A retrospective analysis of prevalence of gastrointestinal parasites among school children in the Palajunoj Valley of Guatemala.

This study retrospectively analyzed demographic factors that may affect the prevalence of intestinal parasites among Guatemalan school children. The findings of the study showed that young age, wet season, female gender, and severe malnutrition all correlated positively with increased rates of infection. Clinical visits were performed on 10,586 school children aged 5-15 years over a four-year period (2004-2007) in the Palajunoj Valley of Guatemala, during which 5,705 viable stool samples were screened for infection with the following parasites: Ascaris lumbricoides, Giardia lamblia, Entamoeba histolytica, Hymenolepis nana, and Blastocystis hominis. The average overall prevalences of infection for specific parasites were A. lumbricoides 17.7%, E. histolytica 16.1%, G. lamblia 10.9%, H. nana 5.4%, and B. hominis 2.8%. Statistical analysis showed significantly higher rates of infection among younger children with G. lamblia (odds ratio [OR]=0.905, 95% confidence interval [CI] 0.871-0.941, p<0.0001) and E. histolytica (p=0.0006), greater prevalence of H. nana among females (OR=1.275, CI 1.010-1.609, p=0.0412), higher infection rates during the wet season for E. histolytica (p=0.0003) and H. nana (OR=0.734, CI 0.557-0.966, p=0.0275), and greater rates of infection with G. lamblia among malnourished children (for moderately malnourished children OR=1.498, CI 1.143-1.963, p<0.0001) and E. histolytica (for mildly malnourished children OR=1.243, CI 1.062-1.455, p=0.0313). The results suggest that the prevalence of gastrointestinal parasites among young Guatemalan children is highly dependent on the specific species of the parasite.


