Dynamics of intestinal multidrug-resistant bacteria colonisation contracted by visitors to a high-endemic setting: a prospective, daily, real-time sampling study

Abstract

Background: Antimicrobial resistance is highly prevalent in low-income and middle-income countries. International travel contributes substantially to the global spread of intestinal multidrug-resistant Gram-negative bacteria. Hundreds of millions of annual visitors to low-income and middle-income countries are all exposed to intestinal multidrug-resistant Gram-negative bacteria resulting in 30-70% of them being colonised at their return. The colonisation process in high-exposure environments is poorly documented because data have only been derived from before travel and after travel sampling. We characterised colonisation dynamics by exploring daily stool samples while visiting a low-income and middle-income countries.

Methods: In this prospective, daily, real-time sampling study 20 European visitors to Laos volunteered to provide daily stool samples and completed daily questionnaires for 22 days. Samples were initially assessed at Mahosot Hospital, Vientiane, Laos, for acquisition of extended-spectrum β-lactamase-producing (ESBL) Gram-negative bacteria followed by whole-genome sequencing of isolates at MicrobesNG, University of Birmingham, Birmingham, UK. The primary outcome of the study was to obtain data on the dynamics of intestinal multidrug-resistant bacteria acquisition.

Findings: Between Sept 18 and Sept 20, 2015, 23 volunteers were recruited, of whom 20 (87%) European volunteers were included in the final study population. Although colonisation rates were 70% at the end of the study, daily sampling revealed that all participants had acquired ESBL-producing Gram-negative bacteria at some point during the study period; the colonisation status varied day by day. Whole-genome sequencing analysis ascribed the transient pattern of colonisation to sequential acquisition of new strains, resulting in a loss of detectable colonisation by the initial multidrug-resistant Gram-negative strains. 19 (95%) participants acquired two to seven strains. Of the 83 unique strains identified (53 Escherichia coli, 10 Klebsiella spp, and 20 other ESBL-producing Gram-negative bacteria), some were shared by as many as four (20%) participants.

Interpretation: To our knowledge, this is the first study to characterise in real-time the dynamics of acquiring multidrug-resistant Gram-negative bacterial colonisation during travel. Our data show multiple transient colonisation events indicative of constant microbial competition and suggest that travellers are exposed to a greater burden of multidrug-resistant bacteria than previously thought. The data emphasise the need for preventing travellers' diarrhoea and limiting antibiotic use, addressing the two major factors predisposing colonisation.

Funding: The Finnish Governmental Subsidy for Health Science Research, The Scandinavian Society for Antimicrobial Chemotherapy, the Sigrid Jusélius Foundation, Biotechnology and Biological Sciences Research Council; Wellcome Trust, Medical Research Council; The Royal Society; Joint Programming Initiative on Antimicrobial Resistance, and European Research Council.

Figure 1

Study design ESBL=extended-spectrum β-lactamase.

Figure 2

Colonisation of participants by ESBL-producing Gram-negative bacteria Strains are shown by participant, with a maximum of five differing strains identified from the samples of a single individual. Because of the large number of constituent strains in the database, the colour designations do not represent the same strains across multiple volunteers. Days where no symbol is present indicates that a participant was unable to produce a sample. ESBL=extended-spectrum β-lactamase. *Dashed lines represent maintenance of a single strain over multiple days interrupted by colonisation by another strain of the same species or a period of no growth.

Figure 3

Resistance determinants identified in ESBL-producing Gram-negative bacteria isolates The different CTX-M subtypes are represented by the different coloured circles.

Figure 4

Abundance of unique STs in the stool samples of volunteers Isolates were examined using a high-resolution single nucleotide polymorphism analysis. Isolates belonging to the same sequence type were genomically deduplicated to avoid a single persistent strain biasing the results. ST=sequence type.

Figure 5

Linkage of isolates between participants High resolution SNP analysis identified several instances of a single strain colonising multiple participants. Most strains did not contain any variation. 5 of the 33 isolates contained between 1-5 SNPs. The most prolifically shared strain (ST-515) found in participants 5, 6, 17, and 33. Participant 11B is a contact of participant 11, who was found to be colonised by a shared strain. SNP=single nucleotide polymorphism. ST=sequence type. *Isolates that were identical, with no SNPs. †Isolates contained 1–5 SNPs.