Bacterial Diversity and Antibiotic Resistance Patterns of Community-Acquired Urinary Tract Infections in Mega Size Clinical Samples of Egyptian Patients: A Cross-Sectional Study.

BACKGROUND:  Community-acquired urinary tract infection (UTI) is one of the most common infectious diseases nowadays. Alarming increased levels of antimicrobial resistance are developing globally which limit treatment options and may lead to life-threatening problems. AIM: Our study aimed to collect surveillance data on non-hospitalized Egyptian UTI cases and to develop strategies against multidrug-resistant pathogens (MDR). According to our knowledge, this is the first study to screen this high number (15,252 urine samples) in a short period (three months), providing valuable data on resistance profiles in non-hospitalized Egyptian UTI patients. METHODS: A total of 15,252 urine samples were collected from different patients. Positive cultures were identified using a semi-quantitative method. Kirby-Bauer's disc diffusion method was used for antibiotic susceptibility testing, the double disc diffusion method was used for extended-spectrum beta-lactamases-producing strains, and the Chi-square test was used for statistical data processing. RESULTS: The results showed 61% positive cultures, females accounted for 67.5%. Infants and elderly patients showed the highest positive cultures (74.4% and 69.2%, respectively). Despite Escherichia coli being the most common uropathogen (47.19%), Klebsiella species(24.42%) were the most MDR and extended-spectrum ß-lactamase (ESBL)-producing organisms. E. coli and Klebsiella spp. displayed increased resistance to cephalosporins (75% and 81%, respectively). In contrast, both organisms displayed high sensitivity to carbapenems. Unlike Klebsiella spp., E. coli was highly sensitive (92%) to first-line treatment (nitrofurantoin) for UTI. Moreover, trimethoprim/sulfamethoxazole showed higher sensitivity rates compared to other nations. CONCLUSION:  Despite Escherichia coli being the most often identified bacteria in our isolates Klebsiella spp. displayed higher resistance to the majority of tested antibiotics. Fortunately, trimethoprim/sulfamethoxazole significantly increased sensitivity, especially against E. coli. However, both species showed high rates of cephalosporin resistance. Moreover, It is important to promote Egypt's national action plan for antimicrobial resistance in collaboration with the World Health Organization, especially in the community to minimize the chance of bacterial resistance in the Egyptian community.

Impact of Vitamin D Supplementation on the Clinical Outcomes and Epigenetic Markers in Patients with Acute Coronary Syndrome.

Vitamin D has recently been found to influence the renin-angiotensin system (RAS); it can reduce the effects of renin-angiotensin system inhibitors (RASI) by decreasing plasma renin. This study examines the effect of vitamin D supplements on cardiac fibrosis markers, echocardiographic parameters, and epigenetic markers in patients with established acute coronary syndrome (ACS). It also looks at the incidence of vitamin D receptor (VDR) gene polymorphisms Apa I (rs7975232), Bsm I (rs1544410), Taq I (rs731236), and Fok I (rs2228570) and its association with the development of secondary major acute cardiovascular events (MACE) and heart failure (HF). A randomized controlled trial in which patients were divided into two groups was performed. Group 1 comprised of 125 ACS patients who received ACS standard therapy alone, while Group 2 consisted of 125 ACS patients who received ACS standard therapy plus vitamin D according to their vitamin D levels. Patients were monitored for 24 months to find subsequent MACE and HF. Vitamin D therapy for ACS patients resulted in a substantial decline in end systolic and end diastolic volumes (p = 0.0075 and 0.002, respectively), procollagen type III N-terminal peptide (PIIINP) and soluble ST2 levels (p = 0.007 and 0.001, respectively), as well as in ejection fraction and vitamin D level (p = 0.0001 and 0.008, respectively). In addition, vitamin D treatment was linked to a significant decline in the levels of noncoding RNA, such as mir361, lncRNA MEG3, and lncRNA Chaer (p = 2.9 × 10-4, 2.2 × 10-6, and 1.2 × 10-5, respectively). Furthermore, patients who suffered MACE had significantly higher levels of the Bsm I CC and Fok I GG genotypes (p = 4.8 × 10-4 and 0.003, respectively), while patients with HF had significantly higher levels of the Taq I AA genotype (p = 4.2 × 10-7). Supplementing ACS patients with vitamin D has been demonstrated to limit cardiac fibrosis and echocardiographic parameters, as well as epigenetic markers. Additionally, MACE and HF among ACS patients may be related to genetic variations among VDR gene polymorphisms.

Shotgun metagenomic analysis of bacterial symbionts associated with "Chromodoris quadricolor" mantle.

