Antimicrobial Susceptibility Profiles among Pseudomonas aeruginosa Isolated from Professional SCUBA Divers with Otitis Externa, Swimming Pools and the Ocean at a Diving Operation in South Africa

SCUBA divers are predisposed to otitis externa caused by Pseudomonas aeruginosa, which is becoming increasingly multi-drug resistant (MDR). The present work assessed the antibiotic resistance profiles of P. aeruginosa obtained from SCUBA divers and their environment in Sodwana Bay, South Africa. Bacterial isolates from a total of 137 random water and ear swab samples were identified using biochemical and molecular methods. P. aeruginosa strains were further evaluated for antibiotic susceptibility using the Kirby–Bauer assay. Double disk synergy test (DDST) to confirm metallo-β-lactamase (MBL) production and PCR amplification of specific antibiotic resistance genes was performed. All (100%) 22 P. aeruginosa isolates recovered were resistant to 6 of the β-lactams tested including imipenem but exhibited susceptibility to trimethoprim–sulfamethoxazole. MBL production was observed in 77% of isolates while the most prevalent extended-spectrum β-lactamase (ESBL) genes present included blaAmpC (86.9%) followed by blaTEM (82.6%). Sulfonamide resistance was largely encoded by sul1 (63.6%) and sul2 (77.3%) genes with a high abundance of class 1 integrons (77.3%) of which 18.2% carried both Intl1 and Intl2. P. aeruginosa found in Sodwana Bay exhibits multi-drug resistance (MDRce) to several pharmaceutically important drugs with the potential to transfer antibiotic resistance to other bacteria if the judicious use of antibiotics for their treatment is not practiced.

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Antimicrobial resistance, plasmids and class 1 and 2 integrons occurring in Pseudomonas aeruginosa isolated from Brazilian aquatic environments

Water Science & Technology ◽

10.2166/wst.2013.676 ◽

2013 ◽

Vol 67 (5) ◽

pp. 1144-1149 ◽

Cited By ~ 9

Author(s):

Maria Olívia Zanetti ◽

Vinicius Vicente Martins ◽

André Pitondo-Silva ◽

Eliana Guedes Stehling

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Antibiotic Resistance Genes ◽

Aquatic Environments ◽

Resistance Plasmids ◽

Class 1 ◽

Potential Reservoir ◽

Polymerase Chain ◽

The University ◽

City Park

Pseudomonas aeruginosa is an important nosocomial pathogen also found in water, soil, plants and in human and animal fecal samples. In this study, 31 isolates from water samples were analyzed by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and PCR to detect integrons and investigated for antibiotic resistance and plasmidial profile. The results demonstrated the presence of plasmids in four isolates. Three of these, isolates from water in a city park (Curupira Park, Ribeirão Preto, Brazil) and a lake at the University of São Paulo (Campus of Ribeirão Preto), had plasmids with the same molecular weight (21MDa) and similar resistance profiles, although they were shown to be genetically different by ERIC-PCR. Class 1 and class 2 integrons were detected in one of these isolates. The results suggest that environmental P. aeruginosa strains may be a potential reservoir of plasmids and antibiotic resistance genes.

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Molecular characterisation of multidrug-resistant Pseudomonas aeruginosa from a private hospital in Durban, South Africa

Southern African Journal of Infectious Diseases ◽

10.4102/sajid.v33i2.19 ◽

2018 ◽

Vol 33 (2) ◽

pp. 38-41 ◽

Cited By ~ 1

Author(s):

Cosmos B. Adjei ◽

Usha Govinden ◽

Krishnee Moodley ◽

Sabiha Y. Essack

Keyword(s):

