Extreme resistance to the novel siderophore-cephalosporin cefiderocol in an extensively drug-resistant Klebsiella pneumoniae strain causing fatal pneumonia with sepsis: genomic analysis and synergistic combinations for resistance reversal.

Cefiderocol (CFDC) is the first-in-class siderophore-cephalosporin. Klebsiella pneumoniae strain that is extremely resistant to CFDC (MIC: 256 µg/ml) was isolated for the first time in the United Arab Emirates from a patient with pneumonia and sepsis. It belonged to sequence-type 14 (ST14), with a novel core genome ST. Resistance was driven by the co-expression of ß-lactamases (blaNDM-1, blaOXA-232 and blaCTX-M-15) and a mutation in catecholate-siderophore receptor, utilized by CFDC to enter the bacterial cell. Synergistic combinations (ß-lactamase inhibitors, aztreonam plus CFDC) re-sensitized the bacteria to CFDC. Although CFDC resistance is multifactorial, the combination with ß-lactamase inhibitors represents a promising approach in resistance reversal for fighting superbugs.

Discovery of Lactomodulin, a Unique Microbiome-Derived Peptide That Exhibits Dual Anti-Inflammatory and Antimicrobial Activity against Multidrug-Resistant Pathogens.

The human body is a superorganism that harbors trillions of microbes, most of which inhabit the gut. To colonize our bodies, these microbes have evolved strategies to regulate the immune system and maintain intestinal immune homeostasis by secreting chemical mediators. There is much interest in deciphering these chemicals and furthering their development as novel therapeutics. In this work, we present a combined experimental and computational approach to identifying functional immunomodulatory molecules from the gut microbiome. Based on this approach, we report the discovery of lactomodulin, a unique peptide from Lactobacillus rhamnosus that exhibits dual anti-inflammatory and antibiotic activities and minimal cytotoxicity in human cell lines. Lactomodulin reduces several secreted proinflammatory cytokines, including IL-8, IL-6, IL-1ß, and TNF-α. As an antibiotic, lactomodulin is effective against a range of human pathogens, and is most potent against antibiotic-resistant strains such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE). The multifunctional activity of lactomodulin affirms that the microbiome encodes evolved functional molecules with promising therapeutic potential.

Boosting the Antibacterial Activity of Azithromycin on Multidrug-Resistant Escherichia coli by Efflux Pump Inhibition Coupled with Outer Membrane Permeabilization Induced by Phenylalanine-Arginine ß-Naphthylamide.

The global spread of multidrug-resistant (MDR) bacteria increases the demand for the discovery of new antibiotics and adjuvants. Phenylalanine-arginine ß-naphthylamide (PAßN) is an inhibitor of efflux pumps in Gram-negative bacteria, such as the AcrAB-TolC complex in Escherichia coli. We aimed to explore the synergistic effect and mechanism of action of PAßN combined with azithromycin (AZT) on a group of MDR E. coli strains. Antibiotic susceptibility was tested for 56 strains, which were screened for macrolide resistance genes. Then, 29 strains were tested for synergy using the checkerboard assay. PAßN significantly enhanced AZT activity in a dose-dependent manner in strains expressing the mphA gene and encoding macrolide phosphotransferase, but not in strains carrying the ermB gene and encoding macrolide methylase. Early bacterial killing (6 h) was observed in a colistin-resistant strain with the mcr-1 gene, leading to lipid remodeling, which caused outer membrane (OM) permeability defects. Clear OM damage was revealed by transmission electron microscopy in bacteria exposed to high doses of PAßN. Increased OM permeability was also proven by fluorometric assays, confirming the action of PAßN on OM. PAßN maintained its activity as an efflux pump inhibitor at low doses without permeabilizing OM. A non-significant increase in acrA, acrB, and tolC expression in response to prolonged exposure to PAßN was noted in cells treated with PAßN alone or with AZT, as a reflection of bacterial attempts to counteract pump inhibition. Thus, PAßN was found to be effective in potentiating the antibacterial activity of AZT on E. coli through dose-dependent action. This warrants further investigations of its effect combined with other antibiotics on multiple Gram-negative bacterial species. Synergetic combinations will help in the battle against MDR pathogens, adding new tools to the arsenal of existing medications.

