Morpholinium-based ionic liquids show antimicrobial activity against clinical isolates of Pseudomonas aeruginosa

Pseudomonas aeruginosa is a multi-drug resistant (MDR) pathogen. It is classified by WHO as one of the most life-threatening pathogens causing nosocomial infections. Some of its clinical isolates and their subpopulations show high persistence to many antibiotics that are recommended by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Thus, there is a need for non-traditional classes of antibiotics to fight the increasing threat of MDR P. aeruginosa. Ionic liquids (IL) are one such promising class of novel antibiotics. We selected four strains of P. aeruginosa and studied the growth inhibition and other effects of 12 different ILs. We used the well-characterized P. aeruginosa PAO1 (ATCC 15692) as model strain and compared it to three other isolates from chronic lung infection (LES B58), skin burn infection (UCBPP-PA14) and keratitis infection (39016), respectively. The ILs consisted of either 4,4-didecylmorpholinium [Dec₂Mor]⁺ or 4-decyl-4-ethylmorpholinium [DecEtMor]⁺ cations combined with different anions. We found that the ILs with 4,4-didecylmorpholinium [Dec₂Mor]⁺ cations most effectively inhibited bacterial growth as well as reduced strain fitness and virulence factor production. Our results indicate that these ILs could be used to treat P. aeruginosa infections.     

Enhanced efficacy of the engineered antimicrobial peptide WLBU2 via direct airway delivery in a murine model of Pseudomonas aeruginosa pneumonia

ObjectivesPseudomonas aeruginosa is a common cause of pneumonia in patients with cystic fibrosis with the property to generate multidrug resistance against clinically used antibiotics. Antimicrobial peptides (AMPs) are a diverse group of effector molecules of the innate immune system that protect the host against pathogens. However, the lack of activity in common biological matrices has hampered efforts towards clinical development. In this study, we evaluated the therapeutic potential of the engineered AMP WLBU2 via direct airway delivery in a murine model of P. aeruginosa infection.MethodsThe human AMPs LL37 and WLBU2 were compared for (i) antibiofilm activity using P. aeruginosa on polarized human bronchial epithelial cells, and (ii) efficacy in P. aeruginosa pneumonia in mice using intratracheal instillation of bacteria and AMPs.ResultsWLBU2 (16 μM) prevents biofilm formation by up to 3-log compared with 1-log reduction by LL37. With a single dose of 1 μg (0.05 mg/kg) delivered intratracheally, the initial effect of LL37 was moderate and transitory, as bacterial load and inflammatory cytokines increased at 24 h with observed signs of disease such as lethargy and hypothermia, consistent with moribund state requiring euthanasia. In sharp contrast, WLBU2 reduced bacterial burden (by 2 logs) and bacteria-induced inflammation (leucocytic infiltrates, cytokine and chemokine gene expression) at 6 h and 24 h post-exposure, with no observed signs of disease or host toxicity.ConclusionThese promising results now establish a much lower minimum therapeutic dose of WLBU2 (a net gain of 80-fold) compared with the previously reported 4 mg/kg systemic minimum therapeutic dose, with significant implications for clinical development.     

Epinecidin-1 antimicrobial activity: In vitro membrane lysis and In vivo efficacy against Helicobacter pylori infection in a mouse model

Helicobacter pylori (H. pylori) infection is highly prevalent, and has a strong association with various gastric diseases, including gastritis, digestive ulcers, and cancer. H. pylori strains with resistance to existing antibiotics have emerged in the past two decades. Currently, treatment of H. pylori infection (involving the use of proton pump inhibitors, followed by triple therapy with broad-spectrum antibiotics) is suboptimal, with high failure rates. As such, there is a clear need for new approaches against H. pylori. Here, we report that Epinecidin-1 (Epi-1) shows effective bactericidal activity against H. Pylori in vitro, and modulates H. Pylori-induced host immune responses in a mouse model. Epi-1 exhibited a low minimum inhibitory concentration (MIC) against antibiotic-sensitive and clinical antibiotic-resistant strains. Moreover, Epi-1 treatment caused 1-N-phenylnaphthylamine (NPN)-fluorescent probe uptake, suggesting it induced membrane lysis; transmission electron micrographs revealed that membranes were destabilized by the generation of saddle-splay membrane curvature. Oral administration of Epi-1 (quaque die dose) in a mouse infection model had strong efficacy (p < 0.00152) against H. pylori, as compared with conventional proton pump inhibitor (PPI)-triple therapeutic antibiotics. Epi-1 inhibited infection through in vivo depletion of CD4+-FOXP3+ T Regulatory and Th17 subset populations, and aided in clearance of persistent H. pylori colonization. Flow cytometry and gene expression analysis of mouse splenic and gastric tissue indicated that Epi-1 inhibits IL-10, and thereby affects FOXP3 expression levels and reduces pro-inflammatory cytokine responses. Crucially, high doses of Epi-1 did not exert toxic effects in oral, dermal, and eye irritation models. Collectively, our results suggest that Epi-1 may be a promising, effective, and safe monotherapeutic agent for the treatment of multi-drug resistant H. pylori infection.     

