Phenotypic and genotypic characteristics of tetracycline resistant Acinetobacter baumannii isolates from nosocomial infections at Tehran hospitals

Objective(s): To date, the most important genes responsible for tetracycline resistance among Acinetobacter baumannii isolates have been identified as tet A and tet B. This study was carried out to determine the rate of resistance to tetracycline and related antibiotics, and mechanisms of resistance. Materials and Methods: During the years 2010 and 2011, a total of 100 A. baumannii isolates were recovered from patients in different hospitals of Tehran, Iran. Antimicrobial susceptibility to tetracycline, minocycline, doxicycline and tigecycline was evaluated by E-test. Polymerase chain reaction (PCR) of the tet A and tet B genes was performed using specific primers, after which the isolates were subjected to Repetitive Extragenic Palindromic-PCR (PCR) to identify the major genotypes. Results: Of all isolates, 89 were resistant to tetracycline (MIC50 = 32 mu g/ml, MIC90 = 512 mu g/ml). Minocycline with the resistant rate of 35 (MIC50 = 16 mu g/ml, MIC90 = 32 mu g/ml) and doxicycline with the resistant rate of 25 (MIC50 = 16 mu g/ml, MIC90= 32 mu g/ml) have a good activity against A. baumannii isolates. All isolates were sensitive to tigecycline. Frequencies of tet B and tet A genes and coexistence of tet A and tet B among the isolates resistant to tetracycline, were 87.6, 2.2 and 1.1, respectively. Distribution of REP-types among A. baumannii isolates was types A (40), B (30), C (10), D (5) and E (5). Conclusion: It seems that tet A and tet B genes play an important role in the induction of resistance towards tetracyclines used in this study. It is suggested that further studies focus on other antimicrobial drugs and combinations in order to achieve a successful therapy against multi drug resistance (MDR) A. baumannii strains in Iran

Identification of Streptococcus agalactiae by fluorescent in situ hybridization compared to culturing and the determination of prevalence of Streptococcus agalactiae colonization among pregnant women in Bushehr, Iran

Background: Pregnant women colonized by Streptococcus agalactiae (group B streptococci [GBS]) may transfer this microorganism to their newborns. S. agalactiae is an important cause of pneumonia, sepsis, and meningitis in newborns. Fluorescent in situ hybridization (FISH) is considered as a method of identification in the field of diagnostic microbiology. In this paper, we have designed a study to compare the DNA FISH after 7 h Lim broth enrichment and culturing for the identification of S. agalactiae and to determine the prevalence of vaginal colonization by S. agalactiae among pregnant women in Bushehr, Iran. Methods: Vaginal swab specimens were obtained from 285 pregnant women at 35 weeks or more than 35 weeks of gestation. The specimens were inoculated into Lim broth. In order to evaluate the sensitivity and specificity of GBS DNA FISH after 7 h Lim broth enrichment, the specimens were tested using both FISH and conventional culture methods. In addition, the prevalence of GBS colonization was determined. Results: Based on the results of this study, both the sensitivity and specificity of FISH were 100%. S. agalactiae was detected by both culture and FISH in 27 of the 285 pregnant women. Thus, the prevalence of GBS colonization was 9.5%. Conclusions: Since short-term (7 h) Lim broth enrichment followed by FISH using oligonucleotide probes showed a high sensitivity and specificity, this protocol is therefore a highly accurate and relatively rapid method for the detection of S. agalactiae. Our analysis suggests that the use of DNA FISH to screen for S. agalactiae colonization in pregnant women may be considered in the absence of GBS culture availability

Phenotypic and genotypic evaluation of fluoroquinolone resistance in clinical isolates of Staphylococcus aureus in Tehran

Background: Fluoroquinolones are broad-spectrum antibiotics widely used in the treatment of bacterial infections such as Staphylococcus aureus isolates. Resistance to these antibiotics is increasing. Material/Methods: The occurrence of mutations in the grlA and gyrA loci were evaluated in 69 fluoroquinolone-resistant S. aureus isolates from 2 teaching hospitals of Tehran University of Medical Sciences. Results: Out of the 165 S. aureus isolates, 87 (52.7) were resistant to methicillin and 69 (41.8) were resistant to fluoroquinolone. Fluoroquinolone-resistant S. atoms isolates had a mutation at codon 80 in the grlA gene and different mutational combinations in the gyrA gene. These mutational combinations included 45 isolates at codons 84 and 86,23 isolates at codons 84,86 and 106 and 1 isolate at codons 84, 86 and 90. Fluoroquinolone-resistant S. aureus isolates were clustered into 33 PFGE types. Conclusions: The findings of this study show that the fluoroquinolone-resistant S. aureus strains isolated in the teaching hospitals in Tehran had multiple mutations in the QRDRs region of both grlA and gyrA genes

