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This is an early access version, the complete PDF, HTML, and XML versions will be available soon.
Study Protocol

Study Protocol for Genomic Epidemiology Investigation of Intensive Care Unit Patient Colonization by Antimicrobial-Resistant ESKAPE Pathogens

by
Andrey Shelenkov
1,*,
Oksana Ni
2,
Irina Morozova
3,
Anna Slavokhotova
1,
Sergey Bruskin
4,
Denis Protsenko
2,
Yulia Mikhaylova
1 and
Vasiliy Akimkin
1
1
Central Research Institute of Epidemiology, Novogireevskaya str., 3a, 111123 Moscow, Russia
2
Moscow Multidisciplinary Clinical Center «Kommunarka», Sosenskiy Stan str., 8с3, 108814 Moscow, Russia
3
Institute for Globally Distributed Open Research and Education (IGDORE), 109651 Moscow, Russia
4
Vavilov Institute of General Genetics, Russian Academy of Sciences, Gubkina str., 3, 119991 Moscow, Russia
*
Author to whom correspondence should be addressed.
Methods Protoc. 2025, 8(6), 151; https://doi.org/10.3390/mps8060151
Submission received: 27 October 2025 / Revised: 5 December 2025 / Accepted: 11 December 2025 / Published: 13 December 2025
(This article belongs to the Section Biomedical Sciences and Physiology)
Review Reports Versions Notes

Abstract

ESKAPE bacteria are a major global threat due to their rapid antibiotic resistance acquisition and severe healthcare-associated infections. Effective countermeasures require epidemiological surveillance and resistance transmission studies, particularly for antimicrobial-resistant (AMR) colonization in intensive care unit (ICU) patients. Whole-genome sequencing (WGS) provides critical information on resistance spread and mechanisms. In the provided protocol, rectal and oropharyngeal swabs, or endotracheal aspirate/bronchoalveolar lavage for intubated patients, are collected at ICU admission and twice weekly. Patient interviews and medical records identify risk factors for resistant microflora. Samples undergo cultivation, species identification, antibiotic susceptibility testing, and DNA extraction. Sequencing is performed using second- and third-generation platforms, with selected isolates subject to hybrid genome assembly. Resistance genes, virulence factors, and typing profiles (MLST, cgMLST) are determined. This protocol characterizes the ICU patient colonization by AMR pathogens, including species distribution, phenotypic and genotypic resistance profiles, clonal structure, and temporal changes. It estimates detection frequency and colonization patterns at each locus, identifies key risk factors, including prior community or inter-facility exposure, and analyzes associations between risk factors and admission colonization. The study aims to estimate AMR infection risk and severity in ICU patients through the comprehensive analysis of colonization dynamics, resistance patterns, and clonal characteristics using WGS data on pathogen composition and AMR trends.
Keywords: antimicrobial resistance; intensive care units; ESKAPE pathogens; bacterial colonization; whole genome sequencing antimicrobial resistance; intensive care units; ESKAPE pathogens; bacterial colonization; whole genome sequencing

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MDPI and ACS Style

Shelenkov, A.; Ni, O.; Morozova, I.; Slavokhotova, A.; Bruskin, S.; Protsenko, D.; Mikhaylova, Y.; Akimkin, V. Study Protocol for Genomic Epidemiology Investigation of Intensive Care Unit Patient Colonization by Antimicrobial-Resistant ESKAPE Pathogens. Methods Protoc. 2025, 8, 151. https://doi.org/10.3390/mps8060151

AMA Style

Shelenkov A, Ni O, Morozova I, Slavokhotova A, Bruskin S, Protsenko D, Mikhaylova Y, Akimkin V. Study Protocol for Genomic Epidemiology Investigation of Intensive Care Unit Patient Colonization by Antimicrobial-Resistant ESKAPE Pathogens. Methods and Protocols. 2025; 8(6):151. https://doi.org/10.3390/mps8060151

Chicago/Turabian Style

Shelenkov, Andrey, Oksana Ni, Irina Morozova, Anna Slavokhotova, Sergey Bruskin, Denis Protsenko, Yulia Mikhaylova, and Vasiliy Akimkin. 2025. "Study Protocol for Genomic Epidemiology Investigation of Intensive Care Unit Patient Colonization by Antimicrobial-Resistant ESKAPE Pathogens" Methods and Protocols 8, no. 6: 151. https://doi.org/10.3390/mps8060151

APA Style

Shelenkov, A., Ni, O., Morozova, I., Slavokhotova, A., Bruskin, S., Protsenko, D., Mikhaylova, Y., & Akimkin, V. (2025). Study Protocol for Genomic Epidemiology Investigation of Intensive Care Unit Patient Colonization by Antimicrobial-Resistant ESKAPE Pathogens. Methods and Protocols, 8(6), 151. https://doi.org/10.3390/mps8060151

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