Diarrhea Among International Travelers in Bali- Indonesia: Clinical and Microbiological Finding

Diarrhea remains the most common illness affecting travelers.1,2 During their stay abroad, at least one episode of diarrhea occurs in 20%-50% of travelers.3 It is estimated that about 15-20 million travelers experience traveler’s diarrhea annually.4 Risk factors of diarrhea for travelers are varied according to the selected destination, age, behavior, genetic susceptibility, and nationality of the travelers.4,5 The incidence rate of traveler’s diarrhea can range from 5% to 50% depending on the destination. Travelers originating from developing countries have a relatively low incidence rate of traveler’s diarrhea, while travelers from developed countries tend to experience higher incidence rates. The most common bacteria causing these diarrhea cases are Escherichia coli, Shigella, Campylobacter, Salmonella, Aeromonas, Plesiomonas, and non-cholera vibrios.6,7 Traveler’s diarrhea significantly impacts patients not only during the course of their travel, but also following their journey home. These cases may develop into persistent diarrhea with the risk of developing into irritable bowel syndrome.8,9 Bali has a fastgrowing tourism industry and has been named one of the busiest travel destinations in Southeast Asia, with around 6.7 million international visits annually.10 Located in a tropical region, Bali may unsurprisingly have a high prevalence of diarrhea among travelers. Currently, there are no published studies that have focused on the profile of diarrhea among travelers in Indonesia. This study aimed to assess the clinical characteristics and etiologies of diarrhea among travelers http://ijtmgh.com Int J Travel Med Glob Health. 2017 Sep;5(3):84-88 doi 10.15171/ijtmgh.2017.18 TMGH IInternational Journal of Travel Medicine and Global Health J

admitted to Kasih Ibu hospital, Bali.The results of this study will hopefully contribute to a better understanding of the characteristics of diarrhea affecting travelers visiting Bali and serve as a basis for recommendations of future study on reducing the effects of travelers diarrhea.

Study Population/Recruitment
This study was a single site, prospective cross-sectional, hospital-based study conducted at Kasih Ibu hospital in Bali, Indonesia, which is the hospital referred to international travelers to Bali.The patients recruited were adult international travelers who suffered diarrhea during their stay in Bali.Consecutive-sampling was conducted from April 2015 to August 2016.The inclusion criteria were adult travelers (age ≥18 years), admission to Kasih Ibu hospital for diarrhea (frequency ≥3 times a day), and agreement to participate in the study by signing a written informed consent form.Exclusion criteria were incomplete data, insufficient stool samples, and food intolerance diarrhea.A total of 71 international travelers who met the inclusion criteria were included in this study.The patients were required to fill out a standardized epidemiological questionnaire; they were also interviewed about clinical symptoms and asked to provide stool samples.

Definitions
Diarrhea is defined as the passage of three or more watery stools or any number of unformed or loose stools within a 24hour period, with or without one of the following symptoms: nausea, vomiting, abdominal pain or cramps, fever.Diarrhea is classified as chronic when its duration is more than 2 weeks.Cases not fulfilling these criteria were defined as acute diarrhea.Secretory diarrhea was defined as feces with large amount of water, whereas inflammatory diarrhea was defined as loose stool with the presentation of mucus and/or blood in the stool.The dehydration grading was classified as mild, moderate, or severe according to Duggan et al, 11 and the World Health Organization. 12Hyponatremia was indicated by a sodium level less than 135 mEq/L, while hypokalemia was indicated by a potassium level less than 3.5 mEq/L.Bacterial diarrhea in travelers was diagnosed based on the detection of leukocytosis in the stool sample (>5 leukocyte/high-power field) or enteric pathogen in the stool sample accompanied by fever (temperature ≥37.5°C). 13The classification of Entamoebic diarrhea in travelers was based on the detection of the cyst or vegetative form of Entamoeba spp.