Comparison of Antibacterial Effect of Four Irrigation Solutions in Primary Root Canal Infections: A Clinical Study

Introduction: Reducing the bacterial count from the root canal system is one of the main stages in root canal treatment. The aim of the present clinical study was to compare the antibacterial effect of four intracanal irrigants in primary endodontic infections using both microbiological culture and quantitative Real-time Polymerase Chain Reaction (qRT-PCR) technique. Methods and Materials: Forty patients with primarily infected single rooted premolars were selected and then randomly divided into 4 groups according to the intra canal irrigant used: 5.25% Sodium hypochlorite (NaOCl), Hypoclean (Ogna Laboratori Farmaceutici, Muggiò, Italy), 2% chlorhexidine glouconate (CHX) and CHX-Plus (Vista Dental Products, Racine, WI, USA). Samples were collected before and after chemomechanical preparation and were evaluated by bacterial culture and RT-PCR technique for Enterococcus faecalis and Fusobacterium nucleatum. Data analyzed by repeated measured ANOVA. The significance level was set at 0.05. Results: Four irrigation solutions significantly reduced the total numbers of cultivable bacteria (P<0.05). No statistically differences were found among the antibacterial effects of 5.25% NaOCl (99.93%), Hypoclean (99.94%), 2% CHX (99.77%) and CHX-Plus (99.83%) in reducing cultivable bacteria. Enterococcus faecalis and Fusobacterium nucleatum were no longer detected after preparation using four irrigants (100% reduction). Conclusions: All tested irrigants including 5.25% NaOCl, Hypoclean, 2% CHX and CHX-Plus significantly reduced the number of bacterial colonies in primary endodontic infections.Keywords: CHX-Plus; Endodontic Infection; Enterococcus faecalis; Fusobacterium nucleatum; Hypoclea

Antibacterial Substantivity of Carvacrol and sodium hypochlorite in infected bovine root dentin

INTRODUCTION: Various methods commonly used for cleaning and shaping root canals have not been successful in completely eradicating bacteria due to anatomic complexity and root canals irregularities. Disinfecting the canals with intracanal irrigants in addition to proper cleaning and shaping can produce a successful outcome. Antimicrobials with sustained antibacterial activity would be desirable for irrigation. The purpose of this study was to compare the antimicrobial substantivity of Carvacrol and 5.25% NaOCl in infected bovine root dentin. MATERIALS AND METHODS: One hundred and twenty dentin tubes prepared from bovine incisors were infected in vitro for 14 days with Enterococcus (E) faecalis. The specimens were divided into four groups including 1) Carvacrol, 2) NaOCl, 3) infected dentin tubes (positive control); and 4) sterile dentin tubes (negative control). Dentin chips were collected at five intervals (days 0, 1, 3, 7 and 28) using round burs with sequentially increasing diameters (which includes five layers of dentin) into Brain Heart Infusion (BHI) broth. In order to compare the pre- and post-irrigation antimicrobial activity of the irrigants the colony-forming units (CFU) were counted and classified as “CFU-before” and “CFU-after”. After culturing, the number of CFU with the various experimental time and dentinal layers was recounted. Two-way ANOVA test was used to analyze the effects of time and materials. One-way ANOVA and supplemental Tukey HSD test were used for pair comparison. RESULTS: CFU was significantly reduced in NaOCl group when compared to all other experimental groups (P<0.05). CONCLUSION: The substantivity of NaOCl was significantly greater than Carvacrol. Further studies are required to investigate and approve Carvacrol as a final irrigant

Methicillin and vancomycin resistant isolates of Staphylococcus aureus and Enterococcus faecalis recovered from bovine mastitis