South Africa ◽

Pseudomonas Aeruginosa ◽

Molecular Mechanisms ◽

Private Hospital ◽

Treatment Options ◽

Multidrug Resistant ◽

Infection Prevention And Control ◽

Multi Drug Resistance ◽

Synergy Test ◽

And Control

Background: Multi-drug resistant Pseudomonas aeruginosa pose a clinical challenge globally. This study delineated the molecular mechanisms of resistance to β-lactam antibiotics in multidrug-resistant P. aeruginosa isolated from a single private hospital in Durban, South Africa and ascertained clonality with regard to the isolates carrying β-lactamase genes.Methods: Seventeen P. aeruginosa isolates recovered from sputum, urine, catheter tips, pus swabs, nasal swabs and endotracheal aspirates underwent MIC determination, and phenotypic screening using the Double Disk Synergy Test (DDST) and Modified Hodge Test (MHT) to identify putative extended-spectrum β-lactamases (ESBLs), metallo-β-lactamases and other carbapenemases. Selected β-lactamase encoding genes were genotypically confirmed by PCR and sequencing. REP-PCR was conducted to determine the clonal relatedness of the 11 isolates carrying β-lactamase genes. Results: Sixteen isolates (94%) were resistant to aztreonam and piperacillin, 15 isolates (88%) were resistant to imipenem and ticarcillin, 14 (82%) were resistant to meropenem, and 13 isolates (76%) were resistant to ceftazidime and piperacillin/tazobactam. Resistance to ciprofloxacin and amikacin were 82% and 29% respectively. Of the 17 isolates tested, GES-2, VIM-2 and OXA-21 were present in 10 (59%) four (24%) and one (6%) of the isolates respectively. Three of the isolates harboured both GES-2 and VIM-2 and one isolate harboured OXA-21 and VIM-2. REP-PCR revealed seven clusters with clusters A and F having two (18%) and four (36%) isolates respectively, while the remaining five isolates were unrelated.Conclusion: GES-2 and VIM-2 enzymes were predominantly responsible for carbapenemase resistance. Clones A and F intimated patient-to-patient spread within the ICU and surgical ICU. This apparent dissemination as well as the multi-drug resistance observed points to sub-optimal infection prevention and control and dwindling antibiotic treatment options for P. aeruginosa respectively in this institution.

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Analysis of Class 1 Integrons and Antibiotic Resistance Genes in Pseudomonas aeruginosa Strains from Benin City, Nigeria

Journal of Applied Sciences and Environmental Management ◽

10.4314/jasem.v24i4.14 ◽

2020 ◽

Vol 24 (4) ◽

pp. 633-637

Author(s):

B.O. Isichei-Ukah ◽

O.I. Enabulele

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Environmental Isolates ◽

Significance Level ◽

Beta Lactamases ◽

Class 1 Integrons ◽

Benin City ◽

Class 1

The presence of integrons and antibiotic resistance genes in the genome of Pseudomonas aeruginosa pose a serious problem in the treatment and control of infections caused by this pathogen in hospitals. This study was carried to analyse the presence of class 1 integrons and some antibiotic resistance genes on selected clinical and environmental strains of Pseudomonas aeruginosa. A total of 120 strains were employed for this study.The strains were confirmed using molecular method and species-specific primers targeting the 16S ribosomal ribonucleic acid (rRNA). Polymerase chain reaction (PCR) was used to detect the presence of class 1 integrons and resistance genes using appropriate primers and conditions. The strains were analysed for the presence of the following antibiotic resistance genes - aadA, blaPSE, blaAMPC, blaIMP and tetC encoding aminoglycosides, betalactamases, metallo-beta-lactamases (MBL) and tetracylines resistance respectively. On screening the isolates for the presence of class 1 integrons, 50/60 (83.3 %) clinical isolates and 46/60 (76.7 %) environmental isolates showed positive results (P > 0.05). In both clinical and environmental isolates, the highest occurring resistance genes were blaAMPC and tetC (encoding beta-lactamases and tetracylines respectively), while the least was observed in blaIMP (encoding metallo-beta-lactamases). In comparison, there was high significance difference (at P<0.01 significance level) in the resistance gene blaPSE between the clinical and environmental strains. The high prevalence of these resistance genes is a great threat in the treatment of Pseudomonas infections. Keywords: Pseudomonas aeruginosa, Resistance genes, Integrons, Beta-lactamases.

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Hypermutator Pseudomonas aeruginosa exploits multiple genetic pathways to develop multidrug resistance during long-term infections in the airways of cystic fibrosis patients

10.1101/809319 ◽

2019 ◽

Author(s):

C.A. Colque ◽

A.G. Albarracín Orio ◽

S. Feliziani ◽

R.L. Marvig ◽

A.R. Tobares ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Multidrug Resistance ◽

