PCR-based assay for the rapid and precise distinction of Pseudomonas aeruginosa from other Pseudomonas species recovered from burns patients

Background. Pseudomonas aeruginosa is an important lifethreatening nosocomial pathogen which plays a prominent role in wound infections in burns patients. We designed this study to identify the isolates of P. aeruginosa recovered from burns patients at the genus and species levels by means of primers targeting oprI and oprL genes. Methods. During a 5-month period, wound samples were taken from burns patients and plated on MacConkey agar. All suspected colonies were screened for P. aeruginosa by means of a combination of phenotype tests. Specific primers for oprI and oprL genes were then used for the molecular identification of colonies. Results. During the 5-month period, bacterial isolates recovered from burn wound infections were analyzed. Phenotype identification tests identified 171 (34.8) P. aeruginosa isolates. However, molecular techniques that used species-specific primers to detect the amplicon of the oprL gene confirmed the exact identification of P. aeruginosa in only 133 cases; in the other isolates, the use of genus-specific primers detected the amplicon of the oprI gene, which confirmed the identification of fluorescent pseudomonads. Conclusions. This study indicates that molecular detection by means of an assay targeting the oprL gene is a useful technique for the rapid and precise detection of P. aeruginosa in burns patients. In addition to phenotype testing, PCR detection should be carried out in order to promptly ascertain the best aggressive antibiotic therapy for P. aeruginosa infections, thereby significantly improving clinical outcomes

Cluster analysis and genetic characterization of enterobacter cloacae complex from blood cultures

Background: Enterobacter cloacae bacteremia is reported as an important cause of morbidity and mortality. The members of E. cloacae complex are clinically involved in nosocomial infections. Objectives: The purpose of this study was to investigate the prevalence of E. cloacae complex and its members in blood samples and conduct the hsp60 cluster analysis and genotyping of the isolates. Methods: Eight isolates of E. cloacae complex were collected from blood cultures of hospitalized patients during the study period (December 2012 to November 2013). The hsp60 sequencing was done for the genetic classification. Pulsed-field gel electrophoresis (PFGE) was used for genotyping of the isolates. Results: Fifty percent of the isolates belonged to two E. hormaechei subspecies. Three isolates (37.5) clustered within genotype III while only one isolate fitted cluster XIII genotype (12.5). Pulsed-field gel electrophoresis analysis revealed four different pulsotypes. Conclusions: Different E. cloacae complex species and subspecies unequally contribute to the pathogenesis of blood infections and the subspecies of E. hormaechei were found to be most prevalent. Moreover, the common E. cloacae pulsotypes were observed to essentially produce identical hsp60 sequence types, indicating the probable clonality of isolates with identical pulsotypes. © 2018, Author(s)

Spread of efflux pump overexpressing-mediated fluoroquinolone resistance and multidrug resistance in Pseudomonas aeruginosa by using an efflux pump inhibitor

Background: Fluoroquinolone resistance in Pseudomonas aeruginosa may be due to efflux pump overexpression and/or target mutations. We designed this study to investigate the efflux pump mediated fluoroquinolone resistance and check the increasing effectiveness of fluoroquinolones in combination with an efflux pumps inhibitor among P. aeruginosa isolates from burn wounds infections. Materials and Methods: A total of 154 consecutive strains of P. aeruginosa were recovered from separate patients hospitalized in a burn hospital, Tehran, Iran. The isolates first were studied by disk diffusion antibiogram for 11 antibiotics and then minimum inhibitory concentration (MIC) experiments were performed to detect synergy between ciprofloxacin and the efflux pump inhibitor, carbonyl cyanide-m-chlorophenyl hydrazone (CCCP). Then to elucidate the inducing of multi drug resistance due to different efflux pumps activation in Fluoroquinolone resistant isolates, synergy experiments were also performed in random ciprofloxacin resistant isolates which have overexpressed efflux pumps phenotypically, using CCCP and selected antibiotics as markers for Beta-lactams and Aminoglycosides. The isolates were also tested by polymerase chain reaction (PCR) for the presence of the MexA, MexC and MexE, which encode the efflux pumps MexAB-OprM, MexCD-OprJ and MexEF-OprN. Results: Most of the isolates were resistant to 3 or more antibiotics tested. More than half of the ciprofloxacin resistant isolates exhibited synergy between ciprofloxacin and CCCP, indicating the efflux pump activity contributed to the ciprofloxacin resistance. Also increased susceptibility of random ciprofloxacin resistant isolates of P. aeruginosa to other selected antibiotics, in presence of CCCP, implied multidrug extrusion by different active efflux pump in fluoroquinolones resistant strains. All of Ciprofloxacin resistant isolates were positive for MexA, MexC and MexE genes simultaneously. Conclusion: In this burn hospital, where multidrug resistant P. aeruginosa isolates were prevalent, ciprofloxacin resistance and multidrug resistance due to the overexpression of fluoroquinolones mediated efflux pumps has also now emerged. Early recognition of this resistance mechanism should allow the use of alternative antibiotics and use an efflux pumps inhibitor in combination with antibiotic therapy. © 2015 by The Korean Society of Infectious Diseases