Prevalence, virulence and antifungal activity of C. albicans isolated from infected root canals.

BACKGROUND: There is limited data on the prevalence of Candida species in infected root canal systems of human teeth. We attempted to investigate the prevalence, genotype, virulence and the antifungal susceptibility of Candida albicans isolated from infected root canals of patients with primary and post-treatment infections in a UAE population. METHODS: Microbiological samples from 71 subjects with infected root canals were aseptically collected, and cultured on Sabouraud dextrose agar, and C. albicans was identified using multiplex polymerase chain reaction, and the isolates were further subtyped using ABC genotyping system. Their relative virulence was compared using further four archival samples of endodontic origin from another geographical region, and four more salivary isolates, as controls. The virulence attributes compared were biofilm formation, and production of phospholipase and haemolysin, and the susceptibility to nystatin, amphotericin B, ketoconazole, and fluoconazole was also tested. RESULTS: 4 out of 71 samples (5.6%) yielded Candida species. On analysis of variance among the groups, the intracanal isolates, mainly Genotype A, possessed a high degree of phospholipase and haemolysin activity (p < 0.05). The UAE and Finland isolates were stronger biofilm formers and had superior phospholipase production capacity compared with the salivary isolates. All isolates were sensitive to the antifungal chemicals used. The salivary isolates were more sensitive to fluoconazole compared to other groups (p < 0.05). CONCLUSION: The prevalence of C. albicans in infected root canals of patients attending a dental hospital in UAE is low. The strains isolated were good biofilm formers, possessed strong phospholipase and haemolysin activity and were mostly of the genotype A. The fact that the root canal isolates possessed significant hydrolase activity, imply that they are equipped with virulence attributes necessary for survival within a harsh intracanal ecosystem.

Sequencing [4 + 1]-Cycloaddition and Aza-Michael Addition Reactions: A Diastereoselective Cascade for the Rapid Access of Pyrido[2',1':2,3]/Thiazolo[2',3':2,3]imidazo[1,5-a]quinolone Scaffolds as Potential Antibacterial and Anticancer Motifs.

The design and synthesis of a quality compound library containing a small number of skeletally diverse scaffolds, whose members rapidly deliver new chemical probes active against multiple phenotypes, is paramount in drug discovery. In this context, an efficient one-pot strategy for the synthesis of a mini library of sp3-enriched hexahydropyrido[2',1':2,3]imidazo[1,5-a]quinolinium and hexahydrothiazolo[2',3':2,3]imidazo[1,5-a]quinolinium architectures, is described. This new one-pot method features a combination of Sc(OTf)3-catalyzed [4 + 1]-cycloaddition with aza-Michael addition reactions. The cascade results in a rapid and diastereoselective formation of these scaffolds via desymmetrization of the oxidative dearomatization products of phenols. Phenotypic screening of the mini library against multiple drug-resistant bacteria and a panel of cancer cell lines identified potential antibacterial and anticancer lead drug candidates. Further investigation of the anticancer leads, indicated by their activity as tubulin-polymerization inhibitors, represents a promising approach for cancer therapy.

Molecular analyses indicate profuse bacterial diversity in primary and post- treatment endodontic infections within a cohort from the United Arab Emirates-A preliminary study.