Nudibranchs are colorful marine invertebrates having a diverse group of understudied animals. Recently, some nudibranch members have acquired some attention while others still have not. Chromodoris quadricolor is a member of the Red Sea nudibranch, which did not have the chance to get significant attention. Unlike various invertebrates, it lacks a shell suggesting that it must defend itself in other ways. Therefore, in the present study, we were concerned about the mantle-associated bacterial communities. Being essential partners of this dorid nudibranch system, we investigated their taxonomic and functional profiles. We performed a whole metagenomic shotgun approach for the mantle bacterial cells after a differential pelleting procedure. In this procedure, we separated most of the prokaryotic cells from the eukaryotic host cells. Our findings showed that the mantle-body part holds a diverse group of bacterial species relating mainly to Proteobacteria and Tenericutes phyla. There were novel findings regarding the bacterial members associated with the nudibranch mollusks group. Various species were not previously recorded as bacterial symbionts with nudibranchs. Those members were Bathymodiolus brooksi thiotrophic gill symbiont (23.2%), Mycoplasma marinum (7.4%), Mycoplasma todarodis (5%), and Solemya velum gill symbiont (2.6%). The presence of these bacterial species assumed a nutritional role to the host. However, some of these species were present in a high abundance, suggesting their important symbiosis with Chromodoris quadricolor. In addition, exploring the bacterial ability to produce valuable products resulted in the prediction of 2088 biosynthetic gene clusters (BGCs). We identified different gene cluster classes. Polyketide BGC class was the most represented. Others were related to fatty acid BGCs, RiPP, saccharide, terpene, and NRP BGC classes. Prediction of the activity of these gene clusters resulted in, mainly, an antibacterial activity. In addition, different antimicrobial secondary metabolites were also detected. These secondary metabolites are considered key components regulating the bacterial species interactions in their ecosystem. This suggested the significant contribution of these bacterial symbionts to protect the nudibranch host against predators and pathogens. Globally, it is the first detailed study concerned with both the taxonomic diversity and functional potentials of the bacterial symbionts associated with Chromodoris quadricolor mantle.

In Vitro and In Vivo Effect of Amikacin and Imipenem Combinations against Multidrug-Resistant E. coli.

INTRODUCTION: The emergence of multidrug-resistant (MDR) E. coli has developed worldwide; therefore, the use of antibiotic combinations may be an effective strategy to target resistant bacteria and fight life-threatening infections. The current study was performed to evaluate the in vitro and in vivo efficacy of amikacin and imipenem alone and in combination against multidrug-resistant E. coli. Methods: The combination treatment was assessed in vitro using a checkerboard technique and time-killing curve and in vivo using a peritonitis mouse model. In resistant isolates, conventional PCR and quantitative real-time PCR techniques were used to detect the resistant genes of Metallo-ß-lactamase gene Imipenemase (bla-IMP) and aminoglycoside 6'-N-acetyltransferase (aac (6')-Ib). Scanning electron microscopy was used to detect the morphological changes in the resistant isolates after treatment with each drug alone and in combination. In vitro and in vivo studies showed a synergistic effect using the tested antibiotic combinations, showing fractional inhibitory concentration indices (FICIs) of ≤0.5. Regarding the in vivo study, combination therapy indicated a bactericidal effect after 24 h. E. coli isolates harboring the resistant genes Metallo-ß-lactamase gene Imipenemase (bla-IMP) and aminoglycoside 6'-N-acetyltransferase (aac (6')-Ib) represented 80% and 66.7%, respectively, which were mainly isolated from wound infections. The lowest effect on Metallo-ß-lactamase gene Imipenemase (bla-IMP) and aminoglycoside 6'-N-acetyltransferase (aac (6')-Ib) gene expression was shown in the presence of 0.25 × MIC of imipenem and 0.5 × MIC of amikacin. The scanning electron microscopy showed cell shrinkage and disruption in the outer membrane of E. coli in the presence of the antibiotic combination. Amikacin and imipenem combination can be expected to be effective in the treatment and control of serious infections caused by multidrug-resistant (MDR) E. coli and the reduction in bacterial resistance emergence.