Data Collection
Data extracted from the questionnaire included demographic information (name, age, gender, nationality), type of restaurant, and type of food consumed in the previous 2 days.Interviews were also conducted to retrieve information on type and frequency of diarrhea and concomitant symptoms.Physical examinations were performed to assess the severity of diarrhea by evaluating blood pressure, pulse rate, presentation of sunken eyes, skin turgor, vox cholerica, facies cholerica, clammy limbs, and urine production in the previous 6 hours.Blood was drawn to test for cell blood count, blood urea nitrogen, creatinine, transaminases, blood glucose, and electrolyte levels.Fresh stool samples were collected on admission in 2 different containers for stool tests and stool cultures.

Microbiological and Parasite Identification
Microbiological and parasite identification tests were conducted at the Clinical Microbiology Laboratory of Kasih Ibu hospital and Quantum Laboratory.Stool samples were stored at 4°C in Cary-Blair transport medium for bacteria culture examination.Each specimen was inoculated in MacConkey agar selective media and Xylose Lysine Deoxycholate agar media, and then incubated at 37°C for 24-48 hours.API test was used to confirm the presence of Escherichia coli/Enterobacter.Antimicrobial resistance tests used the disc diffusion Kirby-Bauer method.The antimicroba used were ampicillin, sulfamethoxazole-trimethoprim, chloramphenicol, tetracycline, cephalothin, ceftriaxone, colistin, kanamycin, neomycin, erythromycin, nalidixic acid, norfloxacin and ciprofloxacin.

Statistical Analysis
The propotional data was described according to their characteristics and the categorical variables were analyzed using a chi-square test.A P value of less than 0.05 was considered statistically significant.

Clinical Features
Patients in this study experienced an average duration of diarrhea of 3 days.The median frequency of diarrhea was 7 times (within the range of 2-30 times).The type of diarrheal mechanism was mostly secretory (88.7%);only 11.3% of the  2).The distribution of body temperature was skewed to the right with 40.8% of patients having lower than normal (36 o C) body temperature.Patients' blood pressure rates and blood  test results for hemoglobin, platelets, white blood cells, and hematocrit were normal on average.As shown in Table 3, only 37 samples were available for pathogen identification; 37.8% of the samples showed no growth.E. coli was the most commonly isolated enteropathogen in the samples (54.1%).
As an additional analysis, the clinical characteristics of patients with and without Entamoeba spp. in their stools were compared.As summarized in Table 2, 39 fecal samples (54.9%) contained Entamoeba spp.Based on chi-square and Fisher exact test results, it was determined that there were statistically significant differences in the proportion of nausea and vomiting symptoms between patients having Entamoebic spp.and those having no Entamoebic spp (P < 0.05).