OBJECTIVE: Endodontic microbiota appears to undergo evolutionary changes during disease progression from inflammation to necrosis and post-treatment. The aim of this study was to compare microbiome composition and diversity in primary and post-treatment endodontic infections from a cohort of patients from the UAE. DESIGN: Intracanal samples were collected from primarily infected (n = 10) and post-treatment infected (n = 10) root canals of human teeth using sterile paper points. Bacterial DNA was amplified from seven hypervariable regions (V2-V4 and V6-V9) of the 16S rRNA gene, then sequenced using next-generation sequencing technology. The data was analyzed using appropriate bioinformatic tools. RESULTS: Analyses of all the samples revealed eight major bacterial phyla, 112 genera and 260 species. Firmicutes was the most representative phylum in both groups and was significantly more abundant in the post-treatment (54.4%) than in primary (32.2%) infections (p>0.05). A total of 260 operational taxonomic units (OTUs) were identified, of which 126 (48.5%) were shared between the groups, while 83 (31.9%) and 51 (19.6%) disparate species were isolated from primary and post-treatment infections, respectively. A significant difference in beta, but not alpha diversity was noted using several different indices (p< 0.05). Differential abundance analysis indicated that, Prevotella maculosa, Streptococcus constellatus, Novosphigobium sediminicola and Anaerococcus octavius were more abundant in primary infections while Enterrococcus faecalis, Bifidobacterium dentium, Olsenella profusa and Actinomyces dentalis were more abundant in post-treatment infections (p <0.05). CONCLUSION: Significant differences in the microbiome composition and diversity in primary and post-treatment endodontic infections were noted in our UAE cohort. Such compositional differences of microbiota at various stages of infection could be due to both intrinsic and extrinsic factors impacting the root canal ecosystem during disease progression, as well as during their therapeutic management. Identification of the key microbiota in primarily and secondarily infected root canals can guide in the management of these infections.

Human microbiome derived synthetic antimicrobial peptides with activity against Gram-negative, Gram-positive, and antibiotic resistant bacteria.

The prevalence of antibacterial resistance has become one of the major health threats of modern times, requiring the development of novel antibacterials. Antimicrobial peptides are a promising source of antibiotic candidates, mostly requiring further optimization to enhance druggability. In this study, a series of new antimicrobial peptides derived from lactomodulin, a human microbiome natural peptide, was designed, synthesized, and biologically evaluated. Within the most active region of the parent peptide, linear peptide LM6 with the sequence LSKISGGIGPLVIPV-NH2 and its cyclic derivatives LM13a and LM13b showed strong antibacterial activity against Gram-positive bacteria, including resistant strains, and Gram-negative bacteria. The peptides were found to have a rapid onset of bactericidal activity and transmission electron microscopy clearly shows the disintegration of the cell membrane, suggesting a membrane-targeting mode of action.

Oral administration of Manuka honey induces IFNγ-dependent resistance to tumor growth that correlates with beneficial modulation of gut microbiota composition.

Background: To investigate the potential of Manuka honey (MH) as an immunomodulatory agent in colorectal cancer (CRC) and dissect the underlying molecular and cellular mechanisms. Methods: MH was administered orally over a 4 week-period. The effect of MH treatment on microbiota composition was studied using 16S rRNA sequencing of fecal pellets collected before and after treatment. Pretreated mice were implanted with CRC cells and followed for tumor growth. Tumors and lymphoid organs were analyzed by flow cytometry (FACS), immunohistochemistry and qRT-PCR. Efficacy of MH was also assessed in a therapeutic setting, with oral treatment initiated after tumor implantation. We utilized IFNγ-deficient mice to determine the importance of interferon signaling in MH-induced immunomodulation. Results: Pretreatment with MH enhanced anti-tumor responses leading to suppression of tumor growth. Evidence for enhanced tumor immunogenicity included upregulated MHC class-II on intratumoral macrophages, enhanced MHC class-I expression on tumor cells and increased infiltration of effector T cells into the tumor microenvironment. Importantly, oral MH was also effective in retarding tumor growth when given therapeutically. Transcriptomic analysis of tumor tissue highlighted changes in the expression of various chemokines and inflammatory cytokines that drive the observed changes in tumor immunogenicity. The immunomodulatory capacity of MH was abrogated in IFNγ-deficient mice. Finally, bacterial 16S rRNA sequencing demonstrated that oral MH treatment induced unique changes in gut microbiota that may well underlie the IFN-dependent enhancement in tumor immunogenicity. Conclusion: Our findings highlight the immunostimulatory properties of MH and demonstrate its potential utilization in cancer prevention and treatment.

Unraveling the Impact of Gut and Oral Microbiome on Gut Health in Inflammatory Bowel Diseases.