Effect of Imipenem and Amikacin Combination against Multi-Drug Resistant Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic nosocomial pathogen associated with high morbidity and mortality rates. Combination of antibiotics has been found to combat multi-drug resistant or extensively drug resistance P. aeruginosa. In this study we investigate the in vitro and in vivo effect of amikacin and imipenem combination against resistant P. aeruginosa. The checkerboard technique and time-killing curve have been performed for in vitro studies showed synergistic effect for combination. A peritonitis mouse model has been used for evaluation of the therapeutic efficacy of this combination which confirmed this synergistic effect. The in vitro and in vivo techniques showed synergistic interaction between tested drugs with fractional inhibitory concentration indices (FICIs) of ≤0.5. Conventional PCR and quantitative real-time PCR techniques were used in molecular detection of bla IMP and aac(6')-Ib as 35.5% and 42.2% of P. aeruginosa harbored bla IMP and aac(6')-Ib respectively. Drug combination viewed statistically significant reduction in bacterial counts (p value < 0.5). The lowest bla IMP and aac(6')-Ib expression was observed after treatment with 0.25 MIC of imipenem + 0.5 MIC of amikacin. Morphological changes in P. aeruginosa isolates were detected by scanning electron microscope (SEM) showing cell shrinkage and disruption in the outer membrane of P. aeruginosa that were more prominent with combination therapy than with monotherapy.

Antifungal activity of silver nanoparticles on Fluconazole resistant Dermatophytes identified by (GACA)4 and isolated from primary school children suffering from Tinea Capitis in Ismailia - Egypt.

Fungal infections caused by dermatophytes recently became more common. Available antifungal drugs are limited because of emergence of resistant strains due to prophylaxis with them, so there is an urgent need for novel antifungals. This study is aimed to detect the antifungal activity of silver nanoparticles (SNPs) on Fluconazole resistant dermatophytes isolated from primary school children clinically suffering from tinea capitis and attending El-Sheikh Zaid Dermatology Center in Ismailia. The study was done on 112 clinical cases. Examination with potassium hydroxide(KOH) of hair samples was done, followed by routine identification using culturing, macroscopical and microscopical examination and biochemical tests, finally molecular identification using Polymerase Chain Reaction (PCR) with (GACA) 4 was done. Fluconazole resistance of these dermatophytes was detected by different methods including agar disc diffusion method and broth microdilution susceptibility testing. Silver nanoparticles susceptibility testing was carried out on these Fluconazole resistant dermatophytes. The Ubiquitin 1 (Ub 1) gene was detected in samples which were Fluconazole resistant but SNPs susceptible. In this study dermatophytes were found only in 70 samples (62.5%). They were belonged to 3 species: Trichophyton violaceum, Microsporum gypseum and Microsporum canis. Fluconazole resistance was found in 58 samples (82.85%). Both M. canis and M. gypseum were resistant to all used concentrations of SNPs, while T. violaceum was susceptible to 50 µg/ml SNPs solution. The Ub1 gene was detected in 1 sample (4.8%). Therefore SNPs can be used for treatment of T. violaceum, while they can't be used for treatment of M. canis or M. gypseum.

Dissemination of metallo-ß-lactamase in Pseudomonas aeruginosa isolates in Egypt: mutation in blaVIM-4.

This study was designed to investigate the prevalence of metallo-ß-lactamase (MBL) in Pseudomonas aeruginosa isolates collected from Suez Canal University Hospital in Ismailia, Egypt. Antibiotic susceptibility testing and phenotypic and genotypic screening for MBLs were performed on 147 isolates of P. aeruginosa. MICs were determined by agar dilution method for carbapenem that was ≥2 µg/mL for meropenem. MBL genes were detected by multiplex and monoplex PCR for P. aeruginosa-harbored plasmids. Mutation profile of sequenced MBL genes was screened using online software Clustal Omega. Out of 147 P. aeruginosa, 39 (26.5%) were carbapenem-resistant isolates and 25 (64%) were confirmed to be positive for MBLs. The susceptibility rate of P. aeruginosa toward polymyxin B and norfloxacin was 99% and 88%, respectively. Identification of collected isolates by API analysis and constructed phylogenetic tree of 16S rRNA showed that the isolates were related to P. aeruginosa species. The frequency of blaGIM-1, blaSIM-1, and blaSPM-1 was 52%, 48%, and 24%, respectively. BlaVIM and blaIMP-like genes were 20% and 4% and the sequences confirm the isolate to be blaVIM-1, blaVIM-2, blaVIM-4, and blaIMP-1. Three mutations were identified in blaVIM-4 gene. Our study emphasizes the high occurrence of multidrug-resistant P. aeruginosa-producing MBL enzymes.

Carbapenem Susceptibility and Multidrug-Resistance in Pseudomonas aeruginosa Isolates in Egypt.