Chronic Infection ◽

Antibiotic Resistance Genes ◽

Resistance Mechanisms ◽

Multi Drug Resistance ◽

Genetic Pathways ◽

Resistance Evolution

ABSTRACTPseudomonas aeruginosa exploits intrinsic and acquired resistance mechanisms to resist almost every antibiotic used in chemotherapy. Antimicrobial resistance in P. aeruginosa isolated from cystic fibrosis (CF) patients is further enhanced by the occurrence of hypermutator strains, a hallmark of chronic CF infections. However, the within-patient genetic diversity of P. aeruginosa populations related to antibiotic resistance remains unexplored. Here, we show the evolution of the mutational resistome profile of a P. aeruginosa hypermutator lineage by performing longitudinal and transversal analyses of isolates collected from a CF patient throughout 20 years of chronic infection. Our results show the accumulation of thousands of mutations with an overall evolutionary history characterized by purifying selection. However, mutations in antibiotic resistance genes appear to be positively selected, driven by antibiotic treatment. Antibiotic resistance increased as infection progressed towards the establishment of a population constituted by genotypically diversified coexisting sub-lineages, all of which converged to multi-drug resistance. These sub-lineages emerged by parallel evolution through distinct evolutionary pathways, which affected genes of the same functional categories. Interestingly, ampC and fstI, encoding the β-lactamase and penicillin-binding protein 3, respectively, were found among the most frequently mutated genes. In fact, both genes were targeted by multiple independent mutational events, which led to a wide diversity of coexisting alleles underlying β-lactam resistance. Our findings indicate that hypermutators, apart from boosting antibiotic resistance evolution by simultaneously targeting several genes, favor the emergence of adaptive innovative alleles by clustering beneficial/compensatory mutations in the same gene, hence expanding P. aeruginosa strategies for persistence.IMPORTANCEBy increasing mutation rates, hypermutators boost antibiotic resistance evolution by enabling bacterial pathogens to fully exploit their genetic potential and achieve resistance mechanisms for almost every known antimicrobial agent. Here, we show how co-existing clones from a P. aeruginosa hypermutator lineage that evolved during 20 years of chronic infection and antibiotic chemotherapy, converged to multidrug resistance by targeting genes from alternative genetic pathways that are part of the broad P. aeruginosa resistome. Within this complex assembly of combinatorial genetic changes, in some specific cases, multiple mutations are needed in the same gene to reach a fine tuned resistance phenotype. Hypermutability enables this genetic edition towards higher resistance profiles by recurrently targeting these genes, thus promoting new epistatic relationships and the emergence of innovative resistance-conferring alleles. Our findings help to understand this link between hypermutability and antibiotic resistance, a key challenge for the design of new therapeutic strategies.

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Novel Chromosome-Borne Accessory Genetic Elements Carrying Multiple Antibiotic Resistance Genes in Pseudomonas aeruginosa

Frontiers in Cellular and Infection Microbiology ◽

10.3389/fcimb.2021.638087 ◽

2021 ◽

Vol 11 ◽

Author(s):

Ting Yu ◽

Huiying Yang ◽

Jun Li ◽

Fangzhou Chen ◽

Lingfei Hu ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Genetic Elements ◽

Integrative And Conjugative Elements ◽

Class 1 ◽

Additional Resistance ◽

First Time ◽

Genetic Comparison

Pseudomonas aeruginosa is noted for its intrinsic antibiotic resistance and capacity of acquiring additional resistance genes. In this study, the genomes of nine clinical P. aeruginosa isolates were fully sequenced. An extensive genetic comparison was applied to 18 P. aeruginosa accessory genetic elements (AGEs; 13 of them were sequenced in this study and located within P. aeruginosa chromosomes) that were divided into four groups: five related integrative and conjugative elements (ICEs), four related integrative and mobilizable elements (IMEs), five related unit transposons, and two related IMEs and their two derivatives. At least 45 resistance genes, involved in resistance to 10 different categories of antibiotics and heavy metals, were identified from these 18 AGEs. A total of 10 β-lactamase genes were identified from 10 AGEs sequenced herein, and nine of them were captured within class 1 integrons, which were further integrated into ICEs and IMEs with intercellular mobility, and also unit transposons with intracellular mobility. Through this study, we identified for the first time 20 novel MGEs, including four ICEs Tn6584, Tn6585, Tn6586, and Tn6587; three IMEs Tn6853, Tn6854, and Tn6878; five unit transposons Tn6846, Tn6847, Tn6848, Tn6849, and Tn6883; and eight integrons In1795, In1778, In1820, In1784, In1775, In1774, In1789, and In1799. This was also the first report of two resistance gene variants blaCARB-53 and catB3s, and a novel ST3405 isolate of P. aeruginosa. The data presented here denoted that complex transposition and homologous recombination promoted the assembly and integration of AGEs with mosaic structures into P. aeruginosa chromosomes.

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Methods for the targeted sequencing and analysis of integrons and their gene cassettes from complex microbial communities

10.1101/2021.09.08.459516 ◽

2021 ◽

Author(s):

Timothy M. Ghaly ◽

Anahit Penesyan ◽

Alexander Pritchard ◽

Qin Qi ◽

Vaheesan Rajabal ◽

...

