anti-biofilm mechanism of ultrasound combined citral nanoemulsion, which provides a safe and efficient sterilization method for the food industry.

Induction of Periodontitis via a Combination of Ligature and Lipopolysaccharide Injection in a Rat Model.

Periodontitis (PD) is a highly prevalent, chronic immune-inflammatory disease of the periodontium, that results in a loss of gingival soft tissue, periodontal ligament, cementum, and alveolar bone. In this study, a simple method of PD induction in rats is described. We provide detailed instructions for placement of the ligature model around the first maxillary molars (M1) and a combination of injections of lipopolysaccharide (LPS), derived from Porphyromonas gingivalis at the mesio-palatal side of the M1. The induction of periodontitis was maintained for 14 days, promoting the accumulation of bacteria biofilm and inflammation. To validate the animal model, IL-1ß, a key inflammatory mediator, was determined by an immunoassay in the gingival crevicular fluid (GCF), and alveolar bone loss was calculated using cone beam computed tomography (CBCT). This technique was effective in promoting gingiva recession, alveolar bone loss, and an increase in IL-1ß levels in the GCF at the end of the experimental procedure after 14 days. This method was effective in inducing PD, thus being able to be used in studies on disease progression mechanisms and future possible treatments.

Antimicrobial and cleaning effects of ultrasonic-mediated plasma-loaded microbubbles on Enterococcus faecalis biofilm: an in vitro study.

BACKGROUND: Enterococcus faecalis (E. faecalis) is the most frequently isolated bacteria from teeth with root canal treatment failure. This study aims to evaluate the disinfection effect of ultrasonic-mediated cold plasma-loaded microbubbles (PMBs) on 7d E. faecalis biofilm, the mechanical safety and the mechanisms. METHODS: The PMBs were fabricated by a modified emulsification process and the key reactive species, nitric oxide (NO) and hydrogen peroxide (H2O2) were evaluated. The 7d E. faecalis biofilm on human tooth disk was constructed and divided into the following groups: PBS, 2.5%NaOCl, 2%CHX, and different concentrations of PMBs (108 mL-1, 107 mL-1). The disinfection effects and elimination effects were verified with confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Microhardness and roughness change of dentin after PMBs treatment were verified respectively. RESULTS: The concentration of NO and H2O2 in PMBs increased by 39.99% and 50.97% after ultrasound treatment (p < 0.05) respectively. The CLSM and SEM results indicate that PMBs with ultrasound treatment could remove the bacteria and biofilm components effectively, especially those living in dentin tubules. The 2.5% NaOCl presented an excellent effect against biofilm on dishes, but the elimination effect on dentin tubules is limited. The 2% CHX group exhibits significant disinfection effect. The biosafety tests indicated that there is no significant changes on microhardness and roughness after PMBs with ultrasound treatment (p > 0.05). CONCLUSION: PMBs combined with ultrasound treatment exhibited significant disinfection effect and biofilm removal effect, the mechanical safety is acceptable.

Antimicrobial and Antibiofilm Activity of Copper-Based Metallic Compounds Against Bacteria Related with Healthcare-Associated Infections.