Role of efflux pump inhibitor in decreasing antibiotic cross-resistance of Pseudomonas aeruginosa in a burn hospital in Iran

Introduction: Multidrug resistance in Pseudomonas aeruginosa may be due to efflux pump overexpression. This study phenotypically examined the role of efflux pump inhibitors in decreasing antibiotic cross-resistance between beta-lactams, fluoroquinolones, and aminoglycosides in P. aeruginosa isolates from burn patients in Iran. Methodology: A total of 91 phenotypically and genotypically confirmed P. aeruginosa samples were studied. Multidrug cross-resistance was determined using the disk diffusion method and minimum inhibitory concentration (MIC) test. The contribution of efflux pumps was determined by investigating MIC reduction assay to markers of beta-lactams, fluoroquinolones, and aminoglycosides in the absence and presence of an efflux pump inhibitor. All the isolates were also tested by polymerase chain reaction for the presence of mexA, mexC, and mexE efflux genes. Results: Of the isolates, 81 (89) and 83 (91.2) were multidrug resistant according to the disk diffusion and MIC method, respectively. Cross-resistance was observed in 67 (73.6) and 68 (74.7) of isolates according to the disk diffusion and MIC method, respectively. In the presence of the efflux pump inhibitor, twofold or higher MIC reduction to imipenem, cefepime, ciprofloxacin, and gentamicin was observed in 59, 65, 55, and 60 isolates, respectively. Except for two isolates that were negative for mexC, all isolates were positive for mexA, mexC, and mexE genes simultaneously. Conclusion: Efflux pumps could cause different levels of resistance based on their expression in clinical isolates. Early detection of different efflux pumps in P. aeruginosa could allow the use of other antibiotics and efflux pump inhibitors in combination with antibiotic therapy. © 2016 Talebi-Taher et al

Wide distribution of carbapenem resistant Acinetobacter baumannii in burns patients in Iran

Antimicrobial resistance in carbapenem non-susceptible Acinetobacter baumannii (CNSAb) is a major public health concern globally. This study determined the antibiotic resistance and molecular epidemiology of CNSAb isolates from a referral burn center in Tehran, Iran. Sixty-nine CNSAb isolates were tested for susceptibility to antimicrobial agents using the E test methodology. Multiple locus variable number tandem repeat analysis (MLVA), Multilocus sequence typing (MLST) and multiplex PCR were performed. PCR assays tested for ambler classes A, B, and D β-lactamases. Detection of ISAba1, characterization of integrons, and biofilm formation were investigated. Fifty-three (77) isolates revealed XDR phenotypes. High prevalence of blaOXA-23-like (88) and blaPER-1 (54) were detected. ISAba1 was detected upstream of blaADC, blaOXA-23-like and blaOXA51-like genes in, 97, 42, and 26 of isolates, respectively. Thirty-one (45) isolates were assigned to international clone (IC) variants. MLVA identified 56 distinct types with six clusters and 53 singleton genotypes. Forty previously known MLST sequence types forming 5 clonal complexes were identified. The Class 1 integron (class 1 integrons) gene was identified in 84 of the isolates. The most prevalent (33) cassette combination was aacA4-catB8-aadA1. The IC variants were predominant in the A. baumannii lineage with the ability to form strong biofilms. The XDR-CNSAb from burned patients in Iran is resistant to various antimicrobials, including tigecycline. This study shows wide genetic diversity in CNSAb. Integrating the new Iranian A. baumannii IC variants into the epidemiologic clonal and susceptibility profile databases can help effective global control measures against the XDR-CNSAb pandemic. � 2015 Farshadzadeh, Hashemi, Rahimi, Pourakbari, Esmaeili, Haghighi, Majidpour, Shojaa, Rahmani, Gharesi, Aziemzadeh and Bahador

