Antimicrobial Evaluation of Two Polycyclic Polyprenylated Acylphloroglucinol Compounds: PPAP23 and PPAP53.

Polycyclic polyprenylated acylphloroglucinols (PPAPs) comprise a large group of compounds of mostly plant origin. The best-known compound is hyperforin from St. John's wort with its antidepressant, antitumor and antimicrobial properties. The chemical synthesis of PPAP variants allows the generation of compounds with improved activity and compatibility. Here, we studied the antimicrobial activity of two synthetic PPAP-derivatives, the water-insoluble PPAP23 and the water-soluble sodium salt PPAP53. In vitro, both compounds exhibited good activity against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium. Both compounds had no adverse effects on Galleria mellonella wax moth larvae. However, they were unable to protect the larvae from infection with S. aureus because components of the larval coelom neutralized the antimicrobial activity; a similar effect was also seen with serum albumin. In silico docking studies with PPAP53 revealed that it binds to the F1 pocket of human serum albumin with a binding energy of -7.5 kcal/mol. In an infection model of septic arthritis, PPAP23 decreased the formation of abscesses and S. aureus load in kidneys; in a mouse skin abscess model, topical treatment with PPAP53 reduced S. aureus counts. Both PPAPs were active against anaerobic Gram-positive gut bacteria such as neurotransmitter-producing Clostridium, Enterococcus or Ruminococcus species. Based on these results, we foresee possible applications in the decolonization of pathogens.

Detection of Vancomycin Resistant Genes in Intrinsically Antibiotic Resistant Bacteria from the Gut Microbiota of Indonesian Individuals.

Background: Antibiotic resistance is a global public health concern that has been exacerbated by the overuse and misuse of antibiotics, leading to the emergence of resistant bacteria. The gut microbiota, often influenced by antibiotic usage, plays a crucial role in overall health. Therefore, this study aimed to investigate the prevalence of antibiotic resistant genes in the gut microbiota of Indonesian coastal and highland populations, as well as to identify vancomycin-resistant bacteria and their resistant genes. Methods: Stool samples were collected from 22 individuals residing in Pacet, Mojokerto, and Kenjeran, Surabaya Indonesia in 2022. The read count of antibiotic resistant genes was analyzed in the collected samples, and the bacterium concentration was counted by plating on the antibiotic-containing agar plate. Vancomycin-resistant strains were further isolated, and the presence of vancomycin-resistant genes was detected using a multiplex polymerase chain reaction (PCR). Results: The antibiotic resistant genes for tetracycline, aminoglycosides, macrolides, beta-lactams, and vancomycin were found in high frequency in all stool samples (100%) of the gut microbiota. Meanwhile, those meant for chloramphenicol and sulfonamides were found in 86% and 16% of the samples, respectively. Notably, vancomycin-resistant genes were found in 16 intrinsically resistant Gram-negative bacterial strains. Among the detected vancomycin-resistant genes, vanG was the most prevalent (27.3%), while vanA was the least prevalent (4.5%). Conclusion: The presence of multiple vancomycin resistance genes in intrinsically resistant Gram-negative bacterial strains demonstrated the importance of the gut microbiota as a reservoir and hub for the horizontal transfer of antibiotic resistant genes.

In vitro and in vivo comparative analysis of antibacterial activity of green-synthesized silver nanoparticles.