NO ABSTRACT AVAILABLEMastitis is the most costly disease in the dairy industry. Selecting the proper antibiotic treatment is beneficial for economic and avoids the emergence of antimicrobial resistance. The objective of the present study was to investigate the prevalence of methicillin and vancomycin resistant isolates of mastitis-causing Staphylococcus aureus and Enterococcus faecalis as a probable source of transferable vancomycin resistance to staphylococci. A total of sixty-one Staphylococcus aureus and eight Enterococcusfaecalis isolates were investigated for genotypic and phenotypic antimicrobial resistance.Presence of the mecA, vanA and vanBgenes were surveyed by PCR. The MIC (Minimum Inhibitory Concentration) of vancomycin was determined bybroth microdilution test for all the isolates.Moreover, the antibiotic resistance patterns of the isolates to the most common classes of antibiotics used in dairy cattle such as β-lactam, macrolides and tetracyclines were determinedusing the disk diffusion method. Among Staphylococcus aureus isolates, one MRSA (methicillin-resistant Staphylococcus aureus) isolate was detected while 47.5% of isolates were detected as multidrug-resistant. Furthermore, no phenotypic and genotypicvancomycin-resistance Staphylococcus aureus was found. Most of the Enterococcus faecalis isolates (6/8) showed high MIC for vancomycin (in the range of 128- 1024 μg/ml) and one vanA-type Enterococcus faecalis was observed. This study indicates thatsince the source of transferable resistance to vancomycin exists in dairy farms, there is a potential for emerging and spreading VRSA (vancomycin-resistant Staphylococcus aureus) in dairy cattle which is a risk to animal and human health

In Vitro Comparison of the Effectiveness of Chlorhexidine and Two Calcium Hydroxide Formulations on Enterococcus Faecalis

INTRODUCTION: The aim of this in vitro study was to compare the effectiveness of three intracanal medicaments in disinfecting the root canal and dentin of experimentally infected human teeth with Enterococcus faecalis (EF). MATERIALS AND METHODS: One hundred extracted human single-rooted teeth were used. After root canal preparation, teeth were mounted in epoxy resin. Following sterilization, the teeth were infected for 28 days with EF. Then root canals were filled with one of three different disinfectants: viscous 2% Chlorhexidine (CHX), calcium hydroxide paste (CH) or a mixture of CH and CHX (n=30 in each group). Antimicrobial assessments were performed at 1, 3 and 7 days (n=10 in each time period). Microbial samples were obtained from root canals before and after the experiment. Also dentin samples were examined. The data was analyzed using Two- Way ANOVA test. RESULTS: The findings showed that there was no difference between experimental groups at different time periods. The mixture of CH/CHX in 7 days was able to eliminate EF completely from root canal system. The most elimination of EF was from dentinal tubules. CONCLUSION: According to the results of this in vitro study, viscous 2% CHX, mixture of CH with distilled water and 2% CHX are all effective disinfectants

Prevalence of high-level gentamicin-resistant Enterococcus faecalis and Enterococcus faecium in an Iranian hospital

This study was designed to determine the molecular characteristics and antimicrobial resistance of enterococcal strains isolated from patients admitted to an Iranian Hospital. Enterococcal strains were isolated from the burn patients. All strains were screened for genes encoding resistance to aminoglycoside aac(6')-Ie-aph(2'')-Ia, aph (3'), ant (4'), resistance to vancomycin (vanA, vanB), resistance to tetracycline (tetK, tetL, tetM, tetO), and resistance to erythromycin (ermA, ermB, ermC) by PCR and multiplex PCR-based methods. Genetic diversity was evaluated via Random Amplified Polymorphic DNA (RAPD)-PCR. All enterococcal isolates showed complete sensitivity to vancomycin with MIC � 0.5μg/ml. Resistance to gentamicin, tetracycline, erythromycin, ciprofloxacin or quinopristin-dalfopristin was detected, whilst more than 96.2% of isolates were high-level gentamicinresistant (HLGR) and multiple drug resistant. The most prevalent aminoglycoside resistance gene was aac(6')-Ie-aph(2'')-Ia, that was found in 96.2% (26/27) of the isolates. The most prevalent tetracycline resistance genes were tetM, found in 85.1% (23/27) followed by tetL and tetO found in 7.4% (2/27) of the isolates. The ermA and ermB genes were detected in 33.3% (9/27) and 44.4% (12/27) of the isolates respectively. RAPD-PCR analysis yielded 17 distinct profiles among 27 investigated isolates. One cluster of isolates shared the same RAPD pattern, while 16 isolates had unique RAPD pattern. Our study showed that during the examination time period one RAPD genotype was the common type and was disseminated among patients in the burn unit. Interestingly, most of these strains had an identical or very similar antibiotic and gene resistance pattern

