1887

Fitness of Antibiotic-Resistant Bacteria in the Environment: A Laboratory Activity

    Authors: Massimiliano Marvasi1,*, Manika Choudhury1, Nimisha Binesh Vala1, Max Teplitski2
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    Affiliations: 1: Department of Natural Sciences, Faculty of Science and Technology, Middlesex University, London, UK; 2: Soil and Water Science Department, University of Florida, Gainesville, FL 32603, USA
    AUTHOR AND ARTICLE INFORMATION AUTHOR AND ARTICLE INFORMATION
    • Received 21 October 2016 Accepted 28 January 2017 Published 21 April 2017
    • ©2017 Author(s). Published by the American Society for Microbiology.
    • [open-access] This is an Open Access article distributed under the terms of the Creative Commons Attribution-Noncommercial-NoDerivatives 4.0 International license (https://creativecommons.org/licenses/by-nc-nd/4.0/ and https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode), which grants the public the nonexclusive right to copy, distribute, or display the published work.

    • Supplemental materials available at http://asmscience.org/jmbe
    • *Corresponding author. Mailing address: Department of Natural Sciences, School of Science and Technology, Middlesex University, The Burroughs, London NW4 4BT, UK. Phone: 0044 208 411 4902. E-mail: m.marvasi@mdx.ac.uk.
    Source: J. Microbiol. Biol. Educ. April 2017 vol. 18 no. 1 doi:10.1128/jmbe.v18i1.1257
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    Abstract:

    In this laboratory experiment, we propose an opportunity for students to broaden their understanding of the ecology of antibiotic-resistant and sensitive waterborne bacteria. Antibiotics can be found in rivers or soil as a consequence of agricultural practices or as a result of human use. Concentrations of antibiotics in the environment may range from a few ng to μg L. Such concentrations can affect the selection and fitness of resistant bacteria. In this laboratory activity, students learn how to set up a fitness experiment by using an isogenic pair of antibiotic-resistant and sensitive bacteria in the presence or absence of selective pressure. Microcosms were generated by using filtered river water containing populations of resistant and sensitive bacteria. Competition of both populations was measured in the presence or absence of antibiotics. Students appreciated the use of microcosms for experiments and the extent to which the fitness of resistant and sensitive bacteria changed in the presence and/or absence of a selective pressure in river water. Student learning was measured by using different types of assessments: multiple-choice, true/false, fill in the blanks, laboratory skills observations, and laboratory reports. After the laboratory activity, the percentage of correct answers significantly rose from ~20% to ~85%. Laboratory skills were also evaluated during the exercises, showing no major issues during the experiment. Students showed proficiency in analyzing the complexity of fitness data by reaching a mean of 5.57 (standard error 0.57) over a maximum score of 7 points.

Key Concept Ranking

Environmental Microbiology
0.5207291
Wastewater Treatment Plants
0.42453676
Water Treatment Plants
0.40790185
0.5207291

References & Citations

1. Marti E, Jofre J, Balcazar JL2013Prevalence of antibiotic resistance genes and bacterial community composition in a river influenced by a wastewater treatment plantPLoS One8e7890610.1371/journal.pone.0078906242053473808343 http://dx.doi.org/10.1371/journal.pone.0078906
2. Petrie B, Barden R, Kasprzyk-Hordern B2015A review on emerging contaminants in wastewaters and the environment: current knowledge, understudied areas and recommendations for future monitoringWater Res7232710.1016/j.watres.2014.08.053 http://dx.doi.org/10.1016/j.watres.2014.08.053
3. Huerta B, Rodriguez-Mozaz S, Nannou C, Nakis L, Ruhí A, Acuña V, Sabater S, Barcelo D2016Determination of a broad spectrum of pharmaceuticals and endocrine disruptors in biofilm from a waste water treatment plant–impacted riverSci Total Environ54024124910.1016/j.scitotenv.2015.05.049 http://dx.doi.org/10.1016/j.scitotenv.2015.05.049
4. Roca I, Akova M, Baquero F, Carlet J, Cavaleri M, Coenen S, Cohen J, Findlay D, Gyssens I, Heure OE, Kahlmeter G, Kruse H, Laxminarayan R, Liébana E, López-Cerero L, MacGowan A, Martins M, Rodríguez-Baño J, Rolain J, Segovia C, Sigauque B, Tacconelli E, Wellington E, Vila J2015The global threat of antimicrobial resistance: science for interventionNew Microbes New Infect6222910.1016/j.nmni.2015.02.007260293754446399 http://dx.doi.org/10.1016/j.nmni.2015.02.007
5. Leclercq R, Oberlé K, Galopin S, Cattoir V, Budzinski H, Petit F2013Changes in enterococcal populations and related antibiotic resistance along a medical center-wastewater treatment plant-river continuumAppl Environ Microbiol792428243410.1128/AEM.03586-12233779463623222 http://dx.doi.org/10.1128/AEM.03586-12
6. Pope CF, McHugh TD, Gillespie SH2010Methods to determine fitness in bacteria113121 Gillespie HS, McHugh DTAntibiotic Resistance Protocols, Second EditionHumana PressTotowa, NJ10.1007/978-1-60327-279-7_9 http://dx.doi.org/10.1007/978-1-60327-279-7_9
7. Deng W, Li N, Zheng H, Lin H2016Occurrence and risk assessment of antibiotics in river water in Hong KongEcotoxicol Environ Safety12512112710.1016/j.ecoenv.2015.12.002 http://dx.doi.org/10.1016/j.ecoenv.2015.12.002
8. Marti E, Jofre J, Balcazar JL2013Prevalence of antibiotic resistance genes and bacterial community composition in a river influenced by a wastewater treatment plantPLoS One8e7890610.1371/journal.pone.0078906242053473808343 http://dx.doi.org/10.1371/journal.pone.0078906
9. Andersson DI, Hughes D2010Antibiotic resistance and its cost: is it possible to reverse resistance?Nat Rev Micro8260271
10. Emmert E2013Biosafety guidelines for handling microorganisms in the teaching laboratory: development and rationaleJ Microbiol Biol Educ14788310.1128/jmbe.v14i1.531238583563706168 http://dx.doi.org/10.1128/jmbe.v14i1.531
11. Marvasi M, Choudhury M, Teplitski M2015Laboratory activity to teach about the proliferation of salmonella in vegetablesJ Microbiol Biol Educ1623023610.1128/jmbe.v16i2.948 http://dx.doi.org/10.1128/jmbe.v16i2.948
12. Marvasi M, Davila-Vazquez YC, Martinez LC2013Laboratory activity to effectively teach introductory geomicrobiology concepts to non-geology majorsJ Microbiol Biol Educ1420621210.1128/jmbe.v14i2.578243583843867758 http://dx.doi.org/10.1128/jmbe.v14i2.578
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2017-04-21
2017-05-29