Keyword(s):

Antibiotic Resistance ◽

Antibiotic Resistance Genes ◽

Pcr Amplification ◽

Gene Cassette ◽

Human Pathogens ◽

Diverse Range ◽

Gene Cassettes ◽

Wide Range ◽

Class 1 ◽

Range Of Functions

AbstractIntegrons are bacterial genetic elements that can integrate mobile gene cassettes. They are mostly known for spreading antibiotic resistance cassettes among human pathogens. However, beyond clinical settings, gene cassettes encode an extraordinarily diverse range of functions important for bacterial adaptation. The recovery and sequencing of cassettes has promising applications, including: surveillance of clinically important genes, particularly antibiotic resistance determinants; investigating the functional diversity of integron-carrying bacteria; and novel enzyme discovery. Although gene cassettes can be directly recovered using PCR, there are no standardised methods for their amplification and, importantly, for validating sequences as genuine integron gene cassettes. Here, we present reproducible methods for the PCR amplification, sequence processing, and validation of gene cassette amplicons from complex communities. We describe two different PCR assays that either amplify cassettes together with integron integrases, or gene cassettes together within cassette arrays. We compare the use of Nanopore and Illumina sequencing, and present bioinformatic pipelines that filter sequences to ensure that they represent amplicons from genuine integrons. Using a diverse set of environmental DNAs, we show that our approach can consistently recover thousands of unique cassettes per sample and up to hundreds of different integron integrases. Recovered cassettes confer a wide range of functions, including antibiotic resistance, with as many as 300 resistance cassettes found in a single sample. In particular, we show that class 1 integrons appear to be collecting and concentrating antibiotic resistance genes out of the broader diversity of cassette functions. The methods described here can be applied to any environmental or clinical microbiome sample.

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Quintuplex PCR to Detect Antibiotic Resistance Genes in Streptococcus pneumoniae

Journal of Clinical and Health Sciences ◽

10.24191/jchs.v2i1.5861 ◽

2016 ◽

Vol 1 (2) ◽

pp. 22 ◽

Cited By ~ 2

Author(s):

Navindra Kumari Palanisamy ◽

Parasakthi Navaratnam ◽

Shamala Devi Sekaran

Keyword(s):

Antibiotic Resistance ◽

Streptococcus Pneumoniae ◽

Resistance Genes ◽

Bacterial Pathogen ◽

Antibiotic Resistance Genes ◽

Pcr Amplification ◽

Penicillin Resistance ◽

Penicillin Binding Proteins ◽

Efflux Mechanism ◽

Mic Value

Introduction: Streptococcus pneumoniae is an important bacterial pathogen, causing respiratory infection. Penicillin resistance in S. pneumoniae is associated with alterations in the penicillin binding proteins, while resistance to macrolides is conferred either by the modification of the ribosomal target site or efflux mechanism. This study aimed to characterize S. pneumoniae and its antibiotic resistance genes using 2 sets of multiplex PCRs. Methods: A quintuplex and triplex PCR was used to characterize the pbp1A, ermB, gyrA, ply, and the mefE genes. Fifty-eight penicillin sensitive strains (PSSP), 36 penicillin intermediate strains (PISP) and 26 penicillin resistance strains (PRSP) were used. Results: Alteration in pbp1A was only observed in PISP and PRSP strains, while PCR amplification of the ermB or mefE was observed only in strains with reduced susceptibility to erythromycin. The assay was found to be sensitive as simulated blood cultures showed the lowest level of detection to be 10cfu. Conclusions: As predicted, the assay was able to differentiate penicillin susceptible from the non-susceptible strains based on the detection of the pbp1A gene, which correlated with the MIC value of the strains.

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Mechanisms of Resistance in Multiple-Antibiotic-Resistant Escherichia coli Strains of Human, Animal, and Food Origins

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.10.3996-4001.2004 ◽

2004 ◽

Vol 48 (10) ◽

pp. 3996-4001 ◽

Cited By ~ 269

Author(s):

Yolanda Sáenz ◽

Laura Briñas ◽

Elena Domínguez ◽

Joaquim Ruiz ◽

Myriam Zarazaga ◽

...

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Amino Acid ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Mechanisms Of Resistance ◽

Antibiotic Resistant ◽

E Coli ◽

Class 1 ◽

Human Animal

ABSTRACT Seventeen multiple-antibiotic-resistant nonpathogenic Escherichia coli strains of human, animal, and food origins showed a wide variety of antibiotic resistance genes, many of them carried by class 1 and class 2 integrons. Amino acid changes in MarR and mutations in marO were identified for 15 and 14 E. coli strains, respectively.

