Frequent Combination of Antimicrobial Multiresistance and Extraintestinal Pathogenicity in Escherichia coli Isolates from Urban Rats (Rattus norvegicus) in Berlin, Germany Sebastian Guenther 1 *, Astrid Bethe 1 , Angelika Fruth 2 , Torsten Semmler 1 , Rainer G. Ulrich 3 , Lothar H. Wieler 1 , Christa Ewers 1,4 1 Freie Universita ¨t Berlin, Department of Veterinary Medicine, Institute of Microbiology and Epizootics, Berlin, Germany, 2 Robert Koch-Institut, Wernigerode, Germany, 3 Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute for Novel and Emerging Infectious Diseases, Greifswald - Insel Riems, Germany, 4 Justus- Liebig-Universita ¨t Giessen, Department of Veterinary Medicine, Institute of Hygiene and Infectious Diseases of Animals, Giessen, Germany Abstract Urban rats present a global public health concern as they are considered a reservoir and vector of zoonotic pathogens, including Escherichia coli. In view of the increasing emergence of antimicrobial resistant E. coli strains and the on-going discussion about environmental reservoirs, we intended to analyse whether urban rats might be a potential source of putatively zoonotic E. coli combining resistance and virulence. For that, we took fecal samples from 87 brown rats (Rattus norvegicus) and tested at least three E. coli colonies from each animal. Thirty two of these E. coli strains were pre-selected from a total of 211 non-duplicate isolates based on their phenotypic resistance to at least three antimicrobial classes, thus fulfilling the definition of multiresistance. As determined by multilocus sequence typing (MLST), these 32 strains belonged to 24 different sequence types (STs), indicating a high phylogenetic diversity. We identified STs, which frequently occur among extraintestinal pathogenic E. coli (ExPEC), such as STs 95, 131, 70, 428, and 127. Also, the detection of a number of typical virulence genes confirmed that the rats tested carried ExPEC-like strains. In particular, the finding of an Extended- spectrum beta-lactamase (ESBL)-producing strain which belongs to a highly virulent, so far mainly human- and avian- restricted ExPEC lineage (ST95), which expresses a serogroup linked with invasive strains (O18:NM:K1), and finally, which produces an ESBL-type frequently identified among human strains (CTX-M-9), pointed towards the important role, urban rats might play in the transmission of multiresistant and virulent E. coli strains. Indeed, using a chicken infection model, this strain showed a high in vivo pathogenicity. Imagining the high numbers of urban rats living worldwide, the way to the transmission of putatively zoonotic, multiresistant, and virulent strains might not be far ahead. The unforeseeable consequences of such an emerging public health threat need careful consideration in the future. Citation: Guenther S, Bethe A, Fruth A, Semmler T, Ulrich RG, et al. (2012) Frequent Combination of Antimicrobial Multiresistance and Extraintestinal Pathogenicity in Escherichia coli Isolates from Urban Rats (Rattus norvegicus) in Berlin, Germany. PLoS ONE 7(11): e50331. doi:10.1371/journal.pone.0050331 Editor: Stephen V. Gordon, University College Dublin, Ireland Received June 19, 2012; Accepted October 17, 2012; Published November 26, 2012 Copyright: ß 2012 Guenther et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by the Federal Ministry of Education (BMBF) and Research Network Zoonosis (FBI-Zoo, Grant 01KI1012A), the German Research Foundation (DFG) funded Indo-German Research Training Group (Grant GRK1673), and the Federal Ministry of Education and Research grant to RGU (FKZ 01KI1018). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: sebastian.guenther@fu-berlin.de Introduction Brown rats (Rattus norvegicus) are commensal rodents found in urban areas worldwide. They are associated with hygienic problems and are considered a reservoir and vector of several zoonotic pathogens. Indeed, until the twentieth century, one of the most feared diseases related to rats was the plague caused by Yersinia pestis [1,2]. Nowadays, a number of other bacterial, viral and parasitic pathogens have been associated with rats, such as Leptospira spp., Shiga toxin producing E. coli, Campylobacter spp., Salmonella spp., or Hantaviruses [1,3,4,5,6]. There are numerous ways, by which rodent-borne pathogens may infect human and animal hosts. Inhalation of aerosols and consumption of contam- inated food are considered the main pathways, while also direct contact, e.g. by bites, or infections via vectors might occur. Even surface water contaminated with droppings and urine from infected rats in recreational areas has been identified as possible infection source [7]. In addition, specific ecological and behavioral characteristics, e.g. a concentration of Brown rats into high-density populations along with their cohabitation with humans, may further promote the spread of zoonotic pathogens [7]. Another aspect of Brown rats synanthropism is their inhabita- tion of areas near anthropogenically created food sources, such as garbage or sewage systems, also providing harborage [8,9]. Although it is well known that rats live in certain parts of the sewage system [10], even continuous baiting programs have failed to eliminate Brown rat populations [11]. Brown rats from rural areas can roam as far as 260–2000 m within a day, while observational studies in city environments identified smaller activity areas of 25–150 m for rats in urban areas [10]. Nevertheless, urban Brown rats also appear to be able to build PLOS ONE | www.plosone.org 1 November 2012 | Volume 7 | Issue 11 | e50331