Abstract:

In this laboratory experiment, we propose an opportunity for students to broaden their understanding of the ecology of antibiotic-resistant and sensitive waterborne bacteria. Antibiotics can be found in rivers or soil as a consequence of agricultural practices or as a result of human use. Concentrations of antibiotics in the environment may range from a few ng to μg L. Such concentrations can affect the selection and fitness of resistant bacteria. In this laboratory activity, students learn how to set up a fitness experiment by using an isogenic pair of antibiotic-resistant and sensitive bacteria in the presence or absence of selective pressure. Microcosms were generated by using filtered river water containing populations of resistant and sensitive bacteria. Competition of both populations was measured in the presence or absence of antibiotics. Students appreciated the use of microcosms for experiments and the extent to which the fitness of resistant and sensitive bacteria changed in the presence and/or absence of a selective pressure in river water. Student learning was measured by using different types of assessments: multiple-choice, true/false, fill in the blanks, laboratory skills observations, and laboratory reports. After the laboratory activity, the percentage of correct answers significantly rose from ~20% to ~85%. Laboratory skills were also evaluated during the exercises, showing no major issues during the experiment. Students showed proficiency in analyzing the complexity of fitness data by reaching a mean of 5.57 (standard error 0.57) over a maximum score of 7 points.

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Figures

Image of FIGURE 2

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FIGURE 2

Main findings reported after the three-week laboratory experiment. Box plots represent the fitness of resistant and sensitive bacteria under different concentrations of tetracycline. On the axis, negative value represents sensitive bacteria that are out-competing the resistant bacteria. Positive values represent resistant bacteria outcompeting the sensitive cells. When the ratio is equal the competition index is 0. (A) the fitness of resistant and sensitive bacteria at day 0 and day 2 in the absence of tetracycline. Microcosms were prepared using Thames River water in which the microbial community was removed via filtration. (B) In the presence of tetracycline (selective pressure), resistant bacteria have an increased fitness. Boxes include the lower and upper quartiles, lines within the box are the medians and whiskers indicate the degree of dispersion of the data.

Source: J. Microbiol. Biol. Educ. April 2017 vol. 18 no. 1 doi:10.1128/jmbe.v18i1.1257
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Image of FIGURE 1

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FIGURE 1

Example of workflow of the experiments conducted over the three-week period. PBS = phosphate-buffered saline.

Source: J. Microbiol. Biol. Educ. April 2017 vol. 18 no. 1 doi:10.1128/jmbe.v18i1.1257
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Image of FIGURE 3

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FIGURE 3

Pre-post test assessment LO 1. Percentages of correct answers are shown. The percentage of correct answers was significantly different in all the tests (-test =0.05). Error bars represent standard error. Please refer to Appendix 4 for the typology of questions.

Source: J. Microbiol. Biol. Educ. April 2017 vol. 18 no. 1 doi:10.1128/jmbe.v18i1.1257
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Image of FIGURE 4

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FIGURE 4

Assessment of student performance (LO 2). The scale ranges from very good (5 pts.) to poor (1 pt.). Tasks assigned: A = The student is able to identify contamination on the plates. B = The student can properly patch colonies. C = The student can choose when to use selective plates and non-selective plates. D = The student is able to record the results in the reporting card. E = The student is able to pipette properly.

Source: J. Microbiol. Biol. Educ. April 2017 vol. 18 no. 1 doi:10.1128/jmbe.v18i1.1257
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