Epidemiology of Soil-Transmitted Helminth Infections among Primary School Children in the States of Chhattisgarh, Telangana, and Tripura, India, 2015–2016

ABSTRACT. Soil-transmitted helminth (STH) infections are highly prevalent in many developing countries, affecting the poorest and most deprived communities. We conducted school-based surveys among children studying in first to fifth standard in government schools in the Indian States of Chhattisgarh, Telangana, and Tripura to estimate the prevalence and intensity of STH infections during November 2015 and January 2016. We adopted a two-stage cluster sampling design, with a random selection of districts within each agro-climatic zone in the first stage. In the second stage, government primary schools were selected by probability proportional to size method from the selected districts. We collected information about demographic details, water, sanitation, and hygiene (WASH) characteristics and stool samples from the school children. Stool samples were tested using Kato-Katz method. Stool samples from 3,313 school children (Chhattisgarh: 1,442, Telangana: 1,443, and Tripura: 428) were examined. The overall prevalence of any STH infection was 80.2% (95% confidence interval [CI]: 73.3–85.7) in Chhattisgarh, 60.7% (95% CI: 53.8–67.2) in Telangana, and 59.8% (95% CI: 49.0–69.7) in Tripura. Ascaris lumbricoides was the most prevalent STH infection in all three states. Most of the STH infections were of light intensity. Our study findings indicate that STH infections were highly prevalent among the school children in Chhattisgarh, Telangana, and Tripura, indicating the need for strengthening STH control program in these states. The prevalence estimates from the survey would serve as a baseline for documenting the impact of the National Deworming Day programs in these states.


INTRODUCTION
Ascaris lumbricoides, Trichuris trichiura, and hookworms Necator americanus and Ancylostoma duodenale are the commonest soil-transmitted helminths (STHs), accounting for loss of nearly two million disability-adjusted life years. 1 STH are some of the most common infections contributing heavily to intestinal damage, anemia, and impaired physical growth and cognitive performance in children. 2 Periodic anthelminthic treatment reduces the number of individuals with heavy infections; reduces environmental contamination and risk of infection for other people; reduces micronutrient loss (e.g., iron loss through intestinal bleeding in hookworm infection); and improves nutritional status, cognitive function, and learning ability. 3 School-based deworming programs are considered as simple, safe, cost-effective, and scalable interventions to reach high-risk populations. 4,5 In 2014, the WHO estimated that by number, India has the highest burden of STH infections in the world, with 223 million children aged 1-14 years at risk. 6 Although the published studies indicate heterogenous burden of STH in the country, with prevalence ranging from 0.6% to 91%, with A. lumbricoides as the predominant species, 7 large-scale surveys estimating the prevalence at the state level are limited. Such estimates are required to determine the frequency of preventive chemotherapy. 8 Results of two multi-site statewide surveys in Bihar (N 5 1,279, conducted in 2011) and Uttar Pradesh (N 5 6,421, conducted in 2015) indicated high STH prevalence, ranging between 68% and 76%. 3,9 Besides these two studies, information about STH prevalence and intensity data were not available from other Indian states.
Ad hoc deworming had been conducted in some states in India, often as part of other initiatives, particularly annual mass drug administration (MDA) using a single dose of 400 mg of albendazole in districts where lymphatic filariasis (LF) was endemic, and provision of deworming tablets within the Weekly Iron and Folic Acid Supplementation (WIFS) Program in some areas. 10 Although the reported coverage of MDA was generally higher (more than 80%), 11 there is no information about validated coverage of LF-MDA, and more importantly compliance. In 2014, the Government of India made concerted efforts to scale up STH control activities to meet the WHO global commitment to overcome the impact of neglected tropical diseases. 12 As a key step toward this, cross-sectional, cluster-sampled, school-based surveys were conducted in several States, including Chhattisgarh and Telangana (late 2015), and Tripura (early 2016)-totalling a geographic area of 314,568 km and population of over 67.5 million people. 13 The objective of these surveys was to estimate the prevalence and intensity of STH infections among school-aged children studying in first to fifth standard in these states. The secondary objectives were to estimate the prevalence according to age group, sex, and by agro-climatic zone and to develop geospatial predictive maps of STH prevalence, encompassing the environmental diversity of each state.

