Natural History of Enteroaggregative and Enterotoxigenic Escherichia coli Infection among US Travelers to Guadalajara, Mexico
- Javier A. Adachi1,
- Charles D. Ericsson1,
- Zhi-Dong Jiang1,
- Margaret W. DuPont1,2,
- Sanjay R. Pallegar1 and
- Herbert L. DuPont1
- 1Center for Infectious Diseases, University of Texas, Houston Medical School and School of Public Health, Houston, Texas
- 2St. Luke's Episcopal Hospital and Baylor College of Medicine, Houston, Texas
- Reprints or correspondence: Dr. Herbert L. DuPont, St. Luke's Episcopal Hospital, Internal Medicine Service, 6720 Bertner Ave., MC 1-164, Houston, TX 77030 (hdupont{at}sleh.com).
-
Presented in part: 49th annual meeting of the American Society of Tropical Medicine and Hygiene, Houston, 1 November 2000 (abstract 553).
Abstract
The natural history of enteroaggregative Escherichia coli (EAEC) and enterotoxigenic E. coli (ETEC) infection was studied among 40 US travelers who provided weekly stool samples for 4 weeks after arrival in Mexico. At enrollment, 5 subjects were colonized by EAEC and 3 by ETEC. During the first 2 weeks after enrollment, 12 developed EAEC diarrhea, 7 developed ETEC diarrhea (5 with mixed EAEC/ETEC diarrhea), 13 had EAEC colonization, and 7 had ETEC colonization. During the third and fourth weeks, 4 experienced EAEC diarrhea, 2 experienced ETEC diarrhea (1 with mixed EAEC/ETEC diarrhea), 31 had EAEC colonization, and none had ETEC colonization. Plasmid DNA analysis showed a high degree of heterogeneity among EAEC isolates. Symptomatic EAEC infection occurred early after arrival in Guadalajara, Mexico, and was as common as ETEC infection. Asymptomatic EAEC infection was recurrent.
Enterotoxigenic Escherichia coli (ETEC) is considered to be the principal cause of traveler's diarrhea. Enteroaggregative E. coli (EAEC), which shows a characteristic (“stacked-brick”) aggregative adherence pattern to HEp-2 cells [1], has been implicated as an etiologic agent in acute traveler's diarrhea [2, 3]. In this prospective study, we studied the natural history of both EAEC and ETEC infection in US travelers during the 4 weeks after arrival in Guadalajara, Mexico.
Materials and Methods
The population comprised US college students who attended summer educational sessions in Guadalajara, Mexico (June to August 1999). Subjects were eligible if they were ⩾18 years of age and were enrolled in the study within the first 3 days after arriving in Mexico. Exclusion criteria included travel to a developing region of the worldwithin the 3 months prior to enrollment or administration of an antimicrobial agent with expected activity against bacterial enteric pathogens within the week previous to enrollment.
After signing a consent form, subjects completed daily diaries that detailed stool form and frequency and other clinical signs or symptoms, and they were instructed to report if diarrhea occurred. Diarrhea was defined as the passage of at least 3 unformed stools in 24 h with at least 1 additional clinical manifestation of enteric disease (nausea, vomiting, abdominal cramps or pain, excessive intestinal gas, tenesmus, fecal urgency, or dysentery).
Four stool samples were obtained from each patient: the first was obtained at enrollment, and then a weekly stool sample was obtained for 3 consecutive weeks. If subjects developed diarrhea, additional stool specimens were collected during the illness. All stool samples were tested for conventional enteric pathogens [4]. Five E. coli—like colonies from MacConkey agar plates from each stool sample were transported in individual peptone stabs to Houston and were tested for the presence of ETEC by use of DNA hybridization [5] and for EAEC by use of HEp-2 cell adherence assay [6]. H10407 and JPN 042 strains were used as quality controls.
The ratio of number of cases of symptomatic infection to asymptomatic infection was examined by week of study. For subjects with mixed EAEC/ETEC infection, the 2 diarrheagenic E. coli were considered separately.