INTRODUCTION
Gastrointestinal parasites contribute significantly to global levels of morbidity and mortality. The World Health Organization (WHO) estimates that over two billion people in the world are infected with at least one form of enteropathogen, the majority of whom reside in developing countries and in areas of poor hygiene (1). School-age children are particularly susceptible to parasitosis, often carrying higher burdens of parasites than adults. According to Brooker et al., the greatest obstacle to effective control of parasites in at-risk populations is inadequate knowledge of the geographical distribution of infection and the demographic variables that influence the prevalence of infection (2). While studies to determine the prevalence of gastrointestinal parasites have been conducted in Guatemala city (3) and in other Latin American countries (4)(5)(6)(7)(8)(9)(10)(11), to the best of our knowledge, no study has been undertaken in the Palajunoj Valley or in the Guatemalan Western Highlands.
Nearly half of all Guatemalan children are malnourished, the highest rate among Latin American countries and the fourth highest in the world (12). Poverty in Guatemala is also extreme, with 56% of the population living below the poverty-line and 16% living in extreme poverty (13). Poverty is the greatest among 43% of Guatemalans who are indigenous and who live in rural areas, where 81% of the country's poor and 93% of its extreme poor live. In the Palajunoj Valley, 92.7% of the population is rural, and 95% is indigenous Quiche Maya. The children there, like those living in similar conditions, are vulnerable to increased parasitic infections, malnutrition, stunted development, anaemia, lack of educational opportunities, and child labour (14,15). Aggressive interventions are needed to alleviate the distressed situation in underserved rural areas of Guatemala, such as increased government funding for hospitals and expanded involvement of non-governmental organizations in the health sector.
The health clinic of Primeros Pasos was founded in 2002 to deal specifically with the public-health issues confronting school children in the rural indigenous communities of the Palajunoj Valley. The valley, situated next to the second largest city of Guatemala-Quetzaltenango-is estimated to have 14,481 inhabitants divided among 10 rural communities: Llano del Pinal, Xecaracoj, Las Majadas, Chuicavioc, Tierra Colorada Baja, Tierra Colorada Alta, Xepaché, Candelaria, Bella Vista, and Chuicaracoj. Each year, the clinic invites all children enrolled in each of the 10 community schools to the clinic for a general health screening. Permission was obtained from parents of children to compile health records which have been kept since 2004. This study retrospectively analyzed those records from 2004 to 2007 as these pertain to levels of prevalence of Ascaris lumbricoides, Giardia lamblia, Entamoeba histolytica, Hymenolepis nana, and Blastocystis hominis. These specific parasites were chosen for study due to their status as the five most commonly-diagnosed parasites at the Primeros Pasos clinic for which medication is required. Data on other parasitic species of interest, such as Entamoeba coli, Endolimax nana, and Trichomonas hominis, were not recorded with a sufficient rigueur to validate their reporting here, as those species were deemed to be less important to the daily functioning of the clinic.
A. lumbricoides, G. lamblia, E. histolytica, H. nana, and B. hominis are a significant public-health problem in rural Guatemala and in the developing world as a whole. A. lumbricoides infects an estimated 1.472 billion people worldwide, causing morbidity to 335 million and 60,000 deaths annually (16). The helminth disproportionately impacts the health of children, potentially causing malabsorption of nutrients, loss of appetite, impaired growth, vomiting, anaemia, anorexia, and, in extreme cases, death through intestinal obstructions (17,18). G. lamblia is among the most common intestinal protozoa in the world, infecting more than 200 million people worldwide, although it is markedly more prevalent in developing countries, including Guatemala (19). Infection requires ingestion of as few as 10 viable cysts and is easily spread through contamination of piped water. Painful gas, bloating, fatty diarrhoea, and general epigastric pain are frequently associated with infection. E. histolytica is the major cause of amoebic dysentery in the world, infecting an estimated 480 million people worldwide and causing about 70,000 deaths annually (20). Symptoms of infection include abdominal pain, bloody stools, severe diarrhoea, weight loss, and fatigue, and fatality may occur if the amoeba reaches the liver and is left untreated. The helminth H. nana infects nearly 75 million people worldwide (16). Symptoms of infection include abdominal pain, loss of appetite, itching around the anus, irritability, and diarrhoea. Little is known for certain of the epidemiology of B. hominis, although it is becoming more accepted as a causative agent of disease (21). B. hominis has a worldwide distribution and is most prevalent in areas of low socioeconomic status. Symptoms associated with infection range from abdominal pain and constipation to diarrhoea, vomiting, fatigue, nausea, and fever.
The aim of this study was to identify demographic variables potentially associated with increased rates of gastrointestinal parasite infection among the school children of the Palajunoj Valley of Guatemala. It is hoped that the results of this study will help improve the effectiveness of the Primeros Pasos clinic in designing and promoting its health initiatives in the valley and in aiding other public-health policy-makers active in the Guatemalan Highlands. The data will also contribute to the overall understanding of the epidemiology of A. lumbricoides, G. lamblia, E. histolytica, H. nana, and B. hominis.

Processing of samples
Health workers from the clinic visited the classrooms of students scheduled for clinic-visits one to two days before their appointments to explain the importance of bringing a stool sample and to distribute small plastic containers with instructions on how to gather faeces in the morning of the day of their visit. A volunteer collected stool samples of children at the clinic; stool samples were labelled and taken to the in-house laboratory for analysis. These were processed immediately upon collection using a standard saline wet mount procedure. Samples for which identification of parasites was difficult or uncertain were stained with an iodine solution (1% I 2 and 2% KI) to facilitate analysis. A trained pathologist screened the slides for eggs of A. lumbricoides or H. nana, cysts of E. histolytica, B. hominis, or G. lamblia, trophozoites of G. lamblia, or amoebas of E. histolytica, and the results were recorded on an Excel spreadsheet.