Inflammatory bowel disease (IBD) is a complex disorder characterized by chronic inflammation of the gastrointestinal tract (GIT). IBD mainly includes two distinct diseases, namely Crohn's disease and ulcerative colitis. To date, the precise etiology of these conditions is not fully elucidated. Recent research has shed light on the significant role of the oral and gut microbiome in the development and progression of IBD and its collective influence on gut health. This review aims to investigate the connection between the oral and gut microbiome in the context of IBD, exploring the intricate interplay between these microbial communities and their impact on overall gut health. Recent advances in microbiome research have revealed a compelling link between the oral and gut microbiome, highlighting their pivotal role in maintaining overall health. The oral cavity and GIT are two interconnected ecosystems that harbor complex microbial communities implicated in IBD pathogenesis in several ways. Reduction in diversity and abundance of beneficial bacterial species with the colonization of opportunistic pathogens can induce gut inflammation. Some of these pathogens can arise from oral origin, especially in patients with oral diseases such as periodontitis. It is essential to discern the mechanisms of microbial transmission, the impact of oral health on the gut microbiome, and the potential role of dysbiosis in disease development. By elucidating this relationship, we can enhance our understanding of IBD pathogenesis and identify potential therapeutic avenues for managing the disease. Furthermore, innovative strategies for modulating the oral and gut microbiome can promote health and prevent disease occurrence and progression.

Dysbiosis of the Subgingival Microbiome and Relation to Periodontal Disease in Association with Obesity and Overweight.

Obesity causes gut dysbiosis; nevertheless, little is known about the oral microbiome. We aimed to identify differences in the subgingival microbiota influenced by body weight and periodontal status. Patients (n = 75) recruited at the University Dental Hospital Sharjah, United Arab Emirates, were distributed into three equal groups (healthy weight, overweight, and obese) sub-divided into having either no-mild (NM) or moderate-severe (MS) periodontitis. Subgingival plaques were collected. Microbiota were identified by 16S rRNA sequencing using nanopore technology. Linear discriminant analysis demonstrated significant bacterial biomarkers for body weight and periodontal health. Unique microbiota signatures were identified, with enrichment of periopathogens in patients with MS periodontitis (Aggregatibacter actinomycetemcomitans in obese, Tannerella forsythia and Treponema denticola in overweight, Porphyromonas gingivalis and Fusobacterium nucleatum in healthy weight), thus reflecting differences in the microbiota affected by body weight. Other pathogenic bacteria, such as Salmonella enterica and Klebsiella pneumoniae, were enriched in overweight subjects with NM periodontitis, suggesting an increase in the relative abundance of pathogens even in patients with good periodontal health if they were overweight. Alpha and beta diversities were significantly different among the groups. Dysbiosis of the subgingival microbiota in obese and overweight individuals was associated with increased prevalence and severity of periodontal disease, which was correlated with the body mass index. This study highlights the immense importance of the oral microbiome and the need for lifestyle and dental interventions to resolve oral dysbiosis and restore normal homeostasis.

High efficacy and enhanced synergistic activity of the novel siderophore-cephalosporin cefiderocol against multidrug-resistant and extensively drug-resistant Klebsiella pneumoniae from inpatients attending a single hospital in the United Arab Emirates.

