Padlock Probe-Based Generation of DNAzymes for the Colorimetric Detection of Antibiotic Resistance Genes

The increasing emergence of multidrug- and pan-resistant pathogens requires rapid and cost-efficient diagnostic tools to contain their further spread in healthcare facilities and the environment. The currently established diagnostic technologies are of limited utility for efficient infection control measures because they are either cultivation-based and time-consuming or require sophisticated assays that are expensive. Furthermore, infectious diseases are unfortunately most problematic in countries with low-resource settings in their healthcare systems. In this study, we developed a cost-efficient detection technology that uses G-quadruplex DNAzymes to convert a chromogenic substrate resulting in a color change in the presence of antibiotic resistance genes. The assay is based on padlock probes capable of high-multiplex reactions and targets 27 clinically relevant antibiotic resistance genes associated with sepsis. In addition to an experimental proof-of-principle using synthetic target DNA, the assay was evaluated with multidrug-resistant clinical isolates.

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Molecular identification of antibiotic resistance genes of bacteria isolates from ready to eat food and drinking water sold in Imo State University

Access Microbiology ◽

10.1099/acmi.ac2020.po1007 ◽

2020 ◽

Vol 2 (7A) ◽

Author(s):

Kelechi Dike ◽

Charles Obiukwu ◽

Moses Ekwomadu ◽

Pauline Nnagbo

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Plasmid Dna ◽

Antibiotic Resistance Gene ◽

Antibiotic Resistance Genes ◽

Total Coliform ◽

Control Measures ◽

Coliform Bacteria ◽

State University ◽

Safety Regulations

Food and water are fundamental need of human as its quality is of great concern to the generality of consumers, despite quality control measures during their production, unacceptable microbiological load often makes them unsafe. In this study, we analyzed various ready to eat food and water for presence of coliform bacteria, fungi count, antibiotic resistance gene and plasmid DNA, Samples were found to be positive for various antibiotic resistance genes namely, Sul1, Sul2, Tet A and TetB, interestingly, considerable total coliform bacteria were found from isolates and this result was further confirmed using illumina sequencing of the 16SrRNA gene. All Samples were negative for plasmid DNA, These finding deserve attention as the presence of coliform bacterial and antibiotic resistance genes potentiate health risk to members of university community and as such calls for strigent supervision and saftety and implementation of food safety regulations.

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Full pathogen characterisation: species identification including the detection of virulence factors and antibiotic resistance genes via multiplex DNA-assays

Scientific Reports ◽

10.1038/s41598-021-85438-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Noa Wolff ◽

Michaela Hendling ◽

Fabian Schroeder ◽

Silvia Schönthaler ◽

Andreas F. Geiss ◽

...

Keyword(s):

Antibiotic Resistance ◽

Virulence Factors ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Padlock Probes ◽

Spreading Dynamics ◽

Dna Assays ◽

Cost Efficient ◽

Eskape Pathogens

AbstractAntibiotic resistances progressively cause treatment failures, and their spreading dynamics reached an alarming level. Some strains have already been classified as highly critical, e.g. the ones summarised by the acronym ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter spp.). To restrain this trend and enable effective medication, as much information as possible must be obtained in the least possible time. Here, we present a DNA microarray-based assay that screens for the most important sepsis-relevant 44 pathogenic species, 360 virulence factors (mediate pathogenicity in otherwise non-pathogenic strains), and 409 antibiotic resistance genes in parallel. The assay was evaluated with 14 multidrug resistant strains, including all ESKAPE pathogens, mainly obtained from clinical isolates. We used a cost-efficient ligation-based detection platform designed to emulate the highly specific multiplex detection of padlock probes. Results could be obtained within one day, requiring approximately 4 h for amplification, application to the microarray, and detection.

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Association of nanoparticles with extracellular genetic material and implications for the fate of antibiotic resistance genes in environmental systems

10.1021/scimeetings.0c07368 ◽

2020 ◽

Author(s):

Nadrat Chowdhury

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Genetic Material ◽

Antibiotic Resistance Genes ◽

Environmental Systems

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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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