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Characterization of the New Metallo-β-Lactamase VIM-13 and Its Integron-Borne Gene from a Pseudomonas aeruginosa Clinical Isolate in Spain

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00465-08 ◽

2008 ◽

Vol 52 (10) ◽

pp. 3589-3596 ◽

Cited By ~ 52

Author(s):

Carlos Juan ◽

Alejandro Beceiro ◽

Olivia Gutiérrez ◽

Sebastián Albertí ◽

Margalida Garau ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Pcr Amplification ◽

Class 1 Integron ◽

Negative Results ◽

New Variant ◽

Class B ◽

Class 1 ◽

Encoding Gene ◽

Good Agreement

ABSTRACT During a survey conducted to evaluate the incidence of class B carbapenemase (metallo-β-lactamase [MBL])-producing Pseudomonas aeruginosa strains from hospitals in Majorca, Spain, five clinical isolates showed a positive Etest MBL screening test result. In one of them, strain PA-SL2, the presence of a new bla VIM derivative (bla VIM-13) was detected by PCR amplification with bla VIM-1-specific primers followed by sequencing. The bla VIM-13-producing isolate showed resistance to all β-lactams (except aztreonam), gentamicin, tobramycin, and ciprofloxacin. VIM-13 exhibited 93% and 88% amino acid sequence identities with VIM-1 and VIM-2, respectively. bla VIM-13 was cloned in parallel with bla VIM-1, and the resistance profile conferred was analyzed both in Escherichia coli and in P. aeruginosa backgrounds. Compared to VIM-1, VIM-13 conferred slightly higher levels of resistance to piperacillin and lower levels of resistance to ceftazidime and cefepime. VIM-13 and VIM-1 were purified in parallel as well, and their kinetic parameters were compared. The k cat/K m ratios for the antibiotics mentioned above were in good agreement with the MIC data. Furthermore, EDTA inhibited the activity of VIM-13 approximately 25 times less than it inhibited the activity of VIM-1. VIM-13 was harbored in a class 1 integron, along with a new variant (Ala108Thr) of the aminoglycoside-modifying enzyme encoding gene aacA4, which confers resistance to gentamicin and tobramycin. Finally, the VIM-13 integron was apparently located in the chromosome, since transformation and conjugation experiments consistently yielded negative results and the bla VIM-13 probe hybridized only with the genomic DNA.

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Salmonella in Indian ready-to-cook poultry: antibiotic resistance and molecular characterization

Microbiology Research ◽

10.4081/mr.2017.6882 ◽

2017 ◽

Vol 8 (1) ◽

Cited By ~ 2

Author(s):

Raj Kamal Gautam ◽

Aarti S. Kakatkar ◽

Manisha N. Karani ◽

Shashidhar R. ◽

Jayant R. Bandekar

Keyword(s):

Antibiotic Resistance ◽

Foodborne Pathogen ◽

Resistance Index ◽

Multi Drug Resistance ◽

High Genetic Diversity ◽

Class 1 Integron ◽

Poultry Products ◽

Class 1 ◽

Salmonella Serovars ◽

Poultry Farming

The availability and popularity of processed, ready-to-cook (RTC) poultry products are increasing in India. Though fresh poultry is known to be contaminated with Salmonella, the prevalence of this foodborne pathogen in RTC poultry products is not reported. Eighty-seven chilled and frozen RTC poultry samples of 4 different brands obtained from supermarkets and departmental stores in Mumbai were analyzed for the presence of Salmonella. The prevalence of Salmonella was higher (51%) in chilled RTC samples as compared to the frozen RTC samples (5%). The frozen RTC samples of one brand were free from Salmonella. S. Typhimurium (75.2%) was the most prevalent serovar, followed by S. Enteritidis (23%) and S. Weltevreden (1.7%). A high percentage (81.4%) of the isolates were found to be resistant to 5 or more antibiotics and class 1 integron, which has been shown to confer multi-drug resistance, was detected in 69.9% of the isolates. Multiple antibiotic resistance index of isolates was high (0.6) indicating the indiscriminate use of antibiotics during poultry farming. High genetic diversity was observed among the Salmonella serovars based on Pulsed Field Gel Electrophoresis profiles. Results showed the presence of multi-drug resistant Salmonella serovars in processed, chilled RTC poultry products marketed in Mumbai, India.

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