BACKGROUND: Cefiderocol (CFDC) is a novel siderophore-cephalosporin, which usually penetrates the bacteria through the iron-uptake pathways. Data is limited on the factors affecting CFDC activity and methods for overcoming resistance development. Synergistic approaches are needed to tackle antimicrobial resistance. This study aimed to determine CFDC activity on Klebsiella pneumoniae isolates from patients attending a single hospital in the United Arab Emirates (UAE), to explore the effect of ß-lactamases on CFDC activity and to enhance CFDC susceptibility in both iron-depleted and iron-enriched conditions. METHODS: We investigated 238 K. pneumoniae strains from diverse clinical sources. ß-lactamase genes were detected by PCR. Susceptibility to CFDC and 12 comparator antibiotics were tested. Combinations of CFDC with ß-lactamase inhibitors (BLIs) and/or an outer membrane (OM) permeabilizer (polymyxin B nonapeptide) were tested in iron-depleted and iron-enriched conditions. RESULTS: CFDC exhibited efficacy of 97.9%, against multidrug-resistant (MDR), and extensively drug-resistant (XDR) strains, in addition to strains resistant to the last resort drugs such as colistin and tigecycline, including dual carbapenemase-producers (blaNDM and blaOXA-48-like) with MIC ≤ 0.06-8 µg/ml. It was effective in killing strains with single and multiple ß-lactamases; however, it lost activity in iron-enriched conditions. Synergy was achieved with dual combination of CFDC and BLIs, especially avibactam, which caused a significant reduction in MICs even in iron-enriched conditions. A significant reduction was seen with the triple combination including an OM permeabilizer plus avibactam. Killing-kinetic studies proved that the combination therapy caused dose reduction and faster killing by CFDC than the monotherapy. CONCLUSIONS: CFDC was deemed effective against MDR and XDR K. pneumoniae. Synergistic combination of CFDC with BLIs and OM permeabilizers could be effective to treat infections in iron-rich sites, but this should be investigated in vivo.

Inverse Association between the Existence of CRISPR/Cas Systems with Antibiotic Resistance, Extended Spectrum ß-Lactamase and Carbapenemase Production in Multidrug, Extensive Drug and Pandrug-Resistant Klebsiella pneumoniae.

Antimicrobial resistance, with the production of extended-spectrum ß-lactamases (ESBL) and carbapenemases, is common in the opportunistic pathogen, Klebsiella pneumoniae. This organism has a genome that can contain clustered regularly interspaced short palindromic repeats (CRISPRs), which operate as a defense mechanism against external invaders such as plasmids and viruses. This study aims to determine the association of the CRISPR/Cas systems with antibiotic resistance in K. pneumoniae isolates from Iraqi patients. A total of 100 K. pneumoniae isolates were collected and characterized according to their susceptibility to different antimicrobial agents. The CRISPR/Cas systems were detected via PCR. The phenotypic detection of ESBLs and carbapenemases was performed. The production of ESBL was detected in 71% of the isolates. Carbapenem-resistance was detected in 15% of the isolates, while only 14% were susceptible to all antimicrobial agents. Furthermore, the bacteria were classified into multidrug (77%), extensively drug-resistant (11.0%) and pandrug-resistant (4.0%). There was an inverse association between the presence of the CRISPR/Cas systems and antibiotic resistance, as resistance was higher in the absence of the CRISPR/Cas system. Multidrug resistance in ESBL-producing and carbapenem-resistant K. pneumoniae occurred more frequently in strains negative for the CRISPR/Cas system. Thus, we conclude that genes for exogenous antibiotic resistance can be acquired in the absence of the CRISPR/Cas modules that can protect the bacteria against acquiring foreign DNA.

Study of vacuolating cytotoxin A (vacA) genotypes of ulcerogenic and non-ulcerogenic strains of Helicobacter pylori and its association with gastric disease.

Globally, Helicobacter pylori (H. pylori), a stomach pathogen, is present in around 50 % of the population. This bacterial infection produces persistent inflammation, which significantly raises the risk of duodenal, gastric ulcer, and stomach cancer. The goal of this study is to identify the vacA genotypes in H. pylori and analyze how they relate to medical conditions brought on by the bacteria and clarithromycin resistance. PCR was used to describe 115 endoscopic stomach samples from infected patients and identify vacA gene. Of the 115 research participants, H. pylori was found in 81 (70.4 %) of them. Of the isolated cultures, only 38 (69.1 %) were resistant to clarithromycin. VacA was discovered in 55 (67.9 %) of the samples that had H. pylori in them. Patients with gastritis were more likely to have s2m2 strains of infection (66.7 %), while those with gastric and duodenal ulcers were more likely to have s1m1 strains (64.7 %). VacA-positive H. pylori strains (60 % n = 33) were more resistant to clarithromycin versus (19.2 % n = 5) for vacA-negative bacteria. Clarithromycin resistance was significantly linked to vacA s2m2 in H. pylori isolates (75.9 %). According to the study's results, the vacA variants s1m1 and s2m2 have a strong connection with the emergence of H. pylori infections that cause peptic ulcer disease in the population of Iraq. Genetic testing is essential in predicting both the course of treatment and the outcome of H. pylori disease.

