Browsing by Author "Milligan, Erin"

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    Antimicrobial Resistance Monitoring of Water Environments: A Framework for Standardized Methods and Quality Control
    Liguori, Krista; Keenum, Ishi M.; Davis, Benjamin C.; Calarco, Jeanette; Milligan, Erin; Harwood, Valerie J.; Pruden, Amy (American Chemical Society, 2022-06)
    Antimicrobial resistance (AMR) is a grand societal challenge with important dimensions in the water environment that contribute to its evolution and spread. Environmental monitoring could provide vital information for mitigating the spread of AMR; this includes assessing antibiotic resistance genes (ARGs) circulating among human populations, identifying key hotspots for evolution and dissemination of resistance, informing epidemiological and human health risk assessment models, and quantifying removal efficiencies by domestic wastewater infrastructure. However, standardized methods for monitoring AMR in the water environment will be vital to producing the comparable data sets needed to address such questions. Here we sought to establish scientific consensus on a framework for such standardization, evaluating the state of the science and practice of AMR monitoring of wastewater, recycled water, and surface water, through a literature review, survey, and workshop leveraging the expertise of academic, governmental, consulting, and water utility professionals.
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    A framework for standardized qPCR-targets and protocols for quantifying antibiotic resistance in surface water, recycled water and wastewater
    Keenum, Ishi M.; Liguori, Krista; Calarco, Jeanette; Davis, Benjamin C.; Milligan, Erin; Harwood, Valerie J.; Pruden, Amy (Taylor & Francis, 2022-01-16)
    Water environments are increasingly recognized as a conduit for the spread of antibiotic resistance, but there is need to standardize antibiotic resistance monitoring protocols to ensure comparability across studies. Quantitative polymerase chain reaction (qPCR) is attractive as a sensitive means of quantifying antibiotic resistance genes (ARGs) and has been applied broadly over the past two decades to various water matrices. QPCR avoids challenges and biases associated with culture-based methods, providing a reproducible and highly sensitive measure of ARGs carried across a bacterial community. However, there are numerous quality assurance and other aspects of protocols that need to be addressed to ensure that measurements are representative and comparable across studies. Here we conducted a critical review to identify gene targets that are most commonly measured by qPCR to quantify antibiotic resistance in surface water, recycled water, and wastewater and to assess corresponding protocols. Identified targets monitored in water samples included sul1, tetA, and intI1, given their abundance and tendency to correlate with anthropogenic inputs, and vanA and blaCTX-M, as more rarely detected, but highly clinically-relevant targets. We identified 117 peer-reviewed studies meeting search criteria for application of these assays to water matrices of interest and systematically assessed the corresponding protocols, including sample collection and concentration, DNA extraction, primer/probe specificity, amplification conditions, amplicon length, PCR inhibition evaluation, and limit of detection/quantification. Gene copy numbers reported across studies were further compared by assay and water matrix. Based on this comprehensive evaluation, we recommend assays, standardized workflows, and reporting for the five target genes.
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    Towards the standardization of Enterococcus culture methods for waterborne antibiotic resistance monitoring: A critical review of trends across studies
    Davis, Benjamin C.; Keenum, Ishi M.; Calarco, Jeannette; Liguori, Krista; Milligan, Erin; Pruden, Amy; Harwood, Valerie J. (Elsevier, 2022-12-01)
    Antibiotic resistance is a major 21st century One Health (humans, animals, environment) challenge whose spread limits options to treat bacterial infections. There is growing interest in monitoring water environments, including surface water and wastewater, which have been identified as key recipients, pathways, and sources of antibiotic resistant bacteria (ARB). Aquatic environments also facilitate the transmission and amplification of ARB. Enterococcus spp. often carry clinically-important antibiotic resistance genes and are of interest as environmental monitoring targets. Enterococcus spp. are Gram-positive bacteria that are typically of fecal origin; however, they are also found in relevant environmental niches, with various species and strains that are opportunistic human pathogens. Although the value of environmental monitoring of antibiotic-resistant Enterococcus has been recognized by both national and international organizations, lack of procedural standardization has hindered generation of comparable data needed to implement integrated surveillance programs. Here we provide a comprehensive methodological review to assess the techniques used for the culturing and characterization of antibiotic-resistant Enterococcus across water matrices for the purpose of environmental monitoring. We analyzed 117 peer-reviewed articles from 33 countries across six continents. The goal of this review is to provide a critical analysis of (i) the various methods applied globally for isolation, confirmation, and speciation of Enterococcus isolates, (ii) the different methods for profiling antibiotic resistance among enterococci, and (iii) the current prevalence of resistance to clinically-relevant antibiotics among Enterococcus spp. isolated from various environments. Finally, we provide advice regarding a path forward for standardizing culturing of Enterococcus spp. for the purpose of antibiotic resistance monitoring in wastewater and wastewater-influenced waters within a global surveillance framework.