Drug resistance patterns of bacteria isolated from patients with nosocomial pneumonia at Tehran hospitals during 2009-2011

INTRODUCTION: Nosocomial pneumonia remains an important cause of mortality and morbidity worldwide. Surveillance programs play an important role in the identification of common etiologic agents and local patterns of antimicrobial resistance. METHODOLOGY: In this study we determined the frequency and antimicrobial susceptibility of pathogens isolated from patients with nosocomial pneumonia during 2009 to 2011. RESULTS: A total of 642 bacteria were isolated from 516 suspected samples. Acinetobacter baumannii (21.1%, n = 136), was the commonest isolated pathogen followed by Pseudomonas aeruginosa (17.4%, n = 112), Staphylococcus aureus (15.8%, n = 102) and enterococci (8.4% n = 54). The most effective therapeutic agents against A. baumannii were polymyxin B (95.5% susceptible), ceftriaxone/tazobactam (72% susceptible) and levofloxacin (52.9% susceptible). Polymixin B (89.2% susceptible), ceftriaxone/tazobactam (89.2% susceptible) and piperacillin-tazobactam (80.3% susceptible) were found to be the most active agents against P. aeruginosa. Extended-spectrum beta-lactamases were detected among isolates of K. pneumoniae (45.4%) and E. coli (20.3%). Overall, the prevalence of methicillin-resistant S. aureus and vancomycin resistant enterococci were 80.4% and 40.7% respectively. Linezolid was found to be the most active antibiotic against these pathogens. The etiology of 50% of the nosocomial infection cases was polymicrobial. CONCLUSIONS: The combination of ceftriaxone/tazobactam seems to be beneficial agent against multidrug-resistant Gram-negative bacilli isolated form respiratory tract infections. The results of our study can be used for guiding appropriate empiric therapy in this geographic region

Molecular epidemiology and nitrofurantoin resistance determinants of nitrofurantoin-non-susceptible Escherichia coli isolated from urinary tract infections

Objectives: The worldwide emergence of multidrug-resistant uropathogens has resulted in the revival of old antibiotics such as nitrofurantoin (NIT) for the treatment of uncomplicated urinary tract infections (UTIs). This study aimed to identify determinants of NIT resistance and to investigate the genetic diversity of NIT-resistant (NIT-R) Escherichia coli isolates. Methods: Six NIT-R and three NIT-susceptible clinical E. coli isolates from patients with UTI were studied. The susceptibility of the isolates to various classes of antibiotics was evaluated by disk diffusion. The presence of plasmid-encoded efflux pump genes (oqxA and oqxB) was investigated by PCR. Nucleotide sequences of the nfsA, nfsB and ribE genes were determined. The genetic relatedness of the NIT-R isolates was evaluated by multilocus sequence typing (MLST). Results: All six NIT-R isolates were characterised with high-level NIT resistance (MIC � 512 mg/L) and they belonged to five distinct STs including ST131 (n = 2), ST73, ST405, ST10 and ST354 (n = 1 each). Amikacin, carbapenems, minocycline, tigecycline and fosfomycin were the most active agents against the studied uropathogens. The oqxA and oqxB genes were not detected in any isolate. All NIT-R isolates harboured inactivating genetic alterations in nfsA and nfsB NfsA H11Y, S33N, S38Y, W212R substitutions, �g638 (frameshift), �a64-g73 (frameshift) and NfsB F84S, P45S, W94Stop, E197Stop substitutions, �nfsB locus. The ribE gene of most isolates was unaffected, except for one isolate co-harbouring a deleterious RibE G85C substitution and NfsA/B alterations. Conclusion: NIT resistance in the studied E. coli isolates was mainly mediated by nfsA and nfsB alterations. © 201

Genotypic characterization of Staphylococcus aureus isolated from a burn centre by using agr, spa and SCCmec typing methods

Infections caused by Staphylococcus aureus remain a major global healthcare problem. We aimed to find the common lineages of S. aureus strains circulating in a burn hospital in Tehran. A total of 167 isolates of S. aureus obtained from patients, healthcare workers (HCWs) and environment in Shahid Motahari burn hospital were genotyped by using spa, agr and staphylococcal cassette chromosome mec (SCCmec) typing methods. Antimicrobial susceptibility testing was performed by using the disc diffusion method. The frequency of methicillin-resistant S. aureus (MRSA) was 64.7 (n = 108), with distribution frequencies among patient, HCW and surface isolates of 64.2 (n = 79), 50 (n = 7) and 73.3 (n = 22), respectively. SCCmec type III (75, n = 81) was found to be the most frequent SCCmec type among MRSA isolates, followed by SCCmec type I (20.4, n = 22) and SCCmec type IV (1.8, n = 2). The remaining MRSA isolates (2.8, n = 3) were nontypeable by this method. About 78.4 (n = 131), 10.2 (n = 17) and 4.8 (n = 8) of all isolates were characterized as agr types I, II and III, respectively, and the other isolates (6.6) were nontypeable. spa types t030 and t037 constituted the first and second most predominant spa types found in 56.4 (n = 57) and 25.6 (n = 26) of isolates, respectively. We also report here a novel spa type, t16471. The most prevalent genotypes of the isolates found among patient, surface and HCW samples were SCCmec type III/t030, t037/agr type I. Continuous tracking of epidemic isolates and better hospital infection control policies are recommended to efficiently prevent the spread of bacteria to inpatients. © 201