MATERIALS AND METHODS
Ethics, consent, and permission. The Institutional Ethics Committee of the Indian Council of Medical Research-National Institute of Epidemiology, Chennai, approved study protocols for each state. Written informed consent from parents of all students assenting to participate in the study was obtained prior to the interviews. In all states, the surveys were conducted prior to MDA. Permission to conduct the survey in schools was obtained from the Department of Health as well as the Department of Education of the respective states.
Study setting. As per the 2011 census, the total population of Chhattisgarh was 25,540,196,Telangana,35,193,978, and Tripura 3,671,032, respectively. 13 The WHO recommends sampling on the basis of agro-climatic zones (regions similar in environmental and climatic conditions), in proportion to the population of these zones. 8 Based on soil type, average annual rainfall, and temperature, Chhattisgarh is divided into three agro-climatic zones: the Bastar plateau consisting of six districts, Chhattisgarh plains consisting of 16 districts, and hilly areas consisting of five districts. 14 Telangana is divided into three agro-climatic zones-the north zone consisting of three districts, central zone consisting of three districts, and south zone consisting of four districts. 15 Tripura state, with eight districts, has only one agro-climatic zone 16 (Figure 1).
Study design and population. To develop state-wide estimates of STH prevalence and intensity, we followed the FIGURE 1. Map of India showing agro-climatic zones and survey locations for the three states. This figure appears in color at www.ajtmh.org.

EPIDEMIOLOGY OF STH INFECTION AMONG SCHOOL CHILDREN
WHO-recommended sentinel site approach. 8 We considered schools as sentinel sites and from each of the selected school, stool specimens from approximately 50 children were investigated for the presence of parasite eggs. The cross-sectional surveys were conducted among primary school children in classes one to five studying in government schools.
A sample size of 492 (rounded to 500) children was required per agro-climatic zone, assuming prevalence of any STH of 20% (chosen as preventive chemotherapy is recommended if STH prevalence is . 20%), absolute precision of 5%, a confidence level of 95%, and design effect of 2. Thus, the total sample size required to estimate the prevalence of STH for each of Chhattisgarh and Telangana states was 1,500, and 500 for Tripura. With 50 children to be sampled from each school, a minimum of 10 schools were needed from each zone. Incorporating a nonresponse of 20%, 60 children from each school were enrolled.
Sampling procedure. A two-stage sampling procedure was followed for selecting schools. In the first stage, districts were randomly selected from each agro-climatic zone. In the second stage, all the government primary schools of the districts selected from each zone were line-listed. Ten schools from each zone were then selected by probability proportional to size from the list of schools in the selected districts. School lists were then examined to determine the student population in each school. In cases where the population of the selected school was less than 60, one nearby additional school was selected to allow for a total of 60 children to be sampled from the area. This additional school was either in the same village/town or in the nearby village/town of the same block (sub-district). Children were then selected from enumerated class lists.
Data collection. The surveys were conducted during November 2015-January 2016. Trained survey teams visited the assigned schools, informed the head of the school about the survey objectives, and obtained permission to conduct the survey. The field teams then enrolled 60 children from the selected class, collected their contact details, entered school characteristics, and recorded the GPS location of the school. The survey teams then visited the houses of enrolled children, explained the purpose of the study, and obtained written informed consent from parents/head of the household. The respondents (mainly the mothers) of the selected children were interviewed to collect information about sociodemographic details, house type, source of drinking water, presence of toilet facilities and their use, hand washing practices, history of deworming, and regular use of footwear. Stool specimen containers were provided and the method of stool collection was explained to the children as well as the respondents. All questionnaires were administered using computer-assisted personal interview devices.
Collection, transport, and processing of stool samples. Single stool samples were collected in the mornings from the houses of the selected children, and transported in cool boxes to the field laboratory within 3-4 hours of sample collection. In the laboratory, samples were kept in cool boxes with ice packs until processed. The field laboratory was set up in the nearest district hospital.
Stool samples were processed using the WHOrecommended Kato-Katz method (Vestergaard Frandsen, New Delhi, India), following the manufacturer's instructions. 8 For each sample, two slides were prepared and independently examined by two parasitologists. The egg counts were used to estimate the prevalence and intensity of STH infections, with intensity of infection measured as eggs per gram of stool, and classified into light, moderate, and heavy infections according to the WHO guidelines. 8 For quality control, 10% of the slides were re-read by exchange between the parasitologists. An independent expert reviewed the laboratory and survey procedures.
Data analysis. Parasitological and questionnaire data were imported into STATA (version 14) and merged to form a dataset linked by child unique identifier. Data were analyzed using the survey data analysis module of STATA to estimate the prevalence and intensity of STH infections in each state after adjusting for clustering. The prevalence was also estimated separately for age group, sex, and different agroclimatic zones. A x 2 test was used to compare proportions. A geographic information system (GIS)-based spatial interpolation-inverse distance weighting method was used to generate predictive maps of the prevalence of STH in the state using the observed prevalence data from the surveyed districts. The locations of the schools surveyed along with the unweighted prevalence of STH infections were integrated into GIS, and ArcGIS v. 10 (ESRI, Redlands, CA) was used for mapping.

RESULTS
The survey was conducted in 95 schools from 26 districts in three states (Table 1). Most schools (n 5 81, 85.3%) were from rural areas, 79 (83.2%) had a source of drinking water, and 82 (86.3%) had toilet facilities within the school    (Table 2).
In all the three states, we did not find any association between the behavioral practices and STH prevalence (data not shown).