In vitro activity of several antimicrobial peptides against colistin-susceptible and colistin-resistant Acinetobacter baumannii

At present, colistin is among the few antibiotics effective against Acinetobacter baumannii clinical isolates. However, in the last few years, colistin-resistant A. baumannii strains have been isolated. Therefore, antibiotics effective against these usually pan-resistant colistin-resistant A. baumannii strains are required. The main objective of this study was to analyse the activity of 15 peptides against colistin-susceptible and colistin-resistant A. baumannii. The MICs were determined by microdilution. Among these 15 antimicrobial peptides (AMPs), melittin, indolicidin and mastoparan showed good activity against both colistin-susceptible and colistin-resistant A. baumannii. Further studies of mastoparan with time-killing curves showed bactericidal activity at MIC x8 for both colistin-susceptible and colistin-resistant A. baumannii. In conclusion, mastoparan may be a potential alternative for the treatment of colistin-resistant A. baumannii infections.     

The antimicrobial activity of liposomal lauric acids against Propionibacterium acnes

This study evaluated the antimicrobial activity of lauric acid (LA) and its liposomal derivatives against Propionibacterium acnes (P. acnes), the bacterium that promotes inflammatory acne. First, the antimicrobial study of three free fatty acids (lauric acid, palmitic acid and oleic acid) demonstrated that LA gives the strongest bactericidal activity against P. acnes. However, a setback of using LA as a potential treatment for inflammatory acne is its poor water solubility. Then the LA was incorporated into a liposome formulation to aid its delivery to P. acnes. It was demonstrated that the antimicrobial activity of LA was not only well maintained in its liposomal derivatives but also enhanced at low LA concentration. In addition, the antimicrobial activity of LA-loaded liposomes (LipoLA) mainly depended on the LA loading concentration per single liposomes. Further study found that the LipoLA could fuse with the membranes of P. acnes and release the carried LA directly into the bacterial membranes, thereby killing the bacteria effectively. Since LA is a natural compound that is the main acid in coconut oil and also resides in human breast milk and liposomes have been successfully and widely applied as a drug delivery vehicle in the clinic, the LipoLA developed in this work holds great potential of becoming an innate, safe and effective therapeutic medication for acne vulgaris and other P. acnes associated diseases.     

In vitro activity of antimicrobial agents against oxacillin resistant staphylococci with special reference to Staphylococcus haemolyticus

One hundred and sixty seven isolates of staphylococci isolated from the inpatients of a tertiary care referral hospital in South India were speciated and activity of oxacillin, glycopeptides, linezolid and quinupristin/dalfopristin against these isolates was tested by broth microdilution method. Of the 114 coagulase negative staphylococci (CoNS), 49.1 % were S. haemolyticus, isolated predominantly from urine (64.6%), while the rest belonged to 11 other species. More than half the isolates of S. aureus (52.8%) and 68.4% of the CoNS were oxacillin resistant. All the strains were uniformly susceptible to vancomycin, linezolid and quinupristin/dalfopristin; but 25.6% isolates of S. haemolyticus showed reduced susceptibility to teicoplanin (MIC: 8-16 mg/L). Our study demonstrates the high prevalence of oxacillin resistance among hospital isolates of S. aureus and CoNS in India. Vancomycin, along with the newer agents like linezolid and quinupristin/dalfopristin remains the drug of choice for treating multi drug resistant staphylococcal infections.     

Activity of MSI-78, h-Lf1-11 and cecropin B antimicrobial peptides alone and in combination with voriconazole and amphotericin B against clinical isolates of Fusarium solani

Fusarium infections have been associated with high mortality rates due to the lack of definition of an ideal treatment strategy. Antimicrobial peptides (AMPs) have potential antifungal activity. Therefore, investigating the in vitro activity of these molecules alone and in association with conventional antifungals against clinical isolates of Fusarium was the aim of this study. Fusarium solani (n = 10) strains were tested against the AMPs, MSI-78, h-Lf1-11 and cecropin B in accordance with CLSI protocol. Further, a checkerboard assay for its combination with amphotericin B or voriconazole, was carried out. MSI-78, h-Lf1-11 and cecropin B exhibited antifungal activity against F. solani strains tested with MICs ranging from 20 to 320 mg/L. Satisfactory percentage of synergism was demonstrated for all evaluated combinations, ranging from 70 to 100%. The use of AMPs combined with amphotericin B and voriconazole antifungals has great synergistic potential and deserve to be evaluated in murine models of fusariosis.     