Survival in amoeba: a major selection pressure on the presence of bacterial copper and zinc resistance determinants?: identification of a "copper pathogenicity island"

The presence of metal resistance determinants in bacteria usually is attributed to geological or anthropogenic metal contamination in different environments or associated with the use of antimicrobial metals in human healthcare or in agriculture. While this is certainly true, we hypothesize that protozoan predation and macrophage killing are also responsible for selection of copper/zinc resistance genes in bacteria. In this review, we outline evidence supporting this hypothesis, as well as highlight the correlation between metal resistance and pathogenicity in bacteria. In addition, we introduce and characterize the "copper pathogenicity island" identified in Escherichia coli and Salmonella strains isolated from copper- and zinc-fed Danish pigs

Expression analysis of 10 efflux pump genes in multidrug-resistant and extensively drug-resistant Mycobacterium tuberculosis clinical isolates

Objectives: Active extrusion of antituberculosis drugs via efflux pumps (EPs) has been suggested as contributing to drug resistance in Mycobacterium tuberculosis. This study was conducted to determine the role of 10 drug efflux transporters in the development of drug resistance in a series of clinical M. tuberculosis isolates. Methods: A total of 31 clinical M. tuberculosis isolates without drug exposure 21 multi/extensively drug-resistant (M/XDR-TB) and 10 drug-susceptible isolates were studied. The expression profile of 10 EP genes, including efpA, mmr, stp, drrA, drrB, mmpL7, Rv1250, Rv1634, Rv2994 and Rv1258c, was investigated against the H37Rv standard strain by quantitative reverse transcription PCR (RT-qPCR). Results: Among the 21 M/XDR-TB isolates, 10 showed significantly increased levels of gene expression (>4-fold) for at least one of the studied EPs. Moreover, of the isolates with overexpressed genes, three and seven lacked genetic alterations in the surveyed regions of the rpoB + katG + inhA and katG + inhA genes, respectively. Whilst no elevation was observed in the expression of mmr, Rv1250, Rv1634 and Rv1258c genes in any of the isolates, drrA, stp and drrB were found to be the most commonly overexpressed, being overexpressed in seven, five and three isolates, respectively. Decreased minimum inhibitory concentrations (MICs) of rifampicin, but not isoniazid, were observed in the presence of the efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP). Conclusion: Overexpression of EP genes can contribute to the emergence of a MDR phenotype in M. tuberculosis. Inhibition of EPs may provide a promising strategy for improving tuberculosis treatment outcomes in patients infected with M/XDR-TB isolates. (C) 2019 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved

Novel vaccine candidates against Mycobacterium tuberculosis

Tuberculosis (TB) is now among the top ten causes of mortality worldwide being resulted in 1.7 million deaths including 0.4 million among people with HIV in 2016. The Bacille Calmette-Guerin (BCG) is the only available TB vaccine which fails to provide consistent protection against pulmonary TB in adults and adolescents despite being efficacious at protecting infants and young children from the most severe, often deadly forms of TB disease. To achieve the goal of global TB elimination by 2050 we will need new interventions including more improved vaccines that are effective in adult individuals who have not been infected with Mycobacterium tuberculosis as well as latently infected or immunocompromised subjects. In recent decades, multiple new vaccine candidates including whole cell vaccines, adjuvanted proteins, and vectored subunit vaccines have entered into the clinical trials. These new TB vaccines are hoped to provide encouraging safety and immunogenicity under various conditions including prevention of TB disease in adolescents and adults, as BCG replacement/boosters, or as therapeutic vaccines to reduce the duration of TB therapy. In this review, we will discuss the status of novel TB vaccine candidates currently under development in preclinical or clinical phases. © 2018 Elsevier B.V

Rapid detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis isolates using high resolution melting curve analysis