Plasmid DNA analysis was done to determine whether all infections were caused by similar or different strains and whether the sequential infections represented reinfection or chronic infection. Plasmid analysis was prepared from EAEC isolates by use of the Wizard plus minipreps DNA purification system (Promega) [7]. Strain bands were treated as binary variableswith 2 possible outcomes: presence or absence of the band. Similarity between 2 strains was calculated using values between 0 and 1 to indicate degree of resemblance (0 indicated that the strains had no similarity, and 1 indicated maximum similarity).
Results
Of the 40 subjects who were enrolled in this study, 28 provided 4 routine weekly stool samples, and 12 provided 3 routine weekly stool samples. Thirty-seven subjects provided routine stool samples during the second week, 35 during the third week, and 32 during the last week of the study. All subjects were enrolled during the first 3 days after arrival in Mexico.
At enrollment, 5 of 40 subjects were colonized by EAEC. An additional subject developed EAEC diarrhea 2 days after arrival in Mexico. Three subjects were colonized by ETEC at enrollment. During the first 2 weeks, 13 subjects had EAEC colonization in their routineweekly stool samples, and 7were colonized by ETEC. A total of 14 subjects developed diarrhea during the first 2 weeks: 12 experienced EAEC diarrhea (5 with mixed EAEC/ ETEC infection and 1 with EAEC and Salmonella infections), and 7 experienced ETEC diarrhea (which included the 5 with mixed EAEC/ETEC infection). For weeks 1 and 2, the ratio of number of subjects with diarrhea to total number of subjects colonized with EAEC was 0.8 and 0.9, respectively, which was similar to the ratio found for ETEC infection (0.7 and 1.1, respectively) during the same period (table 1).
During weeks 3 and 4, the number of subjects with asymptomatic EAEC colonization increased to 31. During the same 2 weeks, the number of cases of diarrhea secondary to EAEC and ETEC dropped to 4 and 2 (including 1 case of mixed EAEC/ ETEC infection), respectively, and the ratio of number of cases of diarrhea to number of subjects colonized decreased to 0.1 for EAEC. There were no subjects with ETEC colonization (table 1).
During the total 4 weeks of the study, 27 (68%) subjects became infected with EAEC, with an almost constant number of patients infected per week (7, 6, 7, and 7 persons each of the 4 weeks). During the 4 weeks, 25 (63%) of the 40 enrolled subjects had 30 episodes of diarrhea. Sixteen (53%) of 30 diarrhea episodes were associated with EAEC infection, 9 (30%) were associated with ETEC infection (6 of both were associated with mixed EAEC/ETEC infection), and 11 (37%) episodes occurred without a recognized pathogen. None of the patients with EAEC diarrhea had a repeat episode of EAEC diarrhea.
One subject was asymptomatically infected by EAEC for each of the 4 weeks of the study, 4 subjects had 3 weekly samples positive for EAEC but did not have symptoms of infection, 6 subjects had 2 weekly stool samples positive for EAEC but did not have symptoms, and 9 had 1 weekly sample positive for EAEC but did not have clinical illness. Five of the subjects with initial asymptomatic EAEC infection subsequently developedEAEC diarrhea. None of the cases of ETEC diarrhea were preceded by asymptomatic ETEC infection.
Plasmid DNA analysis of the 60 EAEC isolates obtained revealed that there were 8 clusters at a similarity of 0.2 (minimum similarity). Among the 16 cases of EAEC diarrhea, 12 (75%) had isolates with similarity values ⩽0.2, and the remaining 4 had similarity values >0.2. Of the 11 subjects with ⩾2 sequential EAEC infections, 8 (73%) had isolates with similarity values ⩽0.2.
Of the 9 ETEC isolates from cases of diarrhea, 6 were heatstable enterotoxin (ST), 2 were heat-labile enterotoxin (LT), and 1 was ST/LT. For the 7 ETEC isolates from persons without symptoms, the breakdown of toxin types was 2 ST, 5 LT, and 0 ST/LT. There was no recognized clinical difference in cases of ETEC diarrhea by toxin type, but the number of ETEC cases in each group was small.