Global assessment of the antimicrobial activity of polymyxin B against 54 731 clinical isolates of Gram-negative bacilli: report from the SENTRY antimicrobial surveillance programme (2001–2004)

In total, 54 731 Gram-negative bacilli isolated worldwide between 2001 and 2004 from diverse sites of infection were tested for susceptibility to polymyxin B by the broth reference microdilution method, with interpretation of results according to CLSI (formerly NCCLS) guidelines. Polymyxin B showed excellent potency and spectrum against 8705 Pseudomonas aeruginosa and 2621 Acinetobacter spp. isolates (MIC50, < or = 1 mg/L and MIC90, 2 mg/L for both pathogens). Polymyxin B resistance rates were slightly higher for carbapenem-resistant P. aeruginosa (2.7%) and Acinetobacter spp. (2.8%), or multidrug-resistant (MDR) P. aeruginosa (3.3%) and Acinetobacter spp. (3.2%), when compared with the entire group (1.3% for P. aeruginosa and 2.1% for Acinetobacter spp.). Among P. aeruginosa, polymyxin B resistance rates varied from 2.9% in the Asia-Pacific region to only 1.1% in Europe, Latin America and North America, while polymyxin B resistance rates ranged from 2.7% in Europe to 1.7% in North America and Latin America among Acinetobacter spp. Polymyxin B also demonstrated excellent activity (MIC90, < or = 1 mg/L; > 98% susceptible) against Citrobacter spp., Escherichia coli and Klebsiella spp., but activity was more variable against Enterobacter spp. (MIC50, < or = 1 mg/L; 83.3% susceptible) and Stenotrophomonas maltophilia (MIC50, < or = 1 mg/L; 72.4% susceptible), and was very limited (MIC50, > 8 mg/L) against Burkholderia cepacia (11.8% susceptible), Serratia spp. (5.4% susceptible), indole-positive Proteus spp. (1.3% susceptible) and Proteus mirabilis (0.7% susceptible).     

In vitro activity of ceftaroline and other antimicrobial agents against Gram positive bacterial isolates: Descriptive study from a university hospital

PurposeCeftaroline is a novel antibiotic approved by USA Food and Drug Administration (FDA) for the treatment of acute bacterial skin and skin structure infections (ABSSSI) and community-acquired bacterial pneumonia (CABP). It has potent in vitro activity against Staphylococci, including methicillin-resistant strains, whose incidence is increasing worldwide and they are often difficult to treat. The present study was done to investigate the susceptibility of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus (MR-CoNS) to ceftaroline and other antimicrobial agents in patient samples and to evaluate clinical profile of these patients.MethodsAll consecutive, nonduplicate isolates of MRSA and MR-CoNS recovered from patient samples between June 2020 to December 2020 were included in the study. Species identification was done by Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectromtery (MALDI-TOF-MS) (BioMérieux, France). Antimicrobial sensitivity of ceftaroline and other comparator antimicrobials was done as per the Clinical and Laboratory Standards Institute (CLSI, 2020). Statistical Package for the Social Sciences (IBM-SPSS) software (Version 25) was used for statistical study.ResultsTotal 134 clinical isolates of the study consisted of MR-CoNS (115 isolates; 85.8%) and MRSA (19 isolates; 14.2%).89.5% MRSA isolates were sensitive to ceftaroline. 44.3% and 32.2% MR-CoNS isolates had ceftaroline MIC ≤1 ?μg/ml and MIC ?= ?2–4 ?μg/ml respectively.ConclusionsCeftaroline exhibited potent in vitro activity against both MRSA and MR-CoNS in the study.     

Activity of Novispirin G-10, a novel antimicrobial peptide against Chlamydia trachomatis and vaginosis-associated bacteria

We tested the activity of Novispirin G-10, a novel antimicrobial alpha-helical octadecapeptide structurally related to cathelicidins and other innate immunity peptides, against Chlamydia trachomatis serovars L2, D, and E and three organisms associated with bacterial vaginosis (BV). The peptide's activity against C. trachomatis was measured in 48-h shell vial assays with McCoy cell targets. Exposure to 100 micro g/ml of Novispirin G-10 reduced the infectivity of serovars D and E by 99.4-100% and serovar L2 by 91.7-99.1%. At the same concentration of 100 micro g/ml, Novispirin G-10 rapidly killed >99% of Mobiluncus curtisii, Gardnerella vaginalis, and Prevotella bivia, in standard colony-forming unit (CFU) assays. Given its simple structure and relative lack of cytotoxic and hemolytic activity, Novispirin G-10 may be a useful component of microbicide preparations designed to prevent chlamydial infection and/or remediate the abnormal vaginal flora associated with BV.     

