Genomic Characterization of Colistin-Resistant Isolates from the King Fahad Medical City, Kingdom of Saudi Arabia

Background: Whole-genome sequencing is one of the best ways to investigate resistance mechanisms of clinical isolates as well as to detect and identify circulating multi-drug-resistant (MDR) clones or sub-clones in a given hospital setting. Methods: Here, we sequenced 37 isolates of Acinetobacter baumannii, 10 Klebsiella pneumoniae, and 5 Pseudomonas aeruginosa collected from the biobank of the hospital setting of the King Fahad Medical City. Complete phenotypic analyses were performed, including MALDI-TOF identification and antibiotic susceptibility testing. After the genome assembly of raw data, exhaustive genomic analysis was conducted including full resistome determination, genomic SNP (gSNP) analysis, and comparative genomics. Results: Almost all isolates were highly resistant to all tested antibiotics, including carbapenems and colistin. Resistome analysis revealed many antibiotic resistance genes, including those with resistance to β-lactams, aminoglycosides, macrolides, tetracyclines, sulfamids, quinolones, and phenicols. In A. baumannii isolates, the endemic carbapenemase blaOXA-23 gene was detected in 36 of the 37 isolates. Non-synonymous mutations in pmrB were detected in almost all of the isolates and likely mediated colistin resistance. Interestingly, while classical analyses, such as MLST, revealed the predominance of an ST2 clone in A. baumannii isolates, the genomic analysis revealed the presence of five circulating sub-clones and identified several isolate transmissions between patients. In the 10 K. pneumoniae isolates, several resistance genes were identified, and the observed carbapenem resistance was likely mediated by overexpression of the detected extended-spectrum-β-lactamase (ESBL) genes associated with low membrane permeability as few carbapenemase genes were detected with just blaOXA-48 in three isolates. Colistin resistance was mediated either by non-synonymous mutations in the MgrB regulator, PmrA, PmrB, and PhoQ proteins or the presence of the MCR-1 protein. Here, gSNP analysis also revealed the existence of bacterial clones and cases of isolate transmissions between patients. The five analyzed P. aeruginosa isolates were highly resistant to all tested antibiotics, including carbapenems mediated by loss or truncated OprD porin, and colistin resistance was associated with mutations in the genes encoding the PmrA, PmrB, or PhoQ proteins. Conclusion: We demonstrate here the usefulness of whole-genome sequencing to exhaustively investigate the dissemination of MDR isolates at the sub-clone level. Thus, we suggest implementing such an approach to monitor the emergence and spread of new clones or sub-clones, which classical molecular analyses cannot detect. Moreover, we recommend increasing the surveillance of the endemic and problematic colistin resistance mcr-1 gene to avoid extensive dissemination

Patrimonial management of old antimicrobial agents by screening against modern multi-drug resistants bacteria

L’émergence des bactéries résistantes aux béta-lactamines et aux carbapénèmes, a abouti à la réintroduction de la colistine comme agent de dernier recours pour traiter les infections dues à ces germes. Cependant, les résistances chromosomique et plus récemment plasmidique à la colistine ont apparu. Ce problème de bactéries multi-résistantes a par la suite déclenché la publication d’articles alarmants sur les dangers de ces germes. Pour répondre à la dramatisation médiatique liée à ce problème, mon projet de thèse vise à proposer des stratégies thérapeutiques pour traiter les infections dues aux bactéries multi-résistantes. Dans un premier temps, nous avons testé l’activité d’un large panel comprenant des anciens antibiotiques contre les bactéries résistantes aux carbapénèmes et d’autres résistantes à la colistine. Plusieurs familles d’antibiotiques ont été efficaces contre ces 2 types de bactéries résistantes.Dans un deuxième temps, nous avons évalué l’activité d’antibiotiques combinés en vue de détecter une synergie d’action. Deux combinaisons synergiques ont été retenues : colistine + sulfadiazine et colistine + acide fusidique. Ces associations d’antibiotiques ont démontré un effet bactéricide sur une collection de bactéries Gram négatives résistantes à la colistine, et ceci indépendamment du mécanisme de résistance.The emergence of beta-lactam and carbapenem resistant bacteria, resulted in the reintroduction of colistin as an agent of last resort to treat infections caused by these bacteria. However, chromosomal resistances and more recently plasmidic to colistin appeared. This problem of multidrug-resistant bacteria subsequently triggered the publication of alarming articles on the dangers of these germs. To answer the media dramatization related to this problem, my thesis project aims to propose therapeutic strategies to treat infections due to multiresistant bacteria.Initially, we tested the activity of a large panel including old antibiotics against carbapenem resistant bacteria and others resistant to colistin. Several families of antibiotics have been effective against these two types of resistant bacteria.In a second step, we evaluated the activity of combined antibiotics in order to detect a synergistic action. Two synergistic combinations were retained: colistin + sulfadiazine and colistin + fusidic acid. These combinations of antibiotics have shown a bactericidal effect on a collection of Gram-negative colistin-resistant bacteria, independent of the resistance mechanism

