Carbapenem- and colistin-resistant Enterobacterales in intensive care unit patients in Mediterranean countries, 2019.

Introduction: The colonization of patients by carbapenemase-producing Enterobacterales (CPE) has been associated with heightened mortality, especially in vulnerable individuals within intensive care units (ICUs). Our study aimed to comprehensively assess CPE prevalence among ICU patients across the Mediterranean region pre-COVID-19, conducting a multicenter prevalence study in the first quarter of 2019. Methods: We collected clinical data and rectal or fecal samples from 256 ICU patients for CPE testing. Additionally, we performed whole-genome sequencing on 40 representative CPE strains to document their molecular characteristics. Results: Among the 256 patients, CPE was detected in 73 samples (28.5%), with prevalence varying from 3.3 to 69.0% across participating centers. We observed 13 colistin-resistant CPE strains, affecting three ICUs. Genetic analysis revealed highly diverse E. coli and K. pneumoniae strains, predominantly from international high-risk clones. Notably, blaOXA-48 and blaNDM-1 were the most prevalent carbapenemase genes. Molecular typing uncovered potential patient clusters in six centers. Significantly, longer hospital stays were associated with increased CPE carriage (p < 0.001). Nine centers across Morocco, Tunisia, Egypt, and Lebanon voluntarily participated. Discussion: Our study provides CPE prevalence in Mediterranean ICUs and reaffirms established CPE presence in this setting but also provides updates on the molecular diversity of CPE strains. These findings highlight the imperative of reinforcing infection control measures in the participating ICUs to curtail escalated mortality rates, and of strictly applying isolation measures around patients originating from the Mediterranean region when transferred to other healthcare institutions.

High Mobility Group Box 1 Gene Polymorphism and Serum High Mobility Group Box 1, Interleukin 1 Beta, and Alpha-Klotho Crosstalk in Severe COVID-19 Patients.

AIM: To evaluate the serum levels of HMGB1, IL1ß, and α-klotho in COVID-19 patients with different disease severity, investigate their association with clinicopathological parameters, and to assess HMGB1 rs1045411 polymorphism and its relation with clinical severity. METHODS: 120 COVID-19 patients (89 critically ill, 15 severe, and 16 moderately severe) along with 80 healthy control were enrolled.The serum levels of HMGB1,IL1ß, and α-klotho were determined by ELISA. The HMGB1 rs1045411 polymorphism was detected by RT- PCR. RESULTS: The serum levels of HMGB1, IL1ß, and α-klotho were significantly higher in critically ill COVID-19 patients compared to other groups. The HMGB1rs1045411 polymorphism revealed a significant decrease in the percentage of T/T genotypes in COVID-19 patients compared to controls. The (ROC) analysis showed moderate diagnostic potential for serum HMGB1, IL1ß, and α-klotho. CONCLUSION: The serum HMGB1, IL1ß, and α-klotho may be severity markers and promising therapeutic targets for COVID-19 patients.

Serum soluble toll-like receptor 4 and risk for clinical severity in COVID-19 patients.

Toll-like receptor 4 (TLR4) signaling mediates sustained systemic inflammation in(COVID)-19 patients. We aimed to assess the serum levels of sTLR4 and sCD14 as negative regulators of Toll like receptor signaling and their association with laboratory markers and clinical severity in covid 19 patients. Ninety-eight patients with COVID-19 (70 severe and 28 non-severe) were enrolled in the study. Serum sCD14 andsTLR4were determined by ELISA. A significant increase in serum sTLR4 and sCD14 levels was detected in severe compared to non severe COVID19 patients.Receiver operating characteristic curve (ROC) analysis revealed significant diagnostic potential of serum sTLR4 and sCD14 in covid19 patients.We conclude that Serum sTLR4 and sCD14 may be promising clinical severity markers for COVID19 patients.

Effect of Colistin, Fosfomycin and Meropenem/Vaborbactam on Carbapenem-Resistant Enterobacterales in Egypt: A Cross-Sectional Study.

