Salivary microRNA 155, 146a/b and 203: A pilot study for potentially non-invasive diagnostic biomarkers of periodontitis and diabetes mellitus

Dysregulated expression of MicroRNAs (miRNAs) plays substantial role in the initiation and progression of both diabetes and periodontitis. The aim of the present study was to validate four miRNAs in saliva as potential predictive biomarkers of periodontal disease among patients with and without diabetes mellitus (DM). MiRNAs were extracted from the saliva of 24 adult subjects with DM and 29 healthy controls. Each group was subdivided into periodontally healthy or having periodontitis. In silico analysis identified 4 miRNAs (miRNA 155, 146 a/b and 203) as immune modulators. The expression of miRNAs-146a/b, 155, and 203 was tested using quantitative PCR. The expression levels in the study groups were compared to explore the effect of diabetes on periodontal status and vice versa. In our cohort, the four miRNAs expression were higher in patients with periodontitis and/or diabetes. miRNA-155 was the most reliable predictors of periodontitis among non-diabetics with an optimum cut-off value of < 8.97 with accuracy = 82.6%. MiRNA 146a, on the other hand, was the only reliable predictor of periodontitis among subjects with diabetes with optimum cut-off value of ≥11.04 with accuracy = 86.1%. The results of the present study concluded that MiRNA-146a and miRNA155 in saliva provide reliable, non-invasive, diagnostic and prognostic biomarkers that can be used to monitor periodontal health status among diabetic and non-diabetic patients

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-Bienayme 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 mu g/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.This work was supported by the generous grants from the IsDB-Transformers Fund and the Research Funding Department, University of Sharjah, UAE (CoV19-0306)

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

Epidemiology and antimicrobial resistance trends of Acinetobacter species in the United Arab Emirates: a retrospective analysis of 12 years of national AMR surveillance data

Introduction: Acinetobacter spp., in particular A. baumannii, are opportunistic pathogens linked to nosocomial pneumonia (particularly ventilator-associated pneumonia), central-line catheter-associated blood stream infections, meningitis, urinary tract infections, surgical-site infections, and other types of wound infections. A. baumannii is able to acquire or upregulate various resistance determinants, making it frequently multidrug-resistant, and contributing to increased mortality and morbidity. Data on the epidemiology, levels, and trends of antimicrobial resistance of Acinetobacter spp. in clinical settings is scarce in the Gulf Cooperation Council (GCC) and Middle East and North Africa (MENA) regions. Methods: A retrospective 12-year analysis of 17,564 non-duplicate diagnostic Acinetobacter spp. isolates from the United Arab Emirates (UAE) was conducted. Data was generated at 317 surveillance sites by routine patient care during 2010-2021, collected by trained personnel and reported by participating surveillance sites to the UAE National AMR Surveillance program. Data analysis was conducted with WHONET. Results: Species belonging to the A. calcoaceticus-baumannii complex were mostly reported (86.7%). They were most commonly isolated from urine (32.9%), sputum (29.0%), and soft tissue (25.1%). Resistance trends to antibiotics from different classes during the surveillance period showed a decreasing trend. Specifically, there was a significant decrease in resistance to imipenem, meropenem, and amikacin. Resistance was lowest among Acinetobacter species to both colistin and tigecycline. The percentages of multidrug-resistant (MDR) and possibly extensively drug-resistant (XDR) isolates was reduced by almost half between the beginning of the study in 2010 and its culmination in 2021. Carbapenem-resistant Acinetobacter spp. (CRAB) was associated with a higher mortality (RR: 5.7), a higher admission to ICU (RR 3.3), and an increased length of stay (LOS; 13 excess inpatient days per CRAB case), as compared to Carbapenem-susceptible Acinetobacter spp. Conclusion: Carbapenem-resistant Acinetobacter spp. are associated with poorer clinical outcomes, and higher associated costs, as compared to carbapenem-susceptible Acinetobacter spp. A decreasing trend of MDR Acinetobacter spp., as well as resistance to all antibiotic classes under surveillance was observed during 2010 to 2021. Further studies are needed to explore the reasons and underlying factors leading to this remarkable decrease of resistance over time

The Relation between Periodontopathogenic Bacterial Levels and Resistin in the Saliva of Obese Type 2 Diabetic Patients

This study aims to investigate the relation between resistin and periodontopathogenic bacterial levels in the saliva of obese adults compared to healthy control and to examine whether salivary resistin can serve as a biomarker of type 2 diabetes in obese patients. A total of 78 saliva samples were collected from patients attending to the University Dental Hospital, Sharjah, UAE. The patients were divided into three equal groups: obese diabetics, obese nondiabetics, and nonobese nondiabetic control. Salivary resistin was measured using ELISA. The levels of bacterial species associated with periodontitis (Treponema denticola, Porphyromonas gingivalis, Tannerella forsythia, and Actinobacillus actinomycetemcomitans) and gingivitis (Fusobacterium spp.) were measured using real-time PCR. Both salivary resistin and periodontopathogenic bacteria including Fusobacterium spp., P. gingivalis, and T. forsythia were detected in significantly higher quantities in the obese patients (diabetics and nondiabetics) than nonobese nondiabetic control. Resistin concentrations were significantly correlated with BMI; however, its level was not correlated with the blood glucose. In this study, high salivary resistin was associated with obesity, which is a major predisposing factor for type 2 diabetes and also a risk factor for oral diseases. The high levels of salivary periodontopathogenic bacteria could upregulate the local release of salivary resistin in obese people