Discussion
This is the first report of travelers diarrhea from a tourism destination in Indonesia.This study presents a number of important findings.First, the most predominant age range of cases in this study was young adulthood.This may be attributable to the fact that people in this age range are usually the most adventurous travelers.They may feel more challenged to try local foods and beverages during their stay abroad, thus increasing their risk of contracting gastrointestinal infection.This result is in agreement with those of Ahn et al, 14 and Gascón, 15 who concluded that the age range with the highest prevalence of diarrhea amongst travelers is 20-39 years old, and children and young adults experience more diarrheal episodes than any other age groups.Second, the social economic status of a person is a factor in one's risk of catching this disease.Moreover, being the most adventurous, younger tourists are commonly less well-off economically compared with older tourists; thus, the likelihood they will contract diarrhea is further increased.Third, the results revealed that the most common nationality to contract diarrhea was Dutch, followed by Russian.Dutch and Russian travelers are possibly more susceptible to diarrhea because of their less frequent visits to developing countries.However, the third nationality with the highest number of patients was Australians.Being one of Indonesia's closest neighbors, Australia naturally sends large numbers of tourists to Indonesia every year.The high proportion of Australians visiting Bali contributes to an equally higher propensity of Australian patients to outnumber patients from other countries.
More than 95% of cases in this study were acute diarrhea.It is suspected that the fast onset of diarrhea was caused by enterotoxigenic E. coli (ETEC) and possibly enteroaggregative E. coli (EAEC).Both E. coli strains are known to have fast onset ranging from 8-48 hours.Other studies 2,16,17 have stated that the major causes of diarrhea in travelers are ETEC, Shigella, and Campylobacter.Only a small portion of cases are caused by intestinal parasites such as the Entamoeba species, Giardia species, and Cryptosporidium parvum species. 18Another case report published by Yu and Sohn 19 provided evidence that Cyclospora is a culprit for traveler's diarrhea among tourists visiting Indonesia.To confirm this, microbiological tests were conducted on the stools in this study; E. coli was found to be the most isolated bacteria.The reason behind this might also be related to the high endemicity of this pathogen in developing countries.There were 37 stool samples available for the microbiological test, of which 37.8% showed no growth.Virus might also be incriminated for the diarrhea cases.However, further research is needed to confirm this suspicion as virological testing was not conducted in this study.
The most common type of diarrhea seen was secretory, the mechanism of which is usually exhibited by ETEC.The diarrhea caused by this pathogen is watery and noninflammatory.Non-inflammatory diarrhea could also be caused by the osmotic mechanism carried by the rotavirus. 20his high case rate of non-inflammatory diarrhea is coherent with the microbiological findings of the current study.
The patients in this study mostly experienced cramps, nausea, and vomiting, which normally happen in patients with acute non-inflammatory diarrhea.These clinical symptoms are the manifestations of small intestine disturbance caused by high volume watery stool and dehydration.There were significant differences between electrolyte imbalance caused by hiponatremia and hipokalemia, which may be related to dehydration.
Patients with Entamoeba spp.present in their stools were less likely to experience nausea and vomiting during their diarrheal episodes, and their diarrheal profile was mostly secretory.Classically, the manifestation of diarrhea related to Entamoeba hystolytica is dysentric or bloody diarrhea due to the invasion of this pathogen into the intestinal mucosal.Further investigation is required to assess whether there was any association between immunity in the travelers (nonimmune) and the different clinical symptoms (secretory manifestation) found in this study.Differences in results obtained in this study compared with previously published data may be due to the possibility that other pathogens besides Entamoeba spp were involved in the infection.The great amount of parasites Entamoeba spp. in stool samples of the present study differs from the results obtained by Murphy and Pandey, 21 who reported Campylobacter, ETEC, Shigella, and Cyclospora as the most commonly found pathogens incriminated for causing diarrhea in travelers in Nepal, while the parasite Giardia was a less common cause.Meanwhile, the findings of the current study match those of Peltola and Gorbach 22 and Taylor et al 23 in which protozoans (Giardia and Entamoeba spp.) were the most common pathogen found to cause traveler's diarrhea. 18In fact, no data as of yet published in scientific journals has shown local cases of infection caused by Entamoeba spp.In the current study, 17 of 37 samples showed multiple pathogens present in the stool (data not shown).However, further analysis and a more robust study with a larger data set are required to confirm this.

Conclusion
The most common diarrhea cases among international travelers in Bali was secretory diarrhea without severe complication.E. coli was the main microorganism found in the patients' stool samples.However, Entamoeba spp., which is usually the main culprit for inflammatory diarrhea, was also found among samples with secretory diarrhea.

Study Limitations
This study has some limitations.First, clinical scoring was used to define the levels of dehydration.This type of scoring may give some error related to clinical judging (more subjective) than of using other more objective tools in defining dehydration.Second, the authors could not unreservedly differentiate between the species of Entamoeba spp. as the cause of diarrhea in travelers using only the microscopic test; this test is too sensitive and dependant on the skills of the laboratorist.More sophisticated tools are needed to differentiate between the species of this amoeba.Reverse line blot hybridization assay or polymerase chain reaction may be a better option for future research to confirm the species of the Entamoeba spp. as responsible for diarrhea in travelers.Detection of virus as the cause of diarrhea in travelers may also be needed in future research.

Table 2 .
Clinical Comparison of Patients With Stools Containing Entamoeba spp.and Patients With Non-Entamoeba spp.
a Fisher exact test.b Statistically significant.Mild is the reference for dehydration variable.