BACKGROUND: Resistant Pseudomonas aeruginosa is a serious concern for antimicrobial therapy, as the common isolates exhibit variable grades of resistance, involving beta-lactamase enzymes, beside native defense mechanisms. OBJECTIVES: The present study was designed to determine the occurrence of Metallo-ß- Lactamases (MBL) and Amp C harboring P. aeruginosa isolates from Suez Canal university hospital in Ismailia, Egypt. METHODS: A total of 147 P. aeruginosa isolates, recovered from 311 patients during a 10-month period, were collected between May 2013 and February 2014; the isolates were collected from urine, wound and sputum. Minimum inhibitory concentration (MIC) determined by agar dilution methods was ≥2 µg/mL for meropenem and imipenem. Identification of P. aeruginosa was confirmed using API 20NE. Metallo-ß- Lactamases and Amp C were detected based on different phenotypic methods. RESULTS: Overall, 26.5% of P. aeruginosa isolates (39/147) were carbapenem resistant isolates. Furthermore, 64.1% (25/39) were MBL producers, these isolates were screened by the combined disc and disc diffusion methods to determine the ability of MBL production. Both MBL and Amp C harbored P. aeruginosa isolates were 28% (7/25). Sixty-four percent of P. aeruginosa isolates were multidrug resistant (MDR) (16/25). The sensitivity toward polymyxin, imipenem, norfloxacin, piperacillin-tazobactam and gentamicin was 99%, 91%, 88%, 82% and 78%, respectively. The resistance rate towards cefotaxime, ceftazidime, cefepime, aztreonam and meropenem was 98.6%, 86%, 71.4%, 34% and 30%, respectively. CONCLUSIONS: Multidrug resistance was significantly associated with MBL production in P. aeruginosa. Early detection of MBL-producing P. aeruginosa and hospital antibiotic policy prescription helps proper antimicrobial therapy and avoidance of dissemination of these multidrug resistance isolates.

In vitro antagonistic activity of Lactobacillus casei against Helicobacter pylori.

Helicobacter pylori is one of the most common causes of chronic infections in humans. Curing H. pylori infection is difficult because of the habitat of the organism below the mucus adherent layer of gastric mucosa. Lactobacilli are known as acid-resistant bacteria and can remain in stomach for a long time than any other organism, we aimed in this study to examine the efficacy of Lactobacillus casei as a probiotic against H. pylori in humans. Particularly, L. casei was opted as it is considered to be one of the widely used probiotics in dairy products. One hundred and seven strains of H. pylori were isolated from dyspeptic patients and were tested for their antibiotic susceptibility to metronidazole (MTZ), clarithromycin (CLR), tetracycline (TET), and amoxicillin (AMX) by the disc diffusion method. The strains were examined for their susceptibility toward L. casei - present in fermented milk products - by well diffusion method. It was found that 74.7% strains were resistant to MTZ; 1.8% to MTZ, TET, and CLR; 3.7% to MTZ and CLR; 4.6% to MTZ and TET; and 0.9% were resistant to MTZ, TET, and AMX. The antibacterial activity of L. casei against H. pylori was determined on all the tested H. pylori isolates including antibiotic resistant strains with different patterns. Our study proposed the use of probiotics for the treatment of H. pylori infection as an effective approach.

In vitro antagonistic activity of Lactobacillus casei against Helicobacter pylori

Helicobacter pylori is one of the most common causes of chronic infections in humans. Curing H. pylori infection is difficult because of the habitat of the organism below the mucus adherent layer of gastric mucosa. Lactobacilli are known as acid-resistant bacteria and can remain in stomach for a long time than any other organism, we aimed in this study to examine the efficacy of Lactobacillus casei as a probiotic against H. pylori in humans. Particularly, L. casei was opted as it is considered to be one of the widely used probiotics in dairy products. One hundred and seven strains of H. pylori were isolated from dyspeptic patients and were tested for their antibiotic susceptibility to metronidazole (MTZ), clarithromycin (CLR), tetracycline (TET), and amoxicillin (AMX) by the disc diffusion method. The strains were examined for their susceptibility toward L. casei - present in fermented milk products - by well diffusion method. It was found that 74.7% strains were resistant to MTZ; 1.8% to MTZ, TET, and CLR; 3.7% to MTZ and CLR; 4.6% to MTZ and TET; and 0.9% were resistant to MTZ, TET, and AMX. The antibacterial activity of L. casei against H. pylori was determined on all the tested H. pylori isolates including antibiotic resistant strains with different patterns. Our study proposed the use of probiotics for the treatment of H. pylori infection as an effective approach.

Macrolide- and tetracycline-resistance determinants of colonizing group B streptococcus in women in Egypt.

