Enteroaggregative Escherichia coli in Daycare – A One-Year Dynamic Cohort Study.

ORIGINAL RESEARCH published: 13 July 2016 doi: 10.3389/fcimb.2016.00075 Enteroaggregative Escherichia coli in Daycare—A 1-Year Dynamic Cohort Study Betina Hebbelstrup Jensen 1*, Christen R. Stensvold 1 , Carsten Struve 1 , Katharina E. P. Olsen 1 , Flemming Scheutz 1 , Nadia Boisen 1 , Dennis Röser 1, 2 , Bente U. Andreassen 3 , Henrik V. Nielsen 1 , Kristian Schønning 4 , Andreas M. Petersen 1, 4, 5 † † and Karen A. Krogfelt 1* 1 Department of Microbiology and Infection Control, Statens Serum Institute, Copenhagen, Denmark, 2 Department of Pediatrics, Copenhagen University Hospital Hvidovre, Copenhagen, Denmark, 3 Department of Pediatrics, H.C. Andersen’s Hospital, University of Odense, Odense, Denmark, 4 Department of Clinical Microbiology, Copenhagen University Hospital Hvidovre, Copenhagen, Denmark, 5 Department of Gastroenterology, Copenhagen University Hospital Hvidovre, Copenhagen, Denmark Edited by: Alfredo G. Torres, University of Texas Medical Branch, USA Reviewed by: Glen C. Ulett, Griffith University, Australia Valerio Iebba, “Sapienza” University of Rome, Italy *Correspondence: Betina Hebbelstrup Jensen Karen A. Krogfelt † These authors have contributed equally to this work. Received: 10 April 2016 Accepted: 29 June 2016 Published: 13 July 2016 Citation: Hebbelstrup Jensen B, Stensvold CR, Struve C, Olsen KEP, Scheutz F, Boisen N, Röser D, Andreassen BU, Nielsen HV, Schønning K, Petersen AM and Krogfelt KA (2016) Enteroaggregative Escherichia coli in Daycare—A 1-Year Dynamic Cohort Study. Front. Cell. Infect. Microbiol. 6:75. doi: 10.3389/fcimb.2016.00075 Enteroaggregative Escherichia coli (EAEC) has been associated with persistent diarrhea, reduced growth acceleration, and failure to thrive in children living in developing countries and with childhood diarrhea in general in industrialized countries. The clinical implications of an EAEC carrier-status in children in industrialized countries warrants clarification. To investigate the pathological significance of an EAEC carrier-state in the industrialized countries, we designed a 1-year dynamic cohort study and performed follow-up every second month, where the study participants submitted a stool sample and answered a questionnaire regarding gastrointestinal symptoms and exposures. Exposures included foreign travel, consumption of antibiotics, and contact with a diseased animal. In the capital area of Denmark, a total of 179 children aged 0–6 years were followed in a cohort study, in the period between 2009 and 2013. This is the first investigation of the incidence and pathological significance of EAEC in Danish children attending daycare facilities. Conventional microbiological detection of enteric pathogens was performed at Statens Serum Institute, Copenhagen, Denmark, and at Hvidovre Hospital, Copenhagen, Denmark. Parents completed questionnaires regarding gastrointestinal symptoms. The EAEC strains were further characterized by serotyping, phylogenetic analysis, and susceptibility testing. EAEC was detected in 25 (14%) of the children during the observational period of 1 year. One or more gastrointestinal symptoms were reported from 56% of the EAEC-positive children. Diarrhea was reported in six (24%) of the EAEC positive children, but no cases of weight loss, and general failure to thrive were observed. The EAEC strains detected comprised a large number of different serotypes, confirming the genetic heterogeneity of this pathotype. EAEC was highly prevalent (n = 25, 14%) in Danish children in daycare centers and was accompanied by gastrointestinal symptoms in 56% of the infected children. No serotype or phylogenetic group was specifically linked to children with disease. Keywords: enteroaggregative Escherichia coli, EAEC, cohort study, carrier-state, daycare Frontiers in Cellular and Infection Microbiology | www.frontiersin.org 1 July 2016 | Volume 6 | Article 75 Hebbelstrup Jensen et al. Cohort Study of EAEC INTRODUCTION provided written informed consent prior to enrollment into the study. The study was performed in accordance with the Declaration of Helsinki and was approved by The National Committee on Health Research Ethics in Denmark. Each child was observed for a 1-year period, and the parents were instructed to submit a stool sample from the child and answer a questionnaire every second month. The questionnaires included questions on symptoms including diarrhea (defined as three of more loose stools per day), exposures (e.g., intake of contaminated food or water), contact with another individual suffering from diarrhea, contact to diseased animals, and recent travel, which was defined as any trip outside of Denmark in a period of 2 months prior to sampling. Long-term carriage of EAEC was defined as testing positive for EAEC in stool samples for 6 months or longer, which has been defined similarly in other studies (Richardson et al., 1981; Lübbert et al., 2014; Ismail et al., 2016). Gastrointestinal symptoms reported included diarrhea, abdominal pain, vomiting, nausea, weight loss, and reduced appetite. An episode with the given symptom was registered, when reported by the parents. Enteroaggregative Escherichia coli (EAEC) is an established pathotype within the group of diarrheagenic E. coli (DEC), which also include enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroinvasive E. coli (EIEC), and Shiga toxinproducing E. coli (STEC; Croxen et al., 2013). EAEC has been associated with cases of persistent diarrhea in children living in developing countries (Steiner et al., 1998), traveler’s diarrhea (Adachi et al., 2001), and, most recently, cases of domestically acquired diarrhea in industrialized countries (Jenkins et al., 2006). Large outbreaks of EAEC-associated diarrhea have been reported in England (Dallman et al., 2014), Germany (Bielaszewska et al., 2011), Japan (Itoh et al., 1997), and South Korea (Shin et al., 2015). In the 2011 German outbreak, which affected thousands of Europeans, the EAEC strain had attained a prophage encoding a potent Shiga toxin, enabling the strain to cause hemolytic uremic syndrome, and bloody diarrhea, proving fatal in 54 cases (Bielaszewska et al., 2011). The pathogenic properties of EAEC are under scrutiny, since EAEC can be isolated from asymptomatic carriers as well as from severe cases of diarrhea (Nüesch-Inderbinen et al., 2013; Tobias et al., 2015). EAEC exhibits substantial genetic diversity, and a wide range of virulence factors have been observed (Hebbelstrup Jensen et al., 2014). In addition, manifestations of disease due to EAEC are believed to depend on host factors (Jiang et al., 2003). Few studies have investigated the pathogenic potential of EAEC in the community in industrialized countries. In previous studies carried out in these settings, EAEC has mostly been associated with cases of travelers’ diarrhea (Adachi et al., 2001). A high number of asymptomatic carriers of EAEC has been reported (Nüesch-Inderbinen (...truncated)