Purpose: The increasing multi-drug carbapenem resistance among Enterobacterales are a severe health problem limiting therapeutic options and worsen the prognosis. This study characterizes carbapenemase genes and integrons among uropathogenic carbapenem resistant Enterobacterales (CRE) isolates recovered from Mansoura University Hospitals and evaluates the effect of colistin, fosfomycin and meropenem-vaborbactam on these isolates. Patients and Methods: A total of 200 Enterobacterales isolates were collected from patients with urinary tract infections. Antimicrobial susceptibility testing was performed by the disc diffusion method. Colistin susceptibility was tested using the broth microdilution method and fosfomycin and meropenem/vaborbactam susceptibility were tested by MIC Test Strips. Carbapenem resistant isolates were screened for carbapenemase activity phenotypically using the modified carbapenem inactivation method and EDTA-modified carbapenem inactivation method and genotypically by multiplex PCR. Integrons class 1 and 2 and fosA gene were assayed by PCR. Data were statistically analyzed using the Statistical Package for Social Sciences (SPSS) version 16. The Chi-square or Fisher's exact test was used to compare groups, as appropriate. Results: Ninety-two Enterobacterales isolates were resistant to meropenem (46%); 52 E. coli and 40 K. pneumoniae strains. All CRE isolates were multi-drug resistant (MDR). Sensitivity of CRE isolates to colistin, fosfomycin and meropenem/vaborbactam were 67.4%, 82.6% and 58.7%, respectively. Carbapenemase genes were detected by multiplex PCR in 69.6% of CRE isolates (Carbapenemase producing Enterobacterales (CPE) mainly blaNDM (37%). CPE isolates were significantly more resistant to meropenem/vaborbactam than non-CPE isolates; 51.6% vs 17.8%, respectively (P = 0.003) especially blaNDM carrying isolates (70.6%). Class 1 integrons and fosA gene were detected in 91.3% and 11.9% of CRE isolates, respectively. Conclusion: This study revealed that about half of the uropathogenic Enterobacterales isolates were MDR CRE. Carbapenemase gene blaNDM was the main gene among CRE isolates. Meropenem/vaborbactam sensitivity was significantly higher on non-CPE than CPE isolates and limited by the predominance of blaNDM .

Characterization of E. coli Phylogroups Causing Catheter-Associated Urinary Tract Infection.

PURPOSE: Characterization of different uropathogenic E. coli (UPEC) phylogroups is crucial to understand pathogenesis of urinary tract infection (UTI). The objective of our study was to evaluate the antibiotic resistance pattern, biofilm formation and pathogenicity islands (PAIs) of UPEC phylogroups isolated from catheter-associated UTI (CAUTI) compared to community UTI (Com-UTI). PATIENTS AND METHODS: This study included 90 UPEC strains recovered from CAUTI and Com-UTI. Antimicrobial susceptibility was tested by the Kirby-Bauer method and extended spectrum beta-lactamase (ESBL) production was confirmed using the combined disk. The biofilm formation was tested using the microtiter plate assay. Main E. coli phylogroups (A, B1, B2 and D) were detected by multiplex PCR and 2 multiplex PCR detected the 8 PAIs. RESULTS: Antibiotic resistance of UPEC strains showed a similar high resistance in CAUTI and Com-UTI. Isolates from CAUTI significantly produced biofilm higher than Com-UTI strains (68.9% vs 44.4%). In CAUTI and Com-UTI isolates, phylogroup A was the commonest (53.3% vs 48.9%, respectively). PAI IV536 was the most common in the strains from CAUTI (71.1%) and Com-UTI (73.3%). No significant relationship was detected between the studied characters and different phylogroups except the significant resistance to cefotaxime, ceftazidime and aztreonam among phylogroups from CAUTI isolates. CONCLUSION: Increased antibiotic resistance and ESBLs were detected in UPEC strains from CAUTI and Com-UTI. The strains from CAUTI significantly produced biofilm higher than Com-UTI strains. Phylogroup A was the predominate phylogroup and PAI IV536 was the most prevalent marker in all phylogroups from both types of UTI.

Virulence Factors Of Carbapenem-Resistant Pseudomonas aeruginosa In Hospital-Acquired Infections In Mansoura, Egypt.