DISCUSSION
This paper highlights the STH prevalence and intensity in the Indian states of Chhattisgarh, Telangana, and Tripura. Findings indicate that the overall prevalence of any STH infection among the children studying in government primary schools in all the three states was very high, with about 60-80% of school children infected with one or more STH. The WHO recommends annual treatment in areas where STH prevalence is between 20% and 50%, and biannual treatment in areas with prevalence rates of over 50%. 8 Therefore, according to these guidelines, these states need two rounds of deworming annually to achieve optimal reductions in the prevalence. In 2015, the Government of India launched the National Deworming Day (NDD) targeting children aged 1-19 years, in government schools, government-aided schools, private schools, and anganwadis (preschools). The results of our surveys will serve as a baseline to measure the impact of the NDD program in these states over time.
Among the three states, STH prevalence was highest in Chhattisgarh, with 70-90% of children from each of the different agro-climatic zones infected with STH. In all the three states, STH prevalence was high in both sexes and across all age groups; with 50-80% of children aged 5-7 years positive for any STH. High STH prevalence in this age group indicates that STH infections are acquired during the preschool and early primary school years. The predominance of A. lumbricoides in all the three states is consistent with the findings of previous school-based surveys conducted in the Indian states of Bihar (51.9%) and Uttar Pradesh (69.6%). 3,9 The prevalence of hookworm infection was higher in Chhattisgarh as compared with Telangana and Tripura. Earlier surveys conducted in Bihar and Uttar Pradesh following the similar methodology indicated the prevalence of hookworm infection of 42% and 23%, respectively. The practice of not wearing foot wear, which is a known risk factor for hookworm infection, 17,18 was prevalent among school children in Chhattisgarh (27.6%) and Telangana (37.9%) as compared with Tripura (10.5%), suggesting that the differences in the prevalence could not be solely on account of this behavior. The delay in processing of stool samples lead to faster disintegration of hookworm eggs compared with Ascaris and Trichuris. 19 Since field transportation and laboratory preparedness were carefully monitored during the surveys, our findings suggest heterogeneity of hookworm as well as STH distribution across different states. Although deworming programs are often school-based because of the generally large burden of morbidity in schoolage children, practical ease of accessing the children, as well as cost-effectiveness of school-based deworming, the agespecific prevalence of STH in the three states emphasizes the importance of deworming in preschool children as well. Children of youngest ages have the greatest growth velocities, and hence impact of STH in these formative years is expected to be highly deleterious. 20 The target age group of 1-19 years for the NDD in India is expected to reduce the overall prevalence of STH in preschool as well as school-aged children.
Although the overall STH prevalence was high, a marked feature of all the three state surveys was generally the light intensity of infection. Overall, across any species, only 51 children, or 1.5% of those surveyed, had moderate-to-heavy intensity of infection. Given these states had not received prior large-scale MDA, these findings could possibly indicate that morbidity because of STH in these areas may be low, as it is the heavy-intensity infections that are linked to higher morbidity in populations. 4,21,22 However, it is increasingly becoming recognized that light-intensity infections can also contribute considerably to morbidity in populations. Some studies have shown that even asymptomatic or light-intensity hookworm infections contribute to anemia in preschool and school-aged children. [23][24][25] The clinical appearance of STH infection often lacks specific symptoms, which may not be recognized by the infected person, even when contributing to significant health damage. 26 With prevalence recorded as high as 90% in some areas, a clear conclusion is that STH are a public health problem in these states of India.
The study had certain limitations. First, the survey was conducted in government schools, with no private schools surveyed. Hence, the actual prevalence of STH infections among school children in these states might have been overestimated. Second, although the WHO recommends use of the Kato-Katz method for STH prevalence surveys, the technique is known to have low sensitivity as a diagnostic test especially for light-intensity infections. Future surveys would benefit from alternative diagnostic techniques such as realtime polymerase chain reaction, if these can be costeffectively implemented on a large scale.
There are particular strengths to this study. This is one of the very few epidemiological studies of STH prevalence and infection intensity, at state-wide scale in India. Given the large population size of these states, this is an important addition to the literature internationally as well as nationally. The use of standard STH survey methodology provided one of the most comprehensive large-scale assessments of STH epidemiology thus far available within India.

CONCLUSIONS
These state-wide surveys provide evidence of high prevalence of STH infections, confirming that STH constitute a public health problem in the states of Chhattisgarh, Telangana, and Tripura. In high STH prevalence settings, augmenting the deworming programs with integrated water, sanitation, and hygiene (WASH) activities is necessary for STH control. The Government of India has initiated biannual mass deworming program in all states. The prevalence estimates from the  survey would serve as baseline for documenting the impact of the NDD program in these states in future years. New Delhi, India, for her valuable contributions for the surveys in the three states. We thank the school