Fecal Microbiota Functional Gene Effects Related to Single-Dose Antibiotic Treatment of Travelers' Diarrhea

Abstract

Background: Travelers' diarrhea (TD) is common among military personnel deployed to tropical and subtropical regions. It remains unclear how TD and subsequent antibiotic treatment impact the resident microflora within the gut, especially given increased prevalence of antibiotic resistance among enteric pathogens and acquisition of multidrug-resistant organisms. We examined functional properties of the fecal microflora in response to TD, along with subsequent antibiotic treatment.

Methods: Fecal samples from US and UK military service members deployed to Djibouti, Kenya, and Honduras who presented with acute watery diarrhea were collected. A sample was collected at acute presentation to the clinic (day 0, before antibiotics), as well as 7 and/or 21 days following a single dose of antibiotics (azithromycin [500 mg], levofloxacin [500 mg], or rifaximin [1650 mg], all with loperamide). Each stool sample underwent culture and TaqMan reverse transcription polymerase chain reaction analyses for pathogen and antibiotic resistance gene detection. Purified DNA from each sample was analyzed using the HumiChip3.1 functional gene array.

Results: In total, 108 day 1 samples, 50 day 7 samples, and 94 day 21 samples were available for analysis from 119 subjects. Geographic location and disease severity were associated with distinct functional compositions of fecal samples. There were no overt functional differences between pre- and postantibiotic treatment samples, nor was there increased acquisition of antibiotic resistance determinants for any of the antibiotic regimens.

Conclusions: These results indicate that single-dose antibiotic regimens may not drastically alter the functional or antibiotic resistance composition of fecal microflora, which should inform clinical practice guidelines and antimicrobial stewardship.

Clinical trials registration number: NCT01618591.

Keywords: antibiotics; microbiome; travelers’ diarrhea.

Published by Oxford University Press on behalf of Infectious Diseases Society of America 2021.

Figures

Figure 1.

Functional composition differences within the fecal microbiota at acute presentation. Principal component analysis (PCA) and response ratios (RRs) were calculated using all functional probes within the HuMiChip2 array. A, PCA analysis of day 0 samples grouped by country of deployment. B, RRs of significant changes in relative abundance of functional subcategories listed along the right side of the plot between the Kenya and Djibouti samples. The various subcategories are grouped into their parental gene category listed along the top of each subplot. RRs with 95% CIs that do not overlap the vertical dashed line at 0 were considered significantly different between groups. RRs to the right of the dashed line represent a higher relative abundance in Kenya samples, and RRs to the left of the dashed line represent a higher relative abundance in Djibouti samples. C, PCA analysis of day 0 samples grouped by disease impact on activity level. D, RRs of significant changes in relative abundance of functional subcategories between disease-impacted and nonimpacted samples. RRs to the right of the dashed line represent a higher relative abundance in “no impact” samples, and RRs to the left of the dashed line represent a higher relative abundance in “impacted” samples. Grouping by country of deployment and disease impact yielded significantly distinct clusters (multiresponse permutation procedure and Adonis P 

Figure 1.

Functional composition differences within the fecal microbiota at acute presentation. Principal component analysis (PCA) and response ratios (RRs) were calculated using all functional probes within the HuMiChip2 array. A, PCA analysis of day 0 samples grouped by country of deployment. B, RRs of significant changes in relative abundance of functional subcategories listed along the right side of the plot between the Kenya and Djibouti samples. The various subcategories are grouped into their parental gene category listed along the top of each subplot. RRs with 95% CIs that do not overlap the vertical dashed line at 0 were considered significantly different between groups. RRs to the right of the dashed line represent a higher relative abundance in Kenya samples, and RRs to the left of the dashed line represent a higher relative abundance in Djibouti samples. C, PCA analysis of day 0 samples grouped by disease impact on activity level. D, RRs of significant changes in relative abundance of functional subcategories between disease-impacted and nonimpacted samples. RRs to the right of the dashed line represent a higher relative abundance in “no impact” samples, and RRs to the left of the dashed line represent a higher relative abundance in “impacted” samples. Grouping by country of deployment and disease impact yielded significantly distinct clusters (multiresponse permutation procedure and Adonis P 

Figure 2.

Functional differences in the fecal microbiota between pre- and postantibiotic treatment samples by treatment group. Principal component analysis (PCA) and response ratios (RRs) were calculated using all functional probes within the HuMiChip2 array. PCA analysis of (A) day 0, (B) day 7, and (C) day 21. Ellipses within the PCA plots encompass 75% of points within groups (multiresponse permutation procedure and Adonis P > .05). RRs of significant changes in relative abundance of functional subcategories listed along the side of the plot between (D) day 0 and day 7 samples and (E) day 0 and day 21 samples. The various subcategories are grouped into their parental gene category listed along the top of each subplot. RRs with 95% CIs that do not overlap the vertical dashed line at 0 were considered significantly different between groups. RRs to the left of the dashed line represent a higher relative abundance in day 0 samples, and RRs to the right of the dashed line represent a higher relative abundance in (D) day 7 or (E) day 21 samples. PCA and RR colors correspond to the specific antibiotic treatment group. Only matched samples were considered in the analysis (must have provided a sample at each time point compared in the plot). Abbreviations: AZI, azithromycin; LEV, levofloxacin; RIF, rifaximin.

Figure 2.

Functional differences in the fecal microbiota between pre- and postantibiotic treatment samples by treatment group. Principal component analysis (PCA) and response ratios (RRs) were calculated using all functional probes within the HuMiChip2 array. PCA analysis of (A) day 0, (B) day 7, and (C) day 21. Ellipses within the PCA plots encompass 75% of points within groups (multiresponse permutation procedure and Adonis P > .05). RRs of significant changes in relative abundance of functional subcategories listed along the side of the plot between (D) day 0 and day 7 samples and (E) day 0 and day 21 samples. The various subcategories are grouped into their parental gene category listed along the top of each subplot. RRs with 95% CIs that do not overlap the vertical dashed line at 0 were considered significantly different between groups. RRs to the left of the dashed line represent a higher relative abundance in day 0 samples, and RRs to the right of the dashed line represent a higher relative abundance in (D) day 7 or (E) day 21 samples. PCA and RR colors correspond to the specific antibiotic treatment group. Only matched samples were considered in the analysis (must have provided a sample at each time point compared in the plot). Abbreviations: AZI, azithromycin; LEV, levofloxacin; RIF, rifaximin.

Figure 3.

Comparison of gene categories associated with antibiotic resistance in acute vs convalescent stool samples. Response ratios (RRs) showing no changes in the relative abundance of antibiotic resistance gene categories between day 0 and day 21 samples. The various antibiotic resistance gene categories are listed along the left of each plot. A detailed description of each category is given in Supplementary Table 1. RRs with 95% CIs that do not overlap the vertical dashed line at 0 were considered significantly different between groups. RRs to the left of the dashed line represent a higher relative abundance in day 0 samples, and RRs to the right of the dashed line represent a higher relative abundance in day 21 samples. Colors correspond to the specific antibiotic treatment group. Only matched samples were considered in the analysis (must have provided a sample at each time point compared in the plot). Abbreviations: AZI, azithromycin; LEV, levofloxacin; RIF, rifaximin.