The tissue factor pathway inhibitor 1 of Sciaenops ocellatus possesses antimicrobial activity and is involved in the immune response against bacterial infection

Tissue factor pathway inhibitor 1 (TFPI-1) is a Kunitz-type serine protease inhibitor that regulates the activation of tissue factor-induced coagulation. In teleosts, TFPI-1-like sequences have been found to exist in two species (Danio rerio and Cyprinus carpio); however, the potential function of fish TFPI-1 has not been investigated. In this study, we identified and analyzed a TFPI-1 homologue, SoTFPI-1, from red drum (Sciaenops ocellatus). The deduced amino acid sequence of SoTFPI-1 is 284 residues in length and contains three Kunitz domains, an acidic N-terminus, and a basic C-terminus. SoTFPI-1 shares 49.5% and 46.9% overall sequence identities with the TFPI-1 of D. rerio and C. carpio, respectively. Quantitative real time RT-PCR analysis showed that constitutive SoTFPI-1 expression occurred, in increasing order, in kidney, brain, liver, gill, blood, spleen, muscle, and heart. Bacterial infection and lipopolysaccharide exposure upregulated SoTFPI-1 expression in kidney in time-dependent manners. Recombinant SoTFPI-1 (rSoTFPI-1) purified from Escherichia coli exhibits not only serine protease inhibitor activity but also bactericidal activity in a manner that is independent of any host factors. A synthetic peptide, TO17, corresponding to the C-terminal basic region of SoTFPI-1 also possesses antibacterial effect that is more potent than that of the full-length rSoTFPI-1. Taken together, these results demonstrate that (i) SoTFPI-1 is a biologically active serine protease inhibitor endowed with bactericidal property; (ii) provide the first indication that teleost TFPI-1 is likely to be involved in anti-microbial infection and thus is linked to innate immune defense.     

Antibacterial activity and mechanism of action of Monarda punctata essential oil and its main components against common bacterial pathogens in respiratory tract

The aim of the current research work was to study the chemical composition of the essential oil of Monarda punctata along with evaluating the essential oil and its major components for their antibacterial effects against some frequently encountered respiratory infection causing pathogens. Gas chromatographic mass spectrometric analysis revealed the presence of 13 chemical constituents with thymol (75.2%), p-cymene (6.7%), limonene (5.4), and carvacrol (3.5%) as the major constituents. The oil composition was dominated by the oxygenated monoterpenes. Antibacterial activity of the essential oil and its major constituents (thymol, p-cymene, limonene) was evaluated against Streptococcus pyogenes, methicillin-resistant Staphylococcus aureus (MRSA), Streptococcus pneumoniae, Haemophilus influenzae and Escherichia coli. The study revealed that the essential oil and its constituents exhibited a broad spectrum and variable degree of antibacterial activity against different strains. Among the tested strains, Streptococcus pyogenes, Escherichia coli and Streptococcus pneumoniae were the most susceptible bacterial strain showing lowest MIC and MBC values. Methicillin-resistant Staphylococcus aureus was the most resistant bacterial strain to the essential oil treatment showing relatively higher MIC and MBC values. Scanning electron microscopy revealed that the essential oil induced potent and dose-dependent membrane damage in S. pyogenes and MRSA bacterial strains. The reactive oxygen species generated by the Monarda punctata essential oil were identified using 2’, 7’-dichlorofluorescein diacetate (DCFDA).This study indicated that the Monarda punctata essential oil to a great extent and thymol to a lower extent triggered a substantial increase in the ROS levels in S. pyogenes bacterial cultures which ultimately cause membrane damage as revealed by SEM results.     

Antimicrobial activity of mosquito cecropin peptides against Francisella

Francisella tularensis is the cause of the zoonotic disease tularemia. In Sweden and Scandinavia, epidemiological studies have implicated mosquitoes as a vector. Prior research has demonstrated the presence of Francisella DNA in infected mosquitoes but has not shown definitive transmission of tularemia from a mosquito to a mammalian host. We hypothesized that antimicrobial peptides, an important component of the innate immune system of higher organisms, may play a role in mosquito host-defense to Francisella. We established that Francisella sp. are susceptible to two cecropin antimicrobial peptides derived from the mosquito Aedes albopictus as well as Culex pipiens. We also demonstrated induced expression of Aedes albopictus antimicrobial peptide genes by Francisella infection C6/36 mosquito cell line. We demonstrate that mosquito antimicrobial peptides act against Francisella by disrupting the cellular membrane of the bacteria. Thus, it is possible that antimicrobial peptides may play a role in the inability of mosquitoes to establish an effective natural transmission of tularemia.