Wide distribution of carbapenem resistant Acinetobacter baumannii in burns patients in Iran

Antimicrobial resistance in carbapenem non-susceptible Acinetobacter baumannii (CNSAb) is a major public health concern globally. This study determined the antibiotic resistance and molecular epidemiology of CNSAb isolates from a referral burn center in Tehran, Iran. Sixty-nine CNSAb isolates were tested for susceptibility to antimicrobial agents using the E test methodology. Multiple locus variable number tandem repeat analysis (MLVA), Multilocus sequence typing (MLST) and multiplex PCR were performed. PCR assays tested for ambler classes A, B, and D β-lactamases. Detection of ISAba1, characterization of integrons, and biofilm formation were investigated. Fifty-three (77) isolates revealed XDR phenotypes. High prevalence of blaOXA-23-like (88) and blaPER-1 (54) were detected. ISAba1 was detected upstream of blaADC, blaOXA-23-like and blaOXA51-like genes in, 97, 42, and 26 of isolates, respectively. Thirty-one (45) isolates were assigned to international clone (IC) variants. MLVA identified 56 distinct types with six clusters and 53 singleton genotypes. Forty previously known MLST sequence types forming 5 clonal complexes were identified. The Class 1 integron (class 1 integrons) gene was identified in 84 of the isolates. The most prevalent (33) cassette combination was aacA4-catB8-aadA1. The IC variants were predominant in the A. baumannii lineage with the ability to form strong biofilms. The XDR-CNSAb from burned patients in Iran is resistant to various antimicrobials, including tigecycline. This study shows wide genetic diversity in CNSAb. Integrating the new Iranian A. baumannii IC variants into the epidemiologic clonal and susceptibility profile databases can help effective global control measures against the XDR-CNSAb pandemic. � 2015 Farshadzadeh, Hashemi, Rahimi, Pourakbari, Esmaeili, Haghighi, Majidpour, Shojaa, Rahmani, Gharesi, Aziemzadeh and Bahador

Balancing exploration and exploitation phases in whale optimization algorithm: an insightful and empirical analysis

Agents of any metaheuristic algorithms are moving in two modes, namely exploration and exploitation. Obtaining robust results in any algorithm is strongly dependent on how to balance between these two modes. Whale optimization algorithm as a robust and well recognized metaheuristic algorithm in the literature, has proposed a novel scheme to achieve this balance. It has also shown superior results on a wide range of applications. Moreover, in the previous chapter, an equitable and fair performance evaluation of the algorithm was provided. However, to this point, only comparison of the final results is considered, which does not explain how these results are obtained. Therefore, this chapter attempts to empirically analyze the WOA algorithm in terms of the local and global search capabilities i.e. the ratio of exploration and exploitation phases. To achieve this objective, the dimension-wise diversity measurement is employed, which, at various stages of the optimization process, statistically evaluates the population's convergence and diversity.Comment: 11 page

Molecular characterization and diagnosis of nosocomial clostridium difficile infection in hospitalized patients