This study aims to compare antibacterial effects of green-synthesized silver nanoparticles (AgNPs) with silver nitrate (AgNO3 ). AgNPs were successfully synthesized using Eucalyptus camaldulensis leaf extract as a reducing and stabilizing agent. Minimum inhibitory concentrations (MIC) of AgNPs and AgNO3 against Staphylococcus aureus and Pseudomonas aeruginosa ranged between 4.8 and 6.75 µg mL-1 . Growth curves demonstrated that inhibition of P. aeruginosa occurred right after AgNPs were added and throughout the period of the study (72 h). Antibacterial effects of both AgNPs and AgNO3 could be abrogated by cysteine and 2-mercaptoethanol, thiol-containing compounds. Galleria mellonella model revealed relatively low toxic effects of both AgNPs and AgNO3 . At 20MIC of AgNPs (≈137.8 mg kg-1 ), more than 80% survival of G. mellonella was observed. Unexpectedly, silver-containing agents could not rescue larvae after S. aureus infection. Further ex vivo experiments in the presence of coelomic larval fluid demonstrated the reduction of antibacterial activity of both AgNPs and AgNO3 . It was speculated that anionic molecules present in the coelomic fluid might neutralize the action of Ag ions. Binding of AgNPs or AgNO3 to albumin, a major protein in human blood which transport several endogenous compounds was not detected, indicating that the silver-containing agents could be applied as an antimicrobial agent.

From an Hsp90 - binding protein to a peptide drug.

The Lpl proteins represent a class of lipoproteins that was first described in the opportunistic bacterial pathogen Staphylococcus aureus, where they contribute to pathogenicity by enhancing F-actin levels of host epithelial cells and thereby increasing S. aureus internalization. The model Lpl protein, Lpl1 was shown to interact with the human heat shock proteins Hsp90α and Hsp90ß, suggesting that this interaction may trigger all observed activities. Here we synthesized Lpl1-derived peptides of different lengths and identified two overlapping peptides, namely, L13 and L15, which interacted with Hsp90α. Unlike Lpl1, the two peptides not only decreased F-actin levels and S. aureus internalization in epithelial cells but they also decreased phagocytosis by human CD14+ monocytes. The well-known Hsp90 inhibitor, geldanamycin, showed a similar effect. The peptides not only interacted directly with Hsp90α, but also with the mother protein Lpl1. While L15 and L13 significantly decreased lethality of S. aureus bacteremia in an insect model, geldanamycin did not. In a mouse bacteremia model L15 was found to significantly decreased weight loss and lethality. Although the molecular bases of the L15 effect is still elusive, in vitro data indicate that simultaneous treatment of host immune cells with L15 or L13 and S. aureus significantly increase IL-6 production. L15 and L13 represent not antibiotics but they cause a significant reduction in virulence of multidrug-resistant S. aureus strains in in vivo models. In this capacity, they can be an important drug alone or additive with other agents.

A new approach against Helicobacter pylori using plants and its constituents: A review study.

Helicobacter pylori is a Gram-negative, spiral-shaped bacillus that colonizes 50% of the world population and is considered a class 1 carcinogen by the World Health Organization. This pathogen is the most common cause of infection-related cancers. Apart from cancer, it also causes several gastric and extra gastric diseases. Eradication of H. pylori using antibiotics is a global challenge because of its drug resistance. Alternative treatment options are gaining more attention to tackle drug-resistant H. pylori infections. Several medicinal plants and their isolated compounds have been reported for their antimicrobial activity against H. pylori. The mechanism of action of many of these plant extracts and plant-derived compounds is different from that of conventional antibiotics. Therefore they are shown to be effective against drug-resistant strains of H. pylori. They act by inhibiting bacterial enzymes, adhesions with gastric mucosa, suppression of nuclear factor-κB and by inhibition of oxidative stress. Extracts from Pistacia lentiscus, Brassica oleracea, Glycyrrhiza glabra, Camellia sinensis, Cinnamomum cassia, Allium sativum and Nigella sativa plants and isolated phyto-compounds such as curcumin, resveratrol, quercetin, allicin and ellagic acid demonstrated antimicrobial activity against H. pylori under in vivo conditions. The plant extracts of Zingiber officinale, Glycyrrhiza glabra; and phytochemical allicin and berberine when combined with standard treatment, result in a dramatic increase in H. pylori eradication. In this review, we highlighted the therapeutic efficacy of different plant extracts and isolated phyto compounds against H. pylori infection and described their role in tackling H. pylori resistance to antibiotics.

Identification of the Natural Transformation Genes in Riemerella anatipestifer by Random Transposon Mutagenesis.