Effect of diethylhexyl phthalate on proliferation of ED1-testicular macrophages in rats

AbstractBis(2-ethylhexyl) phthalate (DEHP) is a commonly used plasticizer in many toys, consumer products and medical devices. The aim of the present immunohistochemical work was to study the effect of DEHP on macrophage proliferation in the testis of pubertal rats. Twenty-day old rats (Rattus norvegicus) were gavaged with DEHP (200mg/kg) or the corn oil vehicle (control) for 10, 20, and 30days. Using the specific antibody ED1 we have determined the number of macrophages (per mm2 in paraffin sections) after the DEHP treatment. The treatment of animals with DEHP significantly increased the macrophage numbers in the testis compared to control. This is true for all three durations of treatment. The macrophages were quantified according to the location in the testis: (1) peritubular, (2) Leydig cell vicinity, (3) perivascular. The data showed that the highest number of macrophages was of peritubular location. To conclude, the significant increase of the macrophage number in the testis of DEHP-treated rats can be interpreted as an expression of the inflammatory and/or apoptotic processes

Histological, ultrastructural, and biochemical studies on the expected role of thyme oil on ameliorating renal toxicity induced by doxorubicin in female albino rats

Doxorubicin is antibiotics anticancer drug which proved to be useful in treatment of cancer, but it has bad effect on healthy body organs. Thymus vulgaris (thyme) cultivated around the world for cosmetic, cooking, and medical uses. So, the present work aims to investigate the possible ameliorating effect of thyme oil on renal toxicity induced by doxorubicin. Adult female albino rats (Rattus norvegicus) were randomly divided into four groups (5 rats/group): control group, thyme group received orally 0.5 mL thyme oil/kg body weight once/week for 6 consecutive weeks, doxorubicin group received intraperitoneally 2 mg doxorubicin/kg body weight once/week for 6 consecutive weeks, and doxorubicin + thyme group received both doxorubicin and thyme oil once/week for 6 consecutive weeks. A significant rising in urea and creatinine level in the sera of rats injected with doxorubicin was recorded. Moreover, their kidney sections showed degenerated, vacuolated and little hemorrhage glomeruli, abnormal renal tubular cells, aggregation of mononuclear leucocytes and hyaline cast. Ultrastructure examination showed degenerated glomerular endothelial, untypical filtrating barrier, degenerated podocyte, mesangial cell, pathological proximal and distal tubular cell with damage microvilli. Treating rats with thyme oil after doxorubicin decreases the elevated levels of urea and creatinine. The glomeruli, Podocyte, intraglomerular mesangial cells and glomerular filtration barrier approximately restored its typical structure, the winding in Bouma’s space is disappeared. Most tubular cells have normal nuclei and cytoplasm. Thyme oil can improve renal functions, histological and ultrastructural changes induced by doxorubicin. So, it can be used to decrease its side effect on kidney. Keywords: Doxorubicin, Histology, Ultrastructure, Thyme oil, urea, creatinine.</jats:p