Role and Function of KPC and MBL Enzymes in Increasing the Pathogenicity of Pseudomonas Aeruginosa Isolated from Burn Wounds

BACKGROUND AND OBJECTIVE: Pseudomonas aeruginosa is one of the main causes of hospital infections. Pathogenic factors in this bacterium may play a role in the resistance to carbapenem and beta-lactam. The purpose of this study was to evaluate the role and function of KPC and MBL enzymes in increasing the pathogenicity of Pseudomonas aeruginosa isolated from burn wounds. METHODS: In this cross-sectional study, 63 isolates of Pseudomonas aeruginosa from burn wounds of different patients were isolated using biochemical tests such as fermentation of sugars in the OF medium, oxidase test, and so on. Determination of resistance pattern and strains with metallobetalactamase and carbapenema was done by disc diffusion method. The oprD gene was used for molecular confirmation of isolates. PCR method was used to detect pathogenicity genes. FINDINGS: Out of 63 isolates of Pseudomonas aeruginosa isolated from burn patients, 10 isolates (15.83%) had KPC enzyme and 13 isolates (20.63%) had MBL enzymes. Doripenem, Ertapenem and meropenem were the most frequent. Also, the lasB gene was observed in 43 isolates (68.25%), plcN gene in 41 isolates (65.07%), lasA gene in 20 isolates (31.74%), apr in 60 isolates (95.23%), phzI gene in 53 isolates (84.12%), the phzII gene in 38 isolates (60.31%), phzH gene in 30 isolates (47.61%) and plcH gene in 56 isolates (88.88%). CONCLUSION: The results of this study showed that the production of Carbapnemase and MBL enzymes increased the pathogenicity of Pseudomonas aeruginosa isolated from burn wounds

Investigation of the Relationship between the Presence of Chromosomal and Plasmid-Encoded AmpC Genes and Type of Clinical Specimen in Pseudomonas Aeruginosa

BACKGROUND AND OBJECTIVE: Different clinical specimens play a decisive role in the type and nature of drug resistance in pathogenic organisms. Occasionally, the presence of certain antibiotic resistance genes is associated with the type of clinical specimen. The aim of this study was to determine the relationship between the presence of chromosomal and plasmid-encoded AmpC genes and type of clinical specimen in Pseudomonas aeruginosa. METHODS: In this descriptive and experimental study, 114 isolates of Pseudomonas aeruginosa, and clinical specimens including blood, urine, wound secretion, burn injuries were collected from teaching hospitals in Hamadan. The presence of chromosomal and plasmid-encoded AmpC genes was evaluated using multiplex PCR technique. FINDINGS: The plasmid-encoded AmpC genes were observed more than chromosomal genes in Pseudomonas aeruginosa isolates. The FOX gene with a value of 29 (37.66%) (p≤0.037) and DHA gene with a value of 5(6.4%) (p≤0.015) in plasmid-encoded AmpC genes, while FOX gene with a value of 39 (48.75%) (p≤0.001) and MOX gene with a value of 2 (7.36%) in chromosomal AmpC genes had the highest and lowest frequency, respectively. CONCLUSION: The results of the study showed that the presence of chromosomal and plasmid-encoded AmpC genes may have various frequencies according to the type of clinical specimen

Identification and Determination of the Relationship between ccr Alleles and Antibiotic Resistance in Clinical Isolates of Methicillin Resistant Staphylococcus aureus

BACKGROUND AND OBJECTIVE: Resistance to methicillin and the presence of the ccr gene in Staphylococcus aureus have provided the basis for the emergence of methicillin resistant strains. The aim of this study was to identify the ccr cassette alleles in methicillin-resistant S.aureus strains and to determine the relationship between the presence of these casts with a multivariate process. METHODS: In this study, 135 clinical isolates of methicillin-resistant S.aureus was isolated by genotypic methods. ccr gene cassette was evaluated qualitatively by multiplex PCR method. Data was analyzed using SPSS version 16 and also, the chi - square test was used. FINDINGS: Out of 135 strains of S.aureus resistant to methicillin, penicillin and erythromycin antibiotic resistance were the most frequent, more than 90%, respectively. Also, ccr gene cassette in the study on genes ccrA/B1, ccrA/B2, ccrA/B3, ccrA/B4, ccrA2/B, ccrC had taken place, respectively, in 2 isolates (1.3%) for gene ccrA/B1, 12 isolates (8.2%) ccrA/B2, 15 isolates (10.34 %) ccrA/B3, 2 isolates (1.3%) ccrA/B4, 4 isolates (8.2 percent) ccrA2/B and 22 isolates (15.87 %) were positive for the gene ccrC. A significant correlation between the presence of these genes and antibiotic distribution was observed (p=0.05). CONCLUSION: The ccr gene cassette can provide a background of resistance to various antibiotics in methicillin-resistant S.aureus strains

Evaluation of Real-time PCR-based DNA melting method for detection of Enterococcus faecalis and Enterococcus faecium in clinical isolates