Aims and objectives: Early detection of multidrug-resistant tuberculosis (MDR-TB) is essential to prevent its transmission in the community and to initiate an effective anti-TB treatment regimen. The objective of this study is to evaluate molecular technique high resolution melting curve (HRM) assay to rapidly detect resistance-conferring mutations in rpoB and katG genes. Methods: HRM analysis was used to screen 95 Mycobacterium tuberculosis (MTB) clinical isolates including 20 rifampin resistant (RIF-R), 21 isoniazid resistant (INH-R), and 54 fully susceptible (S) isolates determined by proportion method of drug susceptibility testing. 19 MTB isolates with known drug susceptibility genotypes were used as control strains for initial validation of the assay. Polymerase chain reaction (PCR) amplicons from rpoB and katG genes were sequenced to investigate the frequency and type of mutations and to confirm HRM results. Results: All RIF-S and INH-S isolates generated wild-type HRM curves and were accurately identified as susceptible by this method. Similarly 19 out of 20 RIF-R and 18 out of 21 INH-R isolates correctly exhibited mutant-type HRM curves. These results were similar to those obtained by direct sequencing. However, 1 RIF-R and 3 INH-R isolates were falsely identified as susceptible by HRM assay. These strains were confirmed as having no mutation in their target regions by sequencing. The main mutations involved in RIF and INH resistance were found at codons rpoB531 (60% of RIF-R isolates) and katG315 (85.7% of INH-R isolates), respectively. Conclusions: HRM was found to be a reliable, rapid and low-cost method to characterize drug susceptibility of clinical TB isolates in resource-limited settings

MgrB Alterations Mediate Colistin Resistance in Klebsiella pneumoniae Isolates from Iran

Colistin is one of the last-resort therapeutic agents to combat multidrug-resistant Gram-negative bacteria (GNB) including Klebsiella pneumoniae. Although it happens rarely, resistance to colistin has been reported for several GNB. A total of 20 colistin resistant (col-R) and three colistin susceptible (col-S) clinical isolates of K. pneumoniae were studied to explore the underlying mechanisms of colistin resistance. The presence of plasmid encoded resistance genes, mcr-1, mcr-2, mcr-3, and mcr-4 genes were examined by PCR. The nucleotide sequences of pmrA, pmrB, phoP, phoQ, and mgrB genes were determined. To evaluate the association between colistin resistance and upregulation of pmrHFIJKLM and pmrCAB operons, transcriptional level of the pmrK and pmrC genes encoding for lipopolysaccharide target modifying enzymes was quantified by RT-qPCR analysis. None of the plasmid encoded resistance genes were detected in the studied isolates. Inactivation of MgrB due to nonsense mutations and insertion of IS elements was observed in 15 col-R isolates (75%). IS elements (IS5-like and IS1-like families) most commonly targeted the coding region and in one case the promoter region of the mgrB. Complementation with wild-type MgrB restored colistin susceptibility in isolates with altered mgrB. All col-R isolates lacked any genetic alterations in the pmrA, phoP, and phoQ genes and substitutions identified in the pmrB were not found to be involved in resistance conferring determined by complementation assay. Colistin resistance linked with upregulation of pmrHFIJKLM and pmrCAB operons with the pmrK and pmrC being overexpressed in 20 and 11 col-R isolates, respectively. Our results demonstrated that MgrB alterations are the major mechanisms contributing to colistin resistance in the tested K. pneumoniae isolates from Iran

Determination of circulating Mycobacterium tuberculosis strains and transmission patterns among TB patients in Iran, using 15 loci MIRU-VNTR

Aims and objectives: Tuberculosis (TB) is considered one of the most important pathogens in the world. Iran has a moderate TB incidence, but borders two high-burden TB countries to the east and one high-burden multidrug-resistant (MDR)-TB country to the north. Limited information is available on the genetic diversity and transmission dynamics of MTB in Iran. To determine the MTB genotypes and their transmission patterns among patients, a collection of isolates from different parts of Iran were genotyped. Methods: Genotypes were generated by means of standard 15-locus variable number tandem repeat (VNTR) for 121 MTB clinical isolates collected from three provinces of Iran, including Sistan–Baluchestan (southeast province of Iran, with the highest rate of TB), Kermanshah (western part of Iran with high TB/human immunodeficiency virus [HIV] cases) and Tehran (the capital of Iran). Results: Sixty-six distinct mycobacterial interspersed repetitive unit (MIRU)-VNTR patterns were detected among 121 isolates. Seventy-five strains were grouped into 20 clusters, and 46 isolates were unique. The genetic diversity of strains from Sistan–Baluchestan was higher than that in the other province. All isolates from Tehran or Kermanshah that were grouped into clusters shared more identical patterns with Sistan–Baluchestan. The Hunter-Gaston discriminatory index (HGDI) was 0.972, indicating a high power of discrimination for MIRU-VNTR typing. The MIRU 16, ETR-A, ETR-E, MTUB04 loci were designated as highly discriminative. Conclusions: MIRU-VNTR typing showed a high genetic diversity and suggests the possibility of transmission from Sistan–Baluchestan to other provinces of Iran. This method could be considered a suitable tool for studying the transmission routes of TB and leading to more appropriate measures for TB control. MIRU-VNTR typing leads to a much better understanding of the bacterial population structure and phylogenetic relationships between strains of MTB in different regions of Iran