<i>Enterobacteriaceae</i> Isolated from the River Danube: Antibiotic Resistances, with a Focus on the Presence of ESBL and Carbapenemases

<div><p>In a clinical setting it seems to be normal these days that a relevant proportion or even the majority of different bacterial species has already one or more acquired antibiotic resistances. Unfortunately, the overuse of antibiotics for livestock breeding and medicine has also altered the wild-type resistance profiles of many bacterial species in different environmental settings. As a matter of fact, getting in contact with resistant bacteria is no longer restricted to hospitals. Beside food and food production, the aquatic environment might also play an important role as reservoir and carrier. The aim of this study was the assessment of the resistance patterns of <i>Escherichia coli</i> and <i>Klebsiella</i> spp. out of surface water without prior enrichment and under non-selective culture conditions (for antibiotic resistance). In addition, the presence of clinically important extended spectrum beta lactamase (ESBL) and carbapenmase harboring <i>Enterobacteriaceae</i> should be investigated. During Joint Danube Survey 3 (2013), water samples were taken over the total course of the River Danube. Resistance testing was performed for 21 different antibiotics. Samples were additionally screened for ESBL or carbapenmase harboring <i>Enterobacteriaceae</i>. 39% of all isolated <i>Escherichia coli</i> and 15% of all <i>Klebsiella</i> spp. from the river Danube had at least one acquired resistance. Resistance was found against all tested antibiotics except tigecycline. Taking a look on the whole stretch of the River Danube the proportion of multiresistances did not differ significantly. In total, 35 ESBL harboring <i>Enterobacteriaceae</i>, 17 <i>Escherichia coli</i>, 13 <i>Klebsiella pneumoniae</i> and five <i>Enterobacter</i> spp. were isolated. One <i>Klebsiella pneumoniae</i> harboring NMD-1 carbapenmases and two <i>Enterobacteriaceae</i> with KPC-2 could be identified. Human generated antibiotic resistance is very common in <i>E</i>. <i>coli</i> and <i>Klebsiella</i> spp. in the River Danube. Even isolates with resistance patterns normally associated with intensive care units are present.</p></div

Proportion of antibiotic resistance.

<p>Proportion of antibiotic resistance.</p

Sampling sites.

<p>Sampling sites.</p

Resistance proportion of <i>Klebsiella</i> spp.

<p>Proportion of <i>Klebsiella</i> spp. with wild type susceptibility pattern (green), resistance to antibiotics out of one or two tested classes (resistant, yellow) and to antibiotics out of three or more classes (multiresistant, red) their total presence in the river and in the three stretches.</p

ESBL and carbapenemase harboring isolates.

<p>ESBL and carbapenemase harboring isolates.</p

JDS 3 overview map.

<p>Overview of the Joint Danube Survey 3 sampling points (JDS3) along the River Danube. Reprinted from Joint Danube Survey Webpage (<a href="http://www.icpdr.org/main/activities-projects/jds3" target="_blank">http://www.icpdr.org/main/activities-projects/jds3</a>) under a CC BY license, with permission from the ICPDR (International Commission for the Protection of the Danube River), original copyright 2013.</p

Resistance proportion of <i>E</i>. <i>coli</i>.

<p>Proportion of <i>E</i>. <i>coli</i> with wild type susceptibility pattern (green), resistance to antibiotics out of one or two tested classes (resistant, yellow) and resistance to antibiotics out of three or more classes (multiresistant, red) their total presence in the river and in the three stretches.</p

Performance Characteristics of qPCR Assays Targeting Human- and Ruminant-Associated <i>Bacteroidetes</i> for Microbial Source Tracking across Sixteen Countries on Six Continents

Numerous quantitative PCR assays for microbial fecal source tracking (MST) have been developed and evaluated in recent years. Widespread application has been hindered by a lack of knowledge regarding the geographical stability and hence applicability of such methods beyond the regional level. This study assessed the performance of five previously reported quantitative PCR assays targeting human-, cattle-, or ruminant-associated <i>Bacteroidetes</i> populations on 280 human and animal fecal samples from 16 countries across six continents. The tested cattle-associated markers were shown to be ruminant-associated. The quantitative distributions of marker concentrations in target and nontarget samples proved to be essential for the assessment of assay performance and were used to establish a new metric for quantitative source-specificity. In general, this study demonstrates that stable target populations required for marker-based MST occur around the globe. Ruminant-associated marker concentrations were strongly correlated with total intestinal <i>Bacteroidetes</i> populations and with each other, indicating that the detected ruminant-associated populations seem to be part of the intestinal core microbiome of ruminants worldwide. Consequently tested ruminant-targeted assays appear to be suitable quantitative MST tools beyond the regional level while the targeted human-associated populations seem to be less prevalent and stable, suggesting potential for improvements in human-targeted methods

Global Distribution of Human-Associated Fecal Genetic Markers in Reference Samples from Six Continents

Numerous bacterial genetic markers are available for the molecular detection of human sources of fecal pollution in environmental waters. However, widespread application is hindered by a lack of knowledge regarding geographical stability, limiting implementation to a small number of well-characterized regions. This study investigates the geographic distribution of five human-associated genetic markers (HF183/BFDrev, HF183/BacR287, BacHum-UCD, BacH, and Lachno2) in municipal wastewaters (raw and treated) from 29 urban and rural wastewater treatment plants (750–4 400 000 population equivalents) from 13 countries spanning six continents. In addition, genetic markers were tested against 280 human and nonhuman fecal samples from domesticated, agricultural and wild animal sources. Findings revealed that all genetic markers are present in consistently high concentrations in raw (median log₁₀ 7.2–8.0 marker equivalents (ME) 100 mL^–1) and biologically treated wastewater samples (median log₁₀ 4.6–6.0 ME 100 mL^–1) regardless of location and population. The false positive rates of the various markers in nonhuman fecal samples ranged from 5% to 47%. Results suggest that several genetic markers have considerable potential for measuring human-associated contamination in polluted environmental waters. This will be helpful in water quality monitoring, pollution modeling and health risk assessment (as demonstrated by QMRAcatch) to guide target-oriented water safety management across the globe