Dietary Fat Effect on the Gut Microbiome, and Its Role in the Modulation of Gastrointestinal Disorders in Children with Autism Spectrum Disorder

Children with autism spectrum disorder (ASD) report a higher frequency and severity of gastrointestinal disorders (GID) than typically developing (TD) children. GID-associated discomfort increases feelings of anxiety and frustration, contributing to the severity of ASD. Emerging evidence supports the biological intersection of neurodevelopment and microbiome, indicating the integral contribution of GM in the development and function of the nervous system, and mental health, and disease balance. Dysbiotic GM could be a contributing factor in the pathogenesis of GID in children with ASD. High-fat diets may modulate GM through accelerated growth of bile-tolerant bacteria, altered bacterial ratios, and reduced bacterial diversity, which may increase the risk of GID. Notably, saturated fatty acids are considered to have a pronounced effect on the increase of bile-tolerant bacteria and reduction in microbial diversity. Additionally, omega-3 exerts a favorable impact on GM and gut health due to its anti-inflammatory properties. Despite inconsistencies in the data elaborated in the review, the dietary fat composition, as part of an overall dietary intervention, plays a role in modulating GID, specifically in ASD, due to the altered microbiome profile. This review emphasizes the need to conduct future experimental studies investigating the effect of diets with varying fatty acid compositions on GID-specific microbiome profiles in children with ASD

Molecular Analysis of Antibiotic Resistance Determinants and Plasmids in Malaysian Isolates of Multidrug Resistant Klebsiella pneumoniae.

Infections caused by multidrug resistant Klebsiella pneumoniae have been increasingly reported in many parts of the world. A total of 93 Malaysian multidrug resistant K. pneumoniae isolated from patients attending to University of Malaya Medical Center, Kuala Lumpur, Malaysia from 2010-2012 were investigated for antibiotic resistance determinants including extended-spectrum beta-lactamases (ESBLs), aminoglycoside and trimethoprim/sulfamethoxazole resistance genes and plasmid replicons. CTX-M-15 (91.3%) was the predominant ESBL gene detected in this study. aacC2 gene (67.7%) was the most common gene detected in aminoglycoside-resistant isolates. Trimethoprim/sulfamethoxazole resistance (90.3%) was attributed to the presence of sul1 (53.8%) and dfrA (59.1%) genes in the isolates. Multiple plasmid replicons (1-4) were detected in 95.7% of the isolates. FIIK was the dominant replicon detected together with 13 other types of plasmid replicons. Conjugative plasmids (1-3 plasmids of ~3-100 kb) were obtained from 27 of 43 K. pneumoniae isolates. An ESBL gene (either CTX-M-15, CTX-M-3 or SHV-12) was detected from each transconjugant. Co-detection with at least one of other antibiotic resistance determinants [sul1, dfrA, aacC2, aac(6')-Ib, aac(6')-Ib-cr and qnrB] was noted in most conjugative plasmids. The transconjugants were resistant to multiple antibiotics including β-lactams, gentamicin and cotrimoxazole, but not ciprofloxacin. This is the first study describing the characterization of plasmids circulating in Malaysian multidrug resistant K. pneumoniae isolates. The results of this study suggest the diffusion of highly diverse plasmids with multiple antibiotic resistance determinants among the Malaysian isolates. Effective infection control measures and antibiotic stewardship programs should be adopted to limit the spread of the multidrug resistant bacteria in healthcare settings

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

Antimicrobial resistance, with the production of extended-spectrum β-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

Molecular Analysis of Ciprofloxacin Resistance Mechanisms in Malaysian ESBL-Producing Klebsiella pneumoniae Isolates and Development of Mismatch Amplification Mutation Assays (MAMA) for Rapid Detection of gyrA and parC Mutations

Ninety-three Malaysian extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae isolates were investigated for ciprofloxacin resistance. Two mismatch amplification mutation (MAMA) assays were developed and used to facilitate rapid detection of gyrA and parC mutations. The isolates were also screened for plasmid-mediated quinolone resistance (PMQR) genes including aac(6′)-Ib-cr, qepA, and qnr. Ciprofloxacin resistance (MICs 4–≥32 μg/mL) was noted in 34 (37%) isolates, of which 33 isolates had multiple mutations either in gyrA alone (n=1) or in both gyrA and parC regions (n=32). aac(6′)-Ib-cr was the most common PMQR gene detected in this study (n=61), followed by qnrB and qnrS (n=55 and 1, resp.). Low-level ciprofloxacin resistance (MICs 1-2 μg/mL) was noted in 40 (43%) isolates carrying qnrB accompanied by either aac(6′)-Ib-cr (n=34) or a single gyrA 83 mutation (n=6). Ciprofloxacin resistance was significantly associated with the presence of multiple mutations in gyrA and parC regions. While the isolates harbouring gyrA and/or parC alteration were distributed into 11 PFGE clusters, no specific clusters were associated with isolates carrying PMQR genes. The high prevalence of ciprofloxacin resistance amongst the Malaysian ESBL-producing K. pneumoniae isolates suggests the need for more effective infection control measures to limit the spread of these resistant organisms in the hospital