BACKGROUND AND OBJECTIVE: Human error and timeless in conventional techniques to identify species Enterococcus faecium and Enterococcus faecalis are two species of the pathogenic species in humans, more accurate techniques is essential. The aim of this study is to design a method of quickly and accurately using DNA melting by Real Time PCR technique to identify and separate the two species in clinical isolates. METHODS: In this experimental study, the bacterial isolates in the Department of Microbiology Bank of Hamadan University of Medical Sciences was used. Design of primers was done by proprietary software and selecting DivIVA gene for Enterococcus faecalis and alanine racemase for Enterococcus faecium was performed. Isolates identification was evaluated by using Real Time PCR test and melting curve temperature of DNA. FINDINGS: Susceptibility of primers designed in divIVA gene (specific for E. faecalis) and alanine-racemase gene (specific for E. faecium) was 15CFU/ml per reaction. Specificity of designed primers by using DNA melting curve analysis was 76.6 for E. faecalis and 80.93 for E. faecium which showed considerable different in comparison with another microorganism. CONCLUSION: Using the results obtained in this study, primers designed were sensitivity and specificity for diagnosis and differentiation of Enterococcus faecium and Enterococcus faecalis species in clinical isolates

Untersuchung der Verteilung und des Antibiotikaresistenzprofils klinischer Isolate von Gruppe-B-Streptokokken (Streptococcus agalactiae)

Aim: The aims of the present study were to determine the antibiotic susceptibility profils with particular emphasis on susceptible or resistant strains to macrolides and lincosamids antibiotics and to determine possible antibiotic resistance mechanisms occurring in group B streptococci (GBS) strains using PCR assay and disk diffusion method.Methods: A total of 62 clinical GBS strains were investigated. Antibacterial susceptibility testing was performed using the disk diffusion method and inducible resistance test for clindamycin by standard double disk diffusion or D-zone test for all isolates to differentiate macrolide resistance phenotype (M), constitutive macrolide-lincosamide-streptogramin B phenotype (cMLSB) and induced macrolide-lincosamide-streptogramin B phenotype (iMLSB). In addition, minimum inhibitory concentrations (MIC) of penicillin were determined for all isolates. Finally, possible existence of antibiotic resistance genes for erythromycin (ermTR , ermB and mefA/E ) and for clindamycin (linB ) were examined among isolates using PCR assay.Results: All 62 isolates were susceptible to penicillin, ampicillin, linezolid, cefazoline and vancomycin. However, 93.5% (n=58) of isolates showed an increased MIC to penicillin. The overall rate of erythromycin resistance was 35.5% (n=22). All erythromycin-resistant isolates displayed the M phenotype (100%, n=22). All three erythromycin resistance genes (i.e. ermTR , ermB and mefA/E ) were found in erythromycin-resistant isolates.Conclusion: It was concluded that prescribing antibiotic without antibacterial susceptibility tests should be prevented because of the high prevalence of erythromycin-resistant GBS strains and the fact that erythromycin-resistant GBS strains has shown an increased MIC to penicillin, as the drug of choice for treating GBS infections.Zielsetzung: In der vorliegenden Studie wurden die Antibiotikaempfindlichkeit mit dem Schwerpunkt Empfindlichkeit gegen Makrolide und Linkosamide bestimmt und mögliche Resistenzmechanismen von Gruppe-B-Streptokokken (GBS) mittels PCR und Plättchendiffusionsmethode untersucht. Methode: Es wurden 62 klinische GBS-Isolate untersucht. Die Resistenztestung wurde mit der Plättchendiffusionsmethode und der induzierbaren Resistenz gegen Clindamycin mittels Doppelplättchen-Diffusion oder D-Zonen-Test durchgeführt, um zwischen der phänotypischen Makrolidresistenz (M), der konstitutiven phänotypischen Makrolid-Linkosamid-Streptogramin B (cMLSB)- und der induzierbaren phänotypischen Makrolid-Linkosamid-Streptogramin B (iMLSB)-Resistenz zu differenzieren. Ergänzend wurde ebenfalls für alle Isolate die minimale Hemmkonzentration (MHK) für Penizillin bestimmt. Schließlich wurde die mögliche Existenz der Erythromycin- (ermTR , ermB und mefA/E ) und Clindamycin-Resistenzgene (linB ) mittels PCR untersucht. Ergebnisse: Alle 62 Isolate waren gegen Penicillin, Ampicillin, Linezolid, Cefazolin und Vancomycin empfindlich. Allerdings zeigten 93,5% (n=58) der Isolate eine erhöhte MHK gegenüber Penicillin. Der Anteil Erythromycin-resistenter Isolate betrug 35,5% (n=22). Alle Erythromycin-resistenten Isolate repräsentierten den M-Phänotyp. In Erythromycin-resistenten Isolaten waren alle 3 Erythromycin-Resistenzgene (ermTR , ermB und mefA/E ) nachweisbar.Schlussfolgerung: Wegen der hohen Prävalenz Erythromycin-resistenter GBS-Stämme und der Tatsache, dass Erythromycin-resistente GBS-Stämme eine erhöhte MHK gegen Penicillin aufwiesen, sollten Infektionen durch GBS nicht ohne vorherige Resistenztestung behandelt werden