Health care-associated infections (HAIs) contribute to a significant rate of morbidity, mortality, and financial burden on health systems. These infections are caused by multidrug-resistant bacteria that produce biofilm as the main virulence factor. This study aimed to evaluate the effect of the copper-based metallic compounds [Cu(phen)(pz)NO2]Cl (I), [Cu(bpy)(pz)(NO2)]Cl (II), and [Cu(phen)(INA)NO2]Cl (III), where phen = phenanthroline, bpy = bipyridine, pz = pyrazinamide, and INA = isonicotinic acid, against planktonic cells and biofilms formation of Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli. The susceptibility of the microorganisms was evaluated by minimum inhibitory concentration (MIC), minimum bacterial concentration (MBC), and time-kill curve assay on planktonic cells. The biofilm formation was evaluated by biomass quantification through staining with crystal violet (CV), colony-forming units (CFUs) quantification, and biofilm metabolic activity determination by XTT assay. The compounds showed bacteriostatic and bactericidal activity on all microorganisms analyzed. Regarding the antibiofilm activity, all metallic compounds were able to reduce significantly the biofilm biomass, colony-forming units, and the metabolic activity of remaining cells, varying the efficient concentration according to the strain analyzed. Interestingly, compounds (I), (II) and (III) did not exhibit DNA degradation activity even with up to 100 µM of these metal complexes. On the other hand, complexes (I) and (III) showed a remarkable capacity to cleave DNA upon addition of glutathione, a reducing agent (CuII/CuI) that leads to reactive oxygen species (ROS) formation. The results presented in this study showed promising antimicrobial and antibiofilm effects.

CsgI (YccT) Is a Novel Inhibitor of Curli Fimbriae Formation in Escherichia coli Preventing CsgA Polymerization and Curli Gene Expression.

Curli fimbriae are amyloids-found in bacteria (Escherichia coli)-that are involved in solid-surface adhesion and bacterial aggregation during biofilm formation. The curli protein CsgA is coded by a csgBAC operon gene, and the transcription factor CsgD is essential to induce its curli protein expression. However, the complete mechanism underlying curli fimbriae formation requires elucidation. Herein, we noted that curli fimbriae formation was inhibited by yccT-i.e., a gene that encodes a periplasmic protein of unknown function regulated by CsgD. Furthermore, curli fimbriae formation was strongly repressed by CsgD overexpression caused by a multicopy plasmid in BW25113-the non-cellulose-producing strain. YccT deficiency prevented these CsgD effects. YccT overexpression led to intracellular YccT accumulation and reduced CsgA expression. These effects were addressed by deleting the N-terminal signal peptide of YccT. Localization, gene expression, and phenotypic analyses revealed that YccT-dependent inhibition of curli fimbriae formation and curli protein expression was mediated by the two-component regulatory system EnvZ/OmpR. Purified YccT inhibited CsgA polymerization; however, no intracytoplasmic interaction between YccT and CsgA was detected. Thus, YccT-renamed CsgI (curli synthesis inhibitor)-is a novel inhibitor of curli fimbriae formation and has a dual role as an OmpR phosphorylation modulator and CsgA polymerization inhibitor.

Antimicrobial and Antibiofilm Activities of Carvacrol, Amoxicillin and Salicylhydroxamic Acid Alone and in Combination vs. Helicobacter pylori: Towards a New Multi-Targeted Therapy.

The World Health Organization has indicated Helicobacter pylori as a high-priority pathogen whose infections urgently require an update of the antibacterial treatments pipeline. Recently, bacterial ureases and carbonic anhydrases (CAs) were found to represent valuable pharmacological targets to inhibit bacterial growth. Hence, we explored the underexploited possibility of developing a multiple-targeted anti-H. pylori therapy by assessing the antimicrobial and antibiofilm activities of a CA inhibitor, carvacrol (CAR), amoxicillin (AMX) and a urease inhibitor (SHA), alone and in combination. Minimal Inhibitory (MIC) and Minimal Bactericidal (MBC) Concentrations of their different combinations were evaluated by checkerboard assay and three different methods were employed to assess their capability to eradicate H. pylori biofilm. Through Transmission Electron Microscopy (TEM) analysis, the mechanism of action of the three compounds alone and together was determined. Interestingly, most combinations were found to strongly inhibit H. pylori growth, resulting in an additive FIC index for both CAR-AMX and CAR-SHA associations, while an indifferent value was recorded for the AMX-SHA association. Greater antimicrobial and antibiofilm efficacy of the combinations CAR-AMX, SHA-AMX and CAR-SHA against H. pylori were found with respect to the same compounds used alone, thereby representing an innovative and promising strategy to counteract H. pylori infections.