High prevalence of direct repeat unit types of 10di, 8 h and 8i among methicillin resistant Staphylococcus aureus strains with staphylococcal cassette chromosome mec type IIIA isolated in Tehran, Iran 11 Medical and Health Sciences 1103 Clinical Sciences

Background: The emergence of methicillin-resistant Staphylococcus aureus (MRSA) is a main concern in burn care centers worldwide. The some reports of MRSA in Iran suggested that MRSA with type SCCmec III is common among burn patients. The aim of this study was to determine the prevalence, virulence genes, and antimicrobial susceptibility of the direct repeat units (dru) types of MRSA with SCCmec IIIA isolated from burn wounds in a burn care center in Tehran, Iran. Methods: In total, 165 S. aureus isolates were collected from clinical samples. In order to detect MRSA isolates, the mecA gene was amplified through the polymerase chain reaction (PCR) method. Antimicrobial susceptibility was tested using the disc agar diffusion test. Moreover, the PCR method was applied to determine SCCmec types, virulence genes, and antimicrobial resistance genes. The dru region was sequenced and thereby, dru types and dru repeats were identified. A similarity matrix was used to create minimum spanning tree (MST). Results: The prevalence of MRSA was 69 (114 out of 165 isolates). Most of MRSA isolates (61 out of 114, 53.5) were SCCmec type IIIA. All MRSA isolates were vancomycin-susceptible and more than 68 of MRSA isolates with SCCmec type IIIA were mupirocin resistant. The successful dru typing of isolates with SCCmec type IIIA revealed fourteen different dru types. There were two new dru types, namely dt10di and dt7aj. MST analysis indicated the presence of the three clusters of dt10di (cluster I), dt8i-dt8 h (cluster II), and dt11c-dt10ao-dt11dd-dt11a-dt10a (cluster III). There were significant differences between clusters I and II respecting antimicrobial resistance pattern and virulence genes. Conclusion: Three main dru clusters are prevalent in the study setting. The main dru types in the setting are dt10di, dt8i, and dt8 h. Dru typing can be used to differentiate MRSA strains with SCCmec IIIA. © 2019 The Author(s)

Distribution of genes encoding virulence factors and molecular analysis of Shigella spp. isolated from patients with diarrhea in Kerman, Iran

Shigella is one of the important causes of diarrhea worldwide. Shigella has several virulence factors contributing in colonization and invasion of epithelial cells and eventually death of host cells. The present study was performed in order to investigate the distribution of virulence factors genes in Shigella spp. isolated from patients with acute diarrhea in Kerman, Iran as well as the genetic relationship of these isolates. A total of 56 isolates including 31 S. flexneri, 18 S. sonnei and 7 S. boydii were evaluated by polymerase chain reaction (PCR) for the presence of 11 virulence genes (ipaH, ial, set1A, set1B, sen, virF, invE, sat, sigA, pic and sepA). Then, the clonal relationship of these strains was analyzed by multilocus variable-number tandem repeat analysis (MLVA) method. All isolates were positive for ipaH gene. The other genes include ial, invE and virF were found in 80.4%, 60.7% and 67.9% of the isolates, respectively. Both set1A and set1B were detected in 32.3% of S. flexneri isolates, whereas 66.1% of the isolates belonging to different serogroup carried sen gene. The sat gene was present in all S. flexneri isolates, but not in the S. sonnei and S. boydii isolates. The result showed, 30.4% of isolates were simultaneously positive and the rest of the isolates were negative for sepA and pic genes. The Shigella isolates were divided into 29 MLVA types. This study, for the first time, investigated distribution of 11 virulence genes in Shigella spp. Our results revealed heterogeneity of virulence genes in different Shigella serogroups. Furthermore, the strains belonging to the same species had little diversity