Deciphering salivary microbiome signature in Crohn's disease patients with different factors contributing to dysbiosis.

Crohn's disease (CD) is a chronic inflammatory bowel disease. An imbalanced microbiome (dysbiosis) can predispose to many diseases including CD. The role of oral dysbiosis in CD is poorly understood. We aimed to explore microbiome signature and dysbiosis of the salivary microbiome in CD patients, and correlate microbiota changes to the level of inflammation. Saliva samples were collected from healthy controls (HC) and CD patients (n = 40 per group). Salivary microbiome was analyzed by sequencing the entire 16S rRNA gene. Inflammatory biomarkers (C-reactive protein and calprotectin) were measured and correlated with microbiome diversity. Five dominant species were significantly enriched in CD, namely Veillonella dispar, Megasphaera stantonii, Prevotella jejuni, Dolosigranulum pigrum and Lactobacillus backii. Oral health had a significant impact on the microbiome since various significant features were cariogenic as Streptococcus mutans or periopathogenic such as Fusobacterium periodonticum. Furthermore, disease activity, duration and frequency of relapses impacted the oral microbiota. Treatment with monoclonal antibodies led to the emergence of a unique species called Simonsiella muelleri. Combining immunomodulatory agents with monoclonal antibodies significantly increased multiple pathogenic species such as Salmonella enterica, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. Loss of diversity in CD was shown by multiple diversity indices. There was a significant negative correlation between gut inflammatory biomarkers (particularly calprotectin) and α-diversity, suggesting more inflammation associated with diversity loss in CD. Salivary dysbiosis was evident in CD patients, with unique microbiota signatures and perturbed species that can serve as disease biomarkers or potential targets for microbiota modulation. The interplay of various factors collectively contributed to dysbiosis, although each factor probably had a unique effect on the microbiome. The emergence of pathogenic bacteria in the oral cavity of CD patients is alarming since they can disturb gut homeostasis and induce inflammation by swallowing, or hematogenous spread of microbiota, their metabolites, or generated inflammatory mediators.

Antimicrobial activity of nature-inspired molecules against multidrug-resistant bacteria.

Multidrug-resistant bacterial infections present a serious challenge to global health. In addition to the spread of antibiotic resistance, some bacteria can form persister cells which are tolerant to most antibiotics and can lead to treatment failure or relapse. In the present work, we report the discovery of a new class of small molecules with potent antimicrobial activity against Gram-positive bacteria and moderate activity against Gram-negative drug-resistant bacterial pathogens. The lead compound SIMR 2404 had a minimal inhibitory concentration (MIC) of 2 µg/mL against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-intermediate Staphylococcus aureus (VISA). The MIC values against Gram-negative bacteria such as Escherichia coli and Actinobacteria baumannii were between 8-32 µg/mL. Time-kill experiments show that compound SIMR 2404 can rapidly kill tested bacteria. Compound SIMR 2404 was also found to rapidly kill MRSA persisters which display high levels of tolerance to conventional antibiotics. In antibiotic evolution experiments, MRSA quickly developed resistance to ciprofloxacin but failed to develop resistance to compound SIMR 2404 even after 24 serial passages. Compound SIMR 2404 was not toxic to normal human fibroblast at a concentration of 4 µg/mL which is twice the MIC concentration against MRSA. However, at a concentration of 8 µg/mL or higher, it showed cytotoxic activity indicating that it is not ideal as a candidate against Gram-negative bacteria. The acceptable toxicity profile and rapid antibacterial activity against MRSA highlight the potential of these molecules for further studies as anti-MRSA agents.

Discerning the role of polymyxin B nonapeptide in restoring the antibacterial activity of azithromycin against antibiotic-resistant Escherichia coli.