Discussion
ETEC strains have been implicated in a number of studies as the major definable cause of traveler's diarrhea [1, 4]. EAEC strains, also implicated as important causes of traveler's diarrhea [2, 3], have been shown to cause persistent diarrhea and malnutrition in children [1] and chronic diarrhea in patients with AIDS [8].
The current study confirms the common finding of EAEC and ETEC infection in Guadalajara [2, 7]. In the present, relatively small study, we found that EAEC was a more common cause of traveler's diarrhea than ETEC. Our study revealed that infections with EAEC and ETEC were rapidly acquired after arrival in Mexico, with 6 subjects infected with EAEC and 3 with ETEC within the first 3 days. Others have found the common occurrence of asymptomatic EAEC infection among international tourists [9]. We found a high level of contamination by EAEC and ETEC in food samples in Guadalajara, which probably explains the early infection [10, 11].
In this study, the number of subjects with EAEC colonization increased over time, whereas the number of cases of EAEC diarrhea decreased with the length of stay. During the first 2 weeks, subjects with EAEC diarrhea and EAEC colonization were more frequently identified than those with ETEC infection. However, the ratio of number of cases of diarrhea to number of asymptomatic subjects by week was found to be similar for both pathogens. During the last 2 weeks, the ratio of diarrhea to asymptomatic infection for EAEC dropped 8-fold, compared with the first week of the study, in which the number of subjects with asymptomatic EAEC colonization doubled in frequency. This suggests that immunity to symptomatic EAEC diarrhea may have occurred early but that resistance to asymptomatic enteric infection did not occur.
The results of the plasmid DNA analysis revealed a high degree of heterogeneity among the infecting EAEC isolates and among the strains from cases with sequential infections, indicating the presence of multiple strains circulating in the population that lead to recurrent infections. This study and others [12–15] provide data to indicate that EAEC strains represent a group of heterogeneous, diarrheagenic E. coli that share the distinctive aggregative adherence pattern to HEp-2 cells but differ by the virulence properties that they possess, as well as by their pulsed-field gel electrophoresis [3] and DNA plasmid content.
The unique aspect of the present study is that a cohort of US adults was followed longitudinally during 4 weeks after entering a high-risk area for EAEC and ETEC infection. Although the prevalence of ETEC infection, with or without diarrhea, by week was lower than that for EAEC, the ratio of number of subjects with diarrhea to number of asymptomatic subjects was similar for both enteropathogens during the first 2 weeks in Mexico. Only 1 subject with ETEC diarrhea was identified during weeks 3 and 4 of the study, indicating the development of anti-ETEC immunity. EAEC appears to show greater heterogeneity than do ETEC strains, probably explaining the chronic and recurrent EAEC infections in our study population.
This prospective observational follow-up study has provided evidence that EAEC infection is prevalent among travelers to Mexico, with asymptomatic colonization occurring more frequently than EAEC diarrhea. Both EAEC and ETEC infection were shown to be common during the first 2 weeks of stay in Mexico, with frequent development of asymptomatic EAEC infections and reinfections during a 4-week stay.
Figures and Tables
Table 1.
Subjects with diarrhea and asymptomatic subjects who have culture-proven enteroaggregative Escherichia coli (EAEC) and enterotoxigenic E. coli (ETEC) infection, by week of study.
Acknowledgments
We thank the members of the field and laboratory teams who carried out this clinical study: Monica A. Gonzalez, Marian C. Holland, Mimi Hu, David B. Huang, Wei Li, Jo Ann Lee, M. Joanna Lyman, Kenneth J.Maverick, Shannon M. Ohnheiser,Greg M. Press, Carmen Pulido, Dorothy Ruelas, Corey S. Scurlock, and Jaqueline Vaca.
Footnotes
-
Written consent was obtained from each patient, and guidelines from the Committee for Protection of Human Subjects of the University of Texas Health Science Center at Houston were followed.
- Received June 25, 2001.
- Revision received January 22, 2002.
- © 2002 by the Infectious Diseases Society of America
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