In our previous study, it was shown that Riemerella anatipestifer, a Gram-negative bacterium, is naturally competent, but the genes involved in the process of natural transformation remain largely unknown. In this study, a random transposon mutant library was constructed using the R. anatipestifer ATCC11845 strain to screen for the genes involved in natural transformation. Among the 3000 insertion mutants, nine mutants had completely lost the ability of natural transformation, and 14 mutants showed a significant decrease in natural transformation frequency. We found that the genes RA0C_RS04920, RA0C_RS04915, RA0C_RS02645, RA0C_RS04895, RA0C_RS05130, RA0C_RS05105, RA0C_RS09020, and RA0C_RS04870 are essential for the occurrence of natural transformation in R. anatipestifer ATCC11845. In particular, RA0C_RS04895, RA0C_RS05130, RA0C_RS05105, and RA0C_RS04870 were putatively annotated as ComEC, DprA, ComF, and RecA proteins, respectively, in the NCBI database. However, RA0C_RS02645, RA0C_RS04920, RA0C_RS04915, and RA0C_RS09020 were annotated as proteins with unknown function, with no homology to any well-characterized natural transformation machinery proteins. The homologs of these proteins are mainly distributed in the members of Flavobacteriaceae. Taken together, our results suggest that R. anatipestifer encodes a unique natural transformation machinery.

Molecular Basis of Rhodomyrtone Resistance in Staphylococcus aureus.

Rhodomyrtone (Rom) is a plant-derived broad-spectrum antibiotic active against many Gram-positive pathogens. A single point mutation in the regulatory farR gene (farR*) confers resistance to Rom in Staphylococcus aureus (RomR). The mutation in farR* alters the activity of the regulator, FarR*, in such a way that not only its own gene, farR*, but also the divergently transcribed farE gene and genes controlled by the global regulator, agr, are highly upregulated. Here, we show that mainly the upregulation of the fatty acid efflux pump FarE causes the RomR phenotype, as farE deletion in either the parent or the RomR strain (RomR ΔfarE) yielded hypersensitivity to Rom. Comparative lipidome analysis of the supernatant (exolipidomics) and the pellet fraction revealed that the RomR strain excreted about 10 times more phospholipids (PGs) than the parent strain or the ΔfarE mutants. Since the PG content in the supernatant (2,244 ng/optical density [OD]) was more than 100-fold higher than that of fatty acids (FA), we assumed that PG interacts with Rom, thereby abrogating its antimicrobial activity. Indeed, by static and dynamic light scattering (SLS and DLS) and isothermal titration calorimetry (ITC) analyses, we could demonstrate that both PG and Rom were vesicular and reacted with each other in milliseconds to form a 1:1.49 [Rom-PG(32:0), where PG(32:0) is PG with C32:0 lipids] complex. The binding is entropically driven and hence hydrophobic and of low specificity in nature. Our results indicate that the cytoplasmic membrane is the actual target of Rom, which is also in agreement with Rom's induced rapid collapse of the membrane potential and decreased membrane integrity. IMPORTANCE Antibiotic resistance is a growing public health problem, and alternative antibiotics are urgently needed. Rhodomyrtone (Rom), an antimicrobial compound originally isolated from Rhodomyrtus tomentosa, is active against multidrug-resistant Gram-positive pathogens. However, Rom-resistant (RomR) mutants occur with low frequency. In this study, we unraveled the underlying resistance mechanism, which is based on a point mutation in the farR regulator gene, causing overexpression of FarE, which most likely acts as a phospholipid (PG) efflux pump, as large amounts of PG were found in the supernatant and the pellet fraction. We show that PG can bind to Rom, thereby abrogating its antimicrobial activity. The direct interaction of Rom with PG suggests that Rom's actual target is the cytoplasmic membrane. Antibiotics that interact with PG are rare. Since Rom can be chemically synthesized, it serves as a lead compound for synthesis of improved variants.