Antimicrobial resistance is a global public health threat. Antibiotic development pipeline has few new drugs; therefore, using antibiotic adjuvants has been envisioned as a successful method to preserve existing medications to fight multidrug-resistant (MDR) pathogens. In this study, we investigated the synergistic effect of a polymyxin derivative known as polymyxin B nonapeptide (PMBN) with azithromycin (AZT). A total of 54 Escherichia coli strains were first characterized for macrolide resistance genes, and susceptibility to different antibiotics, including AZT. A subset of 24 strains was then selected for synergy testing by the checkerboard assay. PMBN was able to re-sensitize the bacteria to AZT, even in strains with high minimum inhibitory concentrations (MIC: 32 to ≥128 µg/ml) for AZT, and in strains resistant to the last resort drugs such as colistin and meropenem. The fractional inhibitory concentration index was lower than 0.5, demonstrating that PMBN and AZT combinations had a synergistic effect. The combinations worked efficiently in strains carrying mphA gene encoding macrolide phosphotransferase which can cause macrolide inactivation. However, the combinations were inactive in strains having an additional ermB gene encoding macrolide methylase which causes ribosomal drug target alteration. Killing kinetics study showed a significant reduction of bacterial growth after 6 h of treatment with complete killing achieved after 24 h. Transmission electron microscopy showed morphological alterations in the bacteria treated with PMBN alone or in combination with AZT, with evidence of damage to the outer membrane. These results suggested that PMBN acted by increasing the permeability of bacterial outer membrane to AZT, which was also evident using a fluorometric assay. Using multiple antimicrobial agents could therefore be a promising strategy in the eradication of MDR bacteria. PMBN is a good candidate for use with other antibiotics to potentiate their activity, but further studies are required in vivo. This will significantly contribute to resolving antimicrobial resistance crisis.

Elucidating the effect of iron acquisition systems in Klebsiella pneumoniae on susceptibility to the novel siderophore-cephalosporin cefiderocol.

BACKGROUND: Cefiderocol (CFDC) is a novel siderophore-cephalosporin, effective against multidrug-resistant Gram-negative bacteria. As it has a siderophore side chain, it can utilize iron acquisition systems for penetration of the bacterial outer membrane. We aimed to elucidate the role of siderophores and iron uptake receptors in defining Klebsiella pneumoniae susceptibility to CFDC. METHODS: Initially, 103 K. pneumoniae strains were characterized for susceptibility to different antibiotics including CFDC. CFDC minimum inhibitory concentrations (MIC) were determined in iron-depleted and iron-enriched conditions. Iron uptake genes including siderophores, their receptors, ferric citrate (fecA) and iron uptake (kfu) receptors were detected by PCR in all the strains. For 10 selected strains, gene expression was tested in iron-depleted media with or without CFDC treatment and compared to expression in iron-enriched conditions. RESULTS: CFDC exhibited 96.1% susceptibility, being superior to all the other antibiotics (MIC50: 0.5 and MIC90: 4 µg/ml). Only three strains (2.9%) were intermediately susceptible and a pandrug resistant strain (0.97%) was resistant to CFDC (MIC: 8 and 256 µg/ml, respectively). The presence of kfu and fecA had a significant impact on CFDC MIC, especially when co-produced, and if coupled with yersiniabactin receptor (fyuA). CFDC MICs were negatively correlated with enterobactin receptor (fepA) expression and positively correlated with expression of kfu and fecA. Thus, fepA was associated with increased susceptibility to CFDC, while kfu and fecA were associated with reduced susceptibility to CFDC. CFDC MICs increased significantly in iron-enriched media, with reduced expression of siderophore receptors, hence, causing less drug uptake. CONCLUSION: Iron acquisition systems have a significant impact on CFDC activity, and their altered expression is a factor leading to reduced susceptibility. Iron concentration is also a major player affecting CFDC susceptibility; therefore, it is essential to explore possible ways to improve the drug activity to facilitate its use to treat infections in iron-rich sites.

Antibacterial Activity of Small Molecules Which Eradicate Methicillin-Resistant Staphylococcus aureus Persisters.