Background: Toxigenic Clostridium difficile is one of the prevalent diarrheagenic pathogens in hospitalized patients. Objectives: The study assessed the ability of three diagnostic methods in identifying C. difficile strains. The genotyping of the isolates was done, as well. Methods: Stool samples were subjected to three different diagnostic methods including direct stool culture, glutamate dehydroge-nase enzyme immunoassay (GDH-EIA), and direct stool PCR for the detection of the tcdA and tcdB genes. The sensitivity and specificity of the tests were evaluated. The genotyping was done by the PFGE method. Results: Of 120 samples, 20 (16) were positive for C. difficile based on PCR, while 15 (12.5) and 12 (10) were positive according to GDH-EIA and direct stool culture. Among patients with C. difficile-associated diseases (CDAD), 11 (61) were more than 65-years-old. The specificity of PCR, GDH-EIA, and direct culture was almost similar and equal to 100, but their sensitivity was 90, 70, and 60, respectively. The positive predictive value (PPV) was lower for GDH-EIA than for the other two methods, and the highest negative predictive value (NPV) was related to the PCR method. The results showed a high similarity between the isolates, and only were three pulsotypes differentiated among the isolates. Conclusions: The specificity and sensitivity of the direct stool PCR method were higher than those of the other two methods. Al-though PCR inhibitors may reduce its ability for the correct diagnosis of negative samples, it seems to be a reliable method for the detection of C. difficile infection. The weakness of the GDH-EIA method was its lower PPV, which can cause false-positive results. Toxin patterns and pulsotypes of C. difficile isolates revealed a high similarity between the strains isolated from the same units. © 2020, Author(s)

Antibiotic susceptibility pattern and virulence genes in Enterococcus spp. Isolated from clinical samples of Milad hospital of Tehran, Iran

Background: Enterococcus spp. are part of the normal flora of humans and animals. The nosocomial pathogenicity of Enterococcus spp. has emerged in recent years and has caused great concern due to developing of resistance to many antimicrobial agents. Objectives: The current study aimed to determine the resistance pattern and the type of virulence genes in Enterococcus spp. isolated from Milad hospital of Tehran, Iran. Materials and Methods: The current observational study was conducted from Apr 2014 to Feb 2015 on a total of 149 Enterococcus species isolated from Milad hospital in Tehran, Iran. The antibiotic susceptibility pattern of the bacteria was determined by the disc diffusion method for eight antibiotics. Minimum inhibitory concentration (MIC) of vancomycin was also done using agar-dilution assay by clinical and laboratory standards institute (CLSI) recommendations. The sodA, esp, cyl, ace and gelE genes were detected by polymerase chain reaction (PCR) assay. Results: About 37.5, 73, 86.6, 35.8, 69, 60.8, 45 and 79 of the isolates were resistant to vancomycin, tetracycline, gentamicin, chloramphenicol, ciprofloxacin, penicillin, ampicillin and erythromycin, respectively. MIC on 38 of the isolates was � 256 μg/mL. Although, the prevalence of vancomycin-resistant Enterococcus (VRE) strains belonged to two species, E. faecium showed high resistance to a broad range of antibiotics. In total, 94 isolates were positive for esp, and 59, 48 and 3 isolates were positive for ace, cylA and gelE, Respectively. Conclusions: The results of the current study designate the important role of medical samples as reservoirs of resistance inducing elements. Early detection of VRE with their virulence trait will help to prevent the spread of vancomycin resistant Enterococcus species. Supervision for antibiotic usage in hospitals, especially for last option antibiotics, can prevent the spread of resistant isolates and losing all treatment options in the future. © 2016, Infectious Diseases and Tropical Medicine Research Center

Invasive candidiasis in intensive care unit; consensus statement from an Iranian panel of experts, July 2013

Invasive candidiasis (IC) is associated with high mortality in intensive care unit (ICU) patients. Timely diagnosis of this potentially fatal condition remains a challenge; on the other hand, the criteria for initiating empirical antifungal therapy in critically ill patients are not well defined in different patient population and ICU settings. Alongside the international guidelines, reaching regional and local consensus on diagnosis and management of IC in ICU setting is essential. This report summarizes our present status of IC management in ICU, considered by a group of Iranian experts in the fields of intensive care and infectious diseases. A round table of 17 experts was held to review the available data and discuss the optimal treatment strategies for IC in critical care setting. Comparative published data on the management of IC were analytically reviewed and the commonly asked questions about the management of IC in ICU were isolated. These questions were interactively discussed by the panel and audience responses were taken to consolidate point-to-point agreement with the panel arriving at consensus in many instances. The responses indicated that patient's risk stratification, clinical discretion, fungal diagnostic techniques and the empirical therapy for IC are likely to save more patients. Treatment options were recommended to be based on the disease severity, prior azole exposure, and the presence of suspected azoleresistant Candida species. This report was reviewed, edited and discussed by all participants to include further evidence-based insights. The panel expects such endorsed recommendations to be soon formulated for implementation across the country. © 2014 The Author(s)