Group B streptococcus (GBS) is a commensal bacterium of the human gastrointestinal and genital tracts. It is a leading cause of neonatal sepsis and meningitis, and has also been recognized as an important pathogen in pregnant women and the elderly. We investigated mechanisms of macrolide and tetracycline resistance in GBS colonizing women in Egypt. A total of 100 isolates were screened using standard antibiotic susceptibility tests. A multiplex PCR assay was used to detect macrolide- and tetracycline-resistance determinants. All isolates were uniformly susceptible to penicillin G, ampicillin, cefotaxime, vancomycin and levofloxacin. The resistance rates to erythromycin, clindamycin, azithromycin, tetracycline and chloramphenicol were 17, 14, 16, 98 and 1 %, respectively. Among the erythromycin-resistant isolates, 82.4 % had constitutive macrolide-lincosamide-streptogramin B (cMLSB) resistance, 5.9 % had inducible MLSB (iMLSB) resistance and 11.8 % had M phenotype resistance. Among the cMLSB phenotypes, 64.3 % of isolates harboured the ermB gene and 35.7 % of isolates harboured none of the investigated macrolide-resistance genes. The single strain expressing the iMLSB phenotype possessed the ermA gene. Of the two strains with the M phenotype, only one possessed the mefA/E gene. Conversely, seven macrolide-sensitive strains (MIC <0.03 µg ml(-1)) were ermB positive and one macrolide-sensitive strain (MIC <0.03 µg ml(-1)) harboured mefA/E. Tetracycline resistance was predominantly due to tetM, which was detected alone (83.7 %) or in association with tetL (12.2 %), tetK (1 %) or tetO (1 %). One strain carried tetM associated with both tetL and tetK, and another strain carried tetO alone. The tetO strains were positive for the mefA/E gene, and the tetL and tetK carrier strains harboured the ermB gene. Susceptible strains harbouring but not expressing an antibiotic-resistance gene should be regarded as potentially resistant. These results emphasize the need to monitor the epidemiology of GBS antibiotic resistance in Egypt.

Comparison of scpB gene and cfb gene polymerase chain reaction assays with culture on Islam medium to detect Group B Streptococcus in pregnancy.

PURPOSE: The purpose of the current study was to evaluate two low-costing PCR assays for rapid detection of Group B Streptococcus (GBS) in comparison to a pigment-based culture method. MATERIALS AND METHODS: One-hundred and fifty vaginal swabs were collected from pregnant women at 35-40 weeks of gestation. Vaginal swabs were inoculated in selective enrichment broth medium, and examined using Islam medium, cfb PCR and scpB PCR assays. The demographic data were analysed to identify independent predictors of GBS colonization (age and gravidity), with GBS status as the dependent variable. RESULTS: There was a significant association of age and gravidity with GBS colonization. GBS was detected in 25.3% of isolates by Islam medium, in 30.6% by using the cfb PCR assay and in 30% by using the scpB PCR assay. CONCLUSION: Older pregnant women (≥30 years) and multigravida (>3 pregnancies) are at higher risk of GBS colonization. Both scpB-gene and cfb-gene-based PCR methods are highly sensitive techniques (100% sensitivity) compared to culture method. However, the specificities of the scpB and cfb PCR assays were 93.75 and 92.85%, respectively.

Vaginal carriage and antibiotic susceptibility profile of group B Streptococcus during late pregnancy in Ismailia, Egypt.

Group B Streptococcus (GBS) infection has long been recognized as a frequent cause of morbidity and mortality in newborn infants. The purpose of this study was to determine the colonization rate with GBS and the antibiotic susceptibility profile in pregnant women attending Gynecological clinics in Egypt. One-hundred and fifty vaginal swabs were collected from pregnant women at 35-40 weeks of gestation. In comparison to culture, direct latex agglutination testing revealed 100% sensitivity and 93.75% specificity. Thirty-eight specimens (25.3%) were found to be positive for GBS. Each isolate was tested for susceptibility to penicillin G, ampicillin, cefotaxime, erythromycin, clindamycin and vancomycin. Erythromycin-resistant isolates were further classified by double-disk method. All isolates were susceptible to penicillin G, ampicillin and vancomycin. Resistance to cefotaxime was detected in three isolates (7.89%). Five isolates (13.15%) were resistant to erythromycin and nine isolates (23.68%) were resistant to clindamycin. Four (80%) isolates had constitutive macrolide-lincosamide-Streptogramin(B) resistance (cMLSB(B)) resistance and one (20%) isolate had inducible resistance (iMLS(B)) resistance. GBS colonization was found to be high in our region. Latex agglutination testing and Islam medium are reliable methods to detect GBS in late pregnancy; however, latex agglutination test is rapid and simpler. Penicillin G remains the first choice antibiotic for treatment of GBS infections.