The serious challenge posed by multidrug-resistant bacterial infections with concomitant treatment failure and high mortality rates presents an urgent threat to the global health. We herein report the discovery of a new class of potent antimicrobial compounds that are highly effective against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA). The compounds were efficiently synthesized in one-pot employing a cascade of Groebke-Blackburn-Bienaymé and aza-Michael addition reactions. Phenotypic screening of the pilot library against various bacterial species including methicillin-sensitive and MRSA strains, has identified potent chemotypes with minimal inhibitory concentrations (MIC) of 3.125-6.25 µg/ml. The most potent compounds were fast-acting at eradicating exponentially growing MRSA, with killing achieved after 30 min of exposure to the compounds. They were also able to kill MRSA persister cells which are tolerant to most available medications. Microscopic analysis using fluorescence microscope and atomic force microscope indicate that these compounds lead to disruption of bacterial cell envelopes. Most notably, bacterial resistance toward these compounds was not observed after 20 serial passages in stark contrast to the significant resistance developed rapidly upon exposure to a clinically relevant antibiotic. Furthermore, the compounds did not induce significant hemolysis to human red blood cells. In vivo safety studies revealed a high safety profile of these motifs. These small molecules hold a promise for further studies and development as new antibacterial agents against MRSA infections.

Supragingival microbiome alternations as a consequence of smoking different tobacco types and its relation to dental caries.

This study aimed to assess the effect of smoking different tobacco types on the supragingival microbiome and its relation to dental caries. Forty supragingival plaque samples were collected from smokers of a single tobacco type and non-smokers seeking treatment at the University Dental Hospital Sharjah, UAE. DMFT (decayed, missing and filled teeth) was determined for all participants who were divided into two groups: no-low caries (NC-LC: DMFT = 0-4; n = 18) and moderate-high caries (MC-HC: DMFT = 5-20; n = 22). 16S rRNA gene was sequenced using third-generation sequencing with Nanopore technology. Microbiome composition and diversity were compared. Caries was most common among cigarette smokers. Supragingival microbiota were significantly altered among smokers of different tobacco types. In cigarette smokers, cariogenic bacteria from genus Streptococcus (including S. mutans) were significantly more among subjects with NC-LC, while Lactobacilli (including L. fermentum) were more among subjects with MC-HC. In medwakh smokers, several periodontopathogens were significantly elevated in subjects with NC-LC, while other pathogenic bacteria (as Klebsiella pneumoniae) were more in those with MC-HC. Cigarette and alternative tobacco smoking had a significant impact on the supragingival microbiome. Indeed, further studies are required to unravel the consequences of oral dysbiosis triggered by smoking. This could pave the way for microbiota-based interventional measures for restoring a healthy oral microbiome which could be a promising strategy to prevent dental caries.

The impact of smoking different tobacco types on the subgingival microbiome and periodontal health: a pilot study.

Smoking is a risk factor for periodontal disease, and a cause of oral microbiome dysbiosis. While this has been evaluated for traditional cigarette smoking, there is limited research on the effect of other tobacco types on the oral microbiome. This study investigates subgingival microbiome composition in smokers of different tobacco types and their effect on periodontal health. Subgingival plaques were collected from 40 individuals, including smokers of either cigarettes, medwakh, or shisha, and non-smokers seeking dental treatment at the University Dental Hospital in Sharjah, United Arab Emirates. The entire (~ 1500 bp) 16S rRNA bacterial gene was fully amplified and sequenced using Oxford Nanopore technology. Subjects were compared for the relative abundance and diversity of subgingival microbiota, considering smoking and periodontal condition. The relative abundances of several pathogens were significantly higher among smokers, such as Prevotella denticola and Treponema sp. OMZ 838 in medwakh smokers, Streptococcus mutans and Veillonella dispar in cigarette smokers, Streptococcus sanguinis and Tannerella forsythia in shisha smokers. Subgingival microbiome of smokers was altered even in subjects with no or mild periodontitis, probably making them more prone to severe periodontal diseases. Microbiome profiling can be a useful tool for periodontal risk assessment. Further studies are recommended to investigate the impact of tobacco cessation on periodontal disease progression and oral microbiome.