Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates

A multiplex assay based on a low-density hydrogel microarray was developed to identify genomic substitutions in <i>N. gonorrhoeae</i> that determine resistance to the currently recommended treatment agents ceftriaxone and azithromycin and the previously used drugs penicillin, tetracyclin...

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Main Authors:	Boris Shaskolskiy, Ilya Kandinov, Dmitry Kravtsov, Alexandra Vinokurova, Sofya Gorshkova, Marina Filippova, Alexey Kubanov, Victoria Solomka, Dmitry Deryabin, Ekaterina Dementieva, Dmitry Gryadunov
Format:	article
Language:	EN
Published:	MDPI AG 2021
Subjects:	hydrogels oligonucleotide microarray DNA hybridization antimicrobial resistance determinants mutations <i>Neisseria gonorrhoeae</i> Organic chemistry QD241-441
Online Access:	https://doaj.org/article/afb8f554e1b64dafaf39ff1d0981580b
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spelling	oai:doaj.org-article:afb8f554e1b64dafaf39ff1d0981580b2021-11-25T18:47:58ZHydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates10.3390/polym132238892073-4360https://doaj.org/article/afb8f554e1b64dafaf39ff1d0981580b2021-11-01T00:00:00Zhttps://www.mdpi.com/2073-4360/13/22/3889https://doaj.org/toc/2073-4360A multiplex assay based on a low-density hydrogel microarray was developed to identify genomic substitutions in <i>N. gonorrhoeae</i> that determine resistance to the currently recommended treatment agents ceftriaxone and azithromycin and the previously used drugs penicillin, tetracycline, and ciprofloxacin. The microarray identifies 74 drug resistance determinants in the <i>N. gonorrhoeae</i> <i>penA</i>, <i>ponA</i>, <i>porB</i>, <i>gyrA</i>, <i>parC</i>, <i>rpsJ</i>, <i>mtrR</i>, <i>bla</i><sub>TEM</sub>, <i>tetM</i>, and 23S rRNA genes. The hydrogel elements were formed by automated dispensing of nanoliter-volume droplets followed by UV-induced copolymerization of NH<sub>2</sub>-containing oligonucleotides with gel-forming monomers. Polybutylene terephthalate plates without special modifications were used as microarray substrates. Sequences and concentrations of immobilized oligonucleotides, gel composition, and hybridization conditions were carefully selected, and the median discrimination ratio ranged from 2.8 to 29.4, allowing unambiguous identification of single-nucleotide substitutions. The mutation identification results in a control sample of 180 <i>N. gonorrhoeae</i> isolates were completely consistent with the Sanger sequencing results. In total, 648 clinical <i>N. gonorrhoeae</i> isolates obtained in Russia during the last 5 years were analyzed and genotyped using these microarrays. The results allowed us to draw conclusions about the present situation with antimicrobial susceptibility of <i>N. gonorrhoeae</i> in Russia and demonstrated the possibility of using hydrogel microarrays to control the spread of antibiotic resistance.Boris ShaskolskiyIlya KandinovDmitry KravtsovAlexandra VinokurovaSofya GorshkovaMarina FilippovaAlexey KubanovVictoria SolomkaDmitry DeryabinEkaterina DementievaDmitry GryadunovMDPI AGarticlehydrogelsoligonucleotide microarrayDNA hybridizationantimicrobial resistance determinantsmutations<i>Neisseria gonorrhoeae</i>Organic chemistryQD241-441ENPolymers, Vol 13, Iss 3889, p 3889 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	hydrogels oligonucleotide microarray DNA hybridization antimicrobial resistance determinants mutations <i>Neisseria gonorrhoeae</i> Organic chemistry QD241-441
spellingShingle	hydrogels oligonucleotide microarray DNA hybridization antimicrobial resistance determinants mutations <i>Neisseria gonorrhoeae</i> Organic chemistry QD241-441 Boris Shaskolskiy Ilya Kandinov Dmitry Kravtsov Alexandra Vinokurova Sofya Gorshkova Marina Filippova Alexey Kubanov Victoria Solomka Dmitry Deryabin Ekaterina Dementieva Dmitry Gryadunov Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
description	A multiplex assay based on a low-density hydrogel microarray was developed to identify genomic substitutions in <i>N. gonorrhoeae</i> that determine resistance to the currently recommended treatment agents ceftriaxone and azithromycin and the previously used drugs penicillin, tetracycline, and ciprofloxacin. The microarray identifies 74 drug resistance determinants in the <i>N. gonorrhoeae</i> <i>penA</i>, <i>ponA</i>, <i>porB</i>, <i>gyrA</i>, <i>parC</i>, <i>rpsJ</i>, <i>mtrR</i>, <i>bla</i><sub>TEM</sub>, <i>tetM</i>, and 23S rRNA genes. The hydrogel elements were formed by automated dispensing of nanoliter-volume droplets followed by UV-induced copolymerization of NH<sub>2</sub>-containing oligonucleotides with gel-forming monomers. Polybutylene terephthalate plates without special modifications were used as microarray substrates. Sequences and concentrations of immobilized oligonucleotides, gel composition, and hybridization conditions were carefully selected, and the median discrimination ratio ranged from 2.8 to 29.4, allowing unambiguous identification of single-nucleotide substitutions. The mutation identification results in a control sample of 180 <i>N. gonorrhoeae</i> isolates were completely consistent with the Sanger sequencing results. In total, 648 clinical <i>N. gonorrhoeae</i> isolates obtained in Russia during the last 5 years were analyzed and genotyped using these microarrays. The results allowed us to draw conclusions about the present situation with antimicrobial susceptibility of <i>N. gonorrhoeae</i> in Russia and demonstrated the possibility of using hydrogel microarrays to control the spread of antibiotic resistance.
format	article
author	Boris Shaskolskiy Ilya Kandinov Dmitry Kravtsov Alexandra Vinokurova Sofya Gorshkova Marina Filippova Alexey Kubanov Victoria Solomka Dmitry Deryabin Ekaterina Dementieva Dmitry Gryadunov
author_facet	Boris Shaskolskiy Ilya Kandinov Dmitry Kravtsov Alexandra Vinokurova Sofya Gorshkova Marina Filippova Alexey Kubanov Victoria Solomka Dmitry Deryabin Ekaterina Dementieva Dmitry Gryadunov
author_sort	Boris Shaskolskiy
title	Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
title_short	Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
title_full	Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
title_fullStr	Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
title_full_unstemmed	Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates
title_sort	hydrogel droplet microarray for genotyping antimicrobial resistance determinants in <i>neisseria gonorrhoeae</i> isolates
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/afb8f554e1b64dafaf39ff1d0981580b
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Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in <i>Neisseria gonorrhoeae</i> Isolates

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