PURPOSE: The problem of carbapenem-resistant Pseudomonas aeruginosa in health-care settings is growing worse. This study was conducted to investigate the rate of carbapenemase genes, antibiotic resistance, and virulence factors in carbapenem-resistant P. aeruginosa associated with hospital-acquired infections. PATIENTS AND METHODS: Isolates of P. aeruginosa were collected from patients with hospital-acquired infections at Mansoura University Hospital in Mansoura. Carbapenem susceptibility was done by broth dilution. The presence of carbapenemase genes and quorum-sensing genes was assessed by PCR. Production of protease, pyocyanin, twitching motility, hemolytic activity and biofilm formation was evaluated. RESULTS: Out of 80 P. aeruginosa isolates, 34 (42.5%) were resistant to carbapenem. Among carbapenem-resistant P. aeruginosa isolates, 21 (61.8%) were carbapenemase producers. The most prevalent gene detected was blaVIM. The frequency of protease, pyocyanin, twitching motility, hemolytic activity and biofilm formation was 76.2%, 58.8%, 83.8%, 93.8% and 77.5%, respectively. Biofilm formation was significantly associated with carbapenem-resistant P. aeruginosa. On the other hand, pyocyanin production was significantly lower in carbapenem-resistant isolates. No correlation existed between carbapenem resistance and any other studied virulence factors or quorum-sensing genes. CONCLUSION: Association of carbapenem-resistant P. aeruginosa with other antibiotic resistance or the presence of virulence factors in hospital-acquired infection may represent a warning that enhances the need for a stringent surveillance program.

Hypervirulent Klebsiella pneumoniae as a hospital-acquired pathogen in the intensive care unit in Mansoura, Egypt.

INTRODUCTION: Hypervirulent Klebsiella pneumoniae (hvKP) are variants of K. pneumoniae that come up worldwide. hvKP is known in community-acquired infections but little is known about its role in hospital-acquired (HA) infections. The aim of this study was to evaluate the frequency of hvKP among HA K. pneumoniae infections in the intensive care unit (ICU) and to compare virulence and antibiotic susceptibility between hvKP and classical K. pneumoniae (cKP). METHODS: String test, biofilm formation, serum bactericidal assay, capsular polysaccharide genes (K1, K2, K5, K20, K54, K57), virulence genes: rmpA, rmpA2, iucA, iroB and antimicrobial susceptibility were assessed in HA K. pneumoniae strains isolated from the ICU in Mansoura, Egypt. RESULTS: Probable hvKP represented 4 out of 65 (6.2%) K. pneumoniae. K1 and K2 genes were present in 2 and 1 isolate respectively in probable hvKP. rmpA genes were significantly associated with hvKP; at the same time biofilm production and serum resistance were not significantly associated with the hypervirulent group. There was no significant difference between hvKP and cKP strains in terms of resistance pattern. CONCLUSION: hvKP in critically ill patients from the ICU may form a new threat especially in the presence of antibiotic resistance. Although the validity of the string test in detecting metastatic Klebsiella is questionable, it is a simple and easy test that can be done in any laboratory indicating the presence of this organism. Serotypes and genomic background may provide helpful and confirmatory tools to diagnose hvKP.

High level aminoglycoside resistant enterococci in hospital-acquired urinary tract infections in Mansoura, Egypt.

INTRODUCTION: Resistance to different antimicrobial agents is increasing in enterococci and effective treatment represents a major health concern. The aim of this study was to determine the antimicrobial resistance patterns and the frequency of high level aminoglycoside resistance (HLAR) among enterococci. METHODS: A total of 80 enterococcal isolates, (73 Enterococcus faecalis, 7 Enterococcus faecium) were collected from patients with hospital acquired urinary tract infections (UTI) at Mansoura University hospitals in Egypt. Antimicrobial susceptibility testing was performed via the disc diffusion method. PCR was used for identification of species and detection of aminoglycoside-modifying enzymes genes (AME). RESULTS: All enterococcal isolates were sensitive to vancomycin and linezolid. Fifty-three isolates exhibited HLAR. Our results show that HLAR was mediated by the presence of multiple AMEs genes. The aac(6')-Ie-aph(2')-Ia gene was associated with aph(3')-IIIa and ant(6)-Ia gene in 69% of HLAR isolates. CONCLUSION: This study showed that enterococci isolated from hospital acquired UTI were resistant to multiple antibiotics. Furthermore, the frequency of high level gentamicin resistance (HLGR) was higher than high level of streptomycin resistance (HLSR). The most common AME genes were aph(3')-IIIa and ant(6)-Ia followed by aac(6')-Ie-aph(2')-Ia.