Data processing
The height, weight, age, and sex of each child was measured and recorded. The severity of malnutrition was calculated using the child's weight-for-age ratio, following the guidelines of the World Health Organization (23). Due to the high incidence of stunting among Guatemalan children, weight-forage was deemed to be a more appropriate measure of malnutrition than standard height-for-weight ratios. Children with weight-for-age ratios at 90% or above the mean of the reference population were assigned a level of 'normal' malnutrition while those between 75% and 89.9% were assigned a level of 'mild' malnutrition, 60% to 74.9% were categorized as moderately malnourished, and those at 59.9% or below were considered to be severely malnourished.

Interventions
A qualified physician or a health professional fluent in Spanish physically examined the children. After their check-up, the children attended classes on parasite prevention, hand-washing, dental care, and basic nutrition. Appropriate medications were administered once a year for any parasites found. Children diagnosed with infection due to A. lumbricoides were given a single dose of 400 mg albendazole while those diagnosed with H. nana were given a single dose of niclosamide dependent on bodyweight (1.5 g for children weighing more than 34 kg, 1 g for children weighing between 11 and 34 kg). Children diagnosed with E. histolytica, G. lamblia, and B. hominis were given metronidazole in weight-dependant dosages (for E. histolytica: 35-50 mg metronidazole per kg of body-weight in three doses a day for 7-10 days; for G. lamblia: 15 mg

Statistical analysis
The gathered data were carefully vetted, and incomplete data points were discarded. Logistic regression analyses (using the SAS software, version 9.

Specific parasites
A logistic regression analysis was performed on individual parasitic infection to examine associations between infection and the child's age, gender, and malnutrition status and the season of the year (wet or dry) in which infection occurred. Results for specific parasites are summarized in Table 2 and 3, which present pertinent odds ratios and model parameter estimates respectively.
Higher rates of infection due to G. lamblia were observed among younger children (p<0.0001) and among children with more severe levels of malnutrition (p=0.0009). G. lamblia was one of two

Demographics
The Guatemalan Ministry of Public Health and Social Assistance in 2004 gathered demographic data for each community (22). Workers went from house to house with a questionnaire to gather information on population size and distribution, number of homes per community, disposition of available potable water, and common methods of sewage disposal. For our purposes, 'adequate' sewage disposal was defined as a flush toilet, a latrine, or other defined areas for placing excrement that limited its contact with the surrounding environment and potential for contamination of soil and food with faeces. Table 4 presents a summary of data collected.

Specific parasites
Our findings highlight the epidemiological variabili-   A large number of the same children were seen during the four-year span of the study, with children turning 16 years being excluded and those turning five years included. Patients who received treatment in previous years were not excluded from the study. There is a high likelihood that the children were initially treated successfully for any infections found. Similar multiple doses, 5-10 days of treatments with metronidazole for children with giardiasis have been found to cure infections at a median rate of 94% across eight studies (38). Treatment with albendazole for ascariasis in children of KwaZulu-Natal, South Africa, was 96.4% effective, with re-infection rates of approximately 40% over a 29-week period (39). Parasitic infections in the Palajunoj Valley should be studied in future to determine the incidence of re-infections.
The simple smear and iodine-staining techniques employed for collecting and processing stool samples do not yield data on parasite loads but only on the prevalence of parasites. The methods were chosen for their efficiency and cost-effectiveness but are less sensitive than other commonly-used procedures. Additional techniques, such as formalin-acetate concentration or simple sedimentation, would need to be used for gathering useful data on the relative intensity of infections.
In conclusion, the results of the study suggest that the prevalence of parasitic infections among young Guatemalan children is highly dependent on the parasite species. Young age, wet season, female gender, and severe malnutrition all correlated positively with increased rates of parasitic infection for at least one of the five parasites examined but no single variable was associated with more than two parasites. Malnutrition was associated with increased rates of infection for G. lamblia and E. histolytica only while infection during the wet season was higher for H. nana and E. histolytica only.
Age was significant only in infections due to G. lamblia and E. histolytica, and infection in females was only greater for H. nana. Many of our findings supported those of previous studies, although some did not. Future research may be conducted using more comprehensive diagnostic techniques to determine not only the prevalence of infection but also the intensity and relative parasite loads of the study population.