Serum and aqueous humor concentrations of interleukin-27 in diabetic retinopathy patients.

PURPOSE: Interleukin (IL)-27 has been reported to possess anti- and proinflammatory properties in several immune related-disorders, but its role in diabetic retinopathy is still elusive. Here, we aimed to (i) evaluate IL-27 concentrations in serum and aqueous humor of diabetic patients with or without retinopathy and (ii) test whether IL-27 is correlated with some risk factors of diabetic retinopathy. METHODS: The study comprised 60 diabetic patients with and without retinopathy along with 20 healthy controls. Serum and aqueous humor concentrations of IL-27 were assessed by ELISA. RESULTS: The mean of IL-27 concentration in aqueous humor in patients with diabetic retinopathy (6.7 ± 2.7 ng/L) was significantly elevated in comparison with either diabetic patients without retinopathy (4.6 ± 0.5 ng/L) or healthy control group (4.1 ± 0.8 ng/L). Besides, IL-27 concentration in aqueous humor was positively correlated with serum glucose, lipid profile and glycated hemoglobin (HbA1c). CONCLUSIONS: Based on this study, IL-27 is implicated in the pathogenesis of diabetic retinopathy and positively correlates with the disorder progression.

The impact of silver nanoparticles integration on biofilm formation and mechanical properties of glass ionomer cement.

OBJECTIVES: To study the effect of silver nanoparticles incorporation to glass ionomer cement (GIC) on the Staphylococcus aureus biofilm in terms of bacterial growth and evaluate the incorporating effect on hardness and compressive strength. METHODS: Silver nanopowder was added in concentration 0, 1, 3, and 5 wt% to the conventional powder of GIC Fuji IX GP and then the powder is added to the liquid and mixed together with the recommended Powder/liquid ratio of 3.6:1 g. One hundred and twenty disc and cylindrical-shaped specimens were prepared using teflon molds. The specimens were put in tissue culture plate wells contained S. aureus in brain-heart infusion broth. The plate was incubated at 37°C for 24 h. Specimens were then washed, fixed, dehydrated, and air dried. The spatial distribution of biofilm was examined via scanning electron microscope. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were also evaluated. After setting, the specimens were stored in distilled water for 24 h before testing for microhardness and compressive strength. RESULTS: Scanning electron photomicrographs of biofilm formed on the control GIC, showed a consistent biofilm with a thick sheet of cells, whereas those formed were less dense at 3 wt% and below the detection limit at 5 wt% silver nanoparticles. MIC and MBC of S. aureus were 25 and 50 µg/mL, respectively. The microhardness and compressive strength values of tested groups showed a nonsignificant decrease from the control group, P = .58 and .82, respectively. CONCLUSION: Incorporation of silver nanoparticles with GIC can limit S. aureus biofilm formation with an insignificant effect on mechanical properties and noticeable influence on its coloration, which restrict its usage in areas where esthetic is not of major concern. CLINICAL SIGNIFICANCE: As the modification of GIC with silver nanoparticles improved the antibiofilm properties without altering its mechanical properties, it could be used as a restoration of root carious lesion mainly in nonesthetic areas, a base under composite restorations in deep posterior cavities and as a core material in caries susceptible patients.

Vancomycin heteroresistance in coagulase negative Staphylococcus blood stream infections from patients of intensive care units in Mansoura University Hospitals, Egypt.

BACKGROUND: Vancomycin heteroresistance in coagulase negative Staphylococci (CoNS) is a recent health concern especially in serious infections like bloodstream infections as it may lead to failure of therapy. Little information is available about the prevalence vancomycin heteroresistance in CoNS causing bloodstream infections in intensive care units (ICUs) patients of Mansoura University Hospitals (MUHs). METHODS: This prospective study enrolled 743 blood samples collected from ICUs patients presented with clinical manifestations of bloodstream infections over the period extending from January 2014 to March 2016. Samples were processed, coagulase negative Staphylococci were identified by routine microbiological methods and the absence of coagulase activity. Species were identified by API Staph 32. Oxacillin resistant CoNS were identified by cefoxitin disc diffusion method. Susceptibility testing of isolated CoNS to vancomycin was carried out using vancomycin agar dilution method. Mec A gene detection by PCR was done for oxacillin resistant isolates. Screening for vancomycin heteroresistance was done on brain heart infusion (BHI) agar containing 4 µg/mL vancomycin. Confirmation of vancomycin heteroresistance was carried out by population analysis profile (PAP). RESULTS: A total of 58 isolates were identified as CoNS from patients of clinically suspected bloodstream infections. The identified species were 33 (56.9%) Staphylococcus epidermidis, 12 (20.7%) Staphylococcus capitis, 7 (12.1%) Staphylococcus haemolyticus, and 3 isolates (5.2%) Staphylococcus lugdunesis. Three isolates were unidentified by API Staph 32. Forty-four (75.9%) isolates were oxacillin resistant. Mec A gene was detected in all oxacillin resistant isolates. All isolates had susceptible vancomycin MICs by agar dilution. Nine isolates (15.5%) could grow on BHI agar containing 4 µg/mL vancomycin. These isolates showed heterogeneous profile of resistance to vancomycin by population analysis profile. CONCLUSIONS: Vancomycin heteroresistant CoNS causing bloodstream infections is growing unrecognized health hazard in ICUs patients. These isolates have susceptible vancomycin MICs. Screening methods are recommended and should be considered to improve clinical outcome in these high risk patients.

Polymorphisms of glutathione S-transferase π 1 and toll-like receptors 2 and 9: Association with breast cancer susceptibility.

Polymorphisms in antioxidant enzymes and innate immune receptors have been implicated in the development of various types of cancer. The present study aimed to investigate whether polymorphisms of glutathione S-transferase π 1 (GSTP1) and toll-like receptors (TLRs) 2 and 9 are associated with susceptibility to breast cancer among females. The study was conducted on 72 Egyptian female patients with breast cancer, along with 100 healthy volunteers. Polymorphisms of GSTP1 (codon 105 Ile/Val) and TLR9 rs187084 (1237T/C) genes were assessed by polymerase chain reaction (PCR)-restriction fragment length polymorphism, while the -196 to -174 deletion/insertion (del/ins) polymorphism of TLR2 was detected by PCR. The results indicated a decrease in GSTP1 Val allele frequency in breast cancer patients compared with healthy controls, at rates of 22.9 vs. 32.5%, respectively. In addition, the breast cancer group demonstrated a decreased TLR9 C allele frequency compared with the control group, at rates of 36.1 vs. 51.5%, respectively (P=0.0047). A non-significant difference was detected in the frequency of the TLR2 -196 to -174 del allele in breast cancer patients when compared to normal controls. In conclusion, these results suggested that the GSTP1 Val and TLR9 1237C alleles, but not TLR2 -196 to -174 del, are likely to be associated with breast cancer development among females.

Serum soluble toll-like receptor 2: a novel biomarker for systemic lupus erythematosus disease activity and lupus-related cardiovascular dysfunction.

AIM: To assess the serum levels of soluble toll-like receptor (sTLR2) as an endogenous negative regulator of TLR2 signaling in systemic lupus erythematosus (SLE) patients, to investigate the correlation between sTLR2 and SLE disease activity index (SELDAI), SLE-related cardiovascular risk factors and ventricular dysfunction and to evaluate the effect of different therapeutic regimens on serum sTLR2 levels. METHODS: Ninety-six SLE patients, along with 30 healthy controls, were enrolled in the study. Echocardiography measurements were performed. Serum levels of (sTLR2) were measured using enzyme-linked immunosorbent assay (ELISA). Serum lipid profiles, uric acid and creatinine were also detected. RESULTS: Mean serum levels of sTLR2 in SLE patients was 3.98 ± 4.4 ng/mL, which was significantly decreased as compared with that of the control group (11.3 ± 4.9 ng/mL; P < 0.0001). sTLR2 was negatively correlated with SELDAI, low-density lipoprotein (LDL) and left ventricular diastolic dysfunction. sTLR2 levels were increased in patients receiving hydroxychloroquine, statins and corticosteroids. CONCLUSION: Serum sTLR2 can attenuate disease activity and negatively impact left ventricular diastolic dysfunction and hypercholersterelemia in SLE patients. Statins, corticosteroids and chloroquine increase sTLR2 levels.

Simple molecular diagnostic method for fragile X syndrome in Egyptian patients: pilot study.

BACKGROUND: Poor knowledge about Fragile X syndrome (FXS) may be a major barrier to early diagnosis that could improve quality of life and prognosis especially in the developing countries. AIM: The aim of this study was to evaluate simple and reproducible method for premutation detection in females of fragile X families for the first time in Egypt. SUBJECTS AND METHODS: We have developed a rapid modified polymerase chain reaction (PCR)-based screening tool for expanded Fragile X mental retardation 1 (FMR1) alleles. This method utilizes betaine as additive to facilitate FMR 1 gene amplification. We screened fifty three males, thirty two first-degree females; twenty normal healthy controls in addition to six reference samples. RESULTS: Simple PCR method showed 16 males with abnormal CGG repeats, where 10 of their mothers and four sisters had FMR 1 premutation. Consanguineous marriage was present in 66.6% percent of the studied families. Studying the correlation between genotype and clinical manifestations showed premature ovarian failure in 40% and learning disability in 50% of the studied female carriers. CONCLUSION: FXS has to be ruled out in families with consanguineous parents, before assuming that familial mental retardation is due to autosomal recessive gene defects. Early carrier detection may reduce the number of affected children. In conclusion, more studies are still needed of much larger sample size with known allele sizes in order to guarantee the accuracy of the method used.

Persistent oxidative stress in patients with chronic active hepatitis-C infection after antiviral therapy failure.

BACKGROUND/AIMS: Oxidative stress and hepatocellular pathological changes are common associations with chronic hepatitis C virus (CHC) disease. The aim of this study was to assess serum antioxidant-oxidant (Redox) balance in patients with CHC infection before and after intake of the traditional antiviral therapy (pegylated interferon α-2b and oral ribavirin). PATIENTS AND METHODS: Blood samples from 50 biopsy-proven CHC patients, with no prior anti-viral treatment and persistently elevated serum transaminase levels for 6 months, as well as 15 age- and sex-matched healthy subjects were used for determination of the antioxidants: reduced glutathione (GSH), superoxide dismutase (SOD), α tocopherol and ascorbic acid as well as lipid peroxidation (LPO) index (malondialdehyde [MDA]). The measurements were repeated in the diseased group 25 weeks after pegylated interferon α-2b and ribavirin combination therapy. RESULTS: Serum levels of bilirubin, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) were significantly higher in CHC patients than in the control group (P < 0.05). Pretreatment serum MDA values were significantly higher in patients with CHC infection than the control group (P < 0.001), while serum antioxidant levels were significantly lower (P < 0.001). Responders (10 patients) had lower pretreatment serum levels of MDA than non-responders (35 patients) (P < 0.001). Both groups were comparable for the antioxidant serum levels. There was significant negative correlation between serum MDA and serum SOD, GSH, α tocopherol, and ascorbic acid concentrations in CHC patients. On the other hand, there was no correlation between the studied parameters and serum bilirubin, albumin, ALT, and AST. CONCLUSIONS: Redox imbalance was detected in patients with CHC. Responders had significantly lower levels of MDA than non-responders. Serum MDA may be used as a pretreatment predictor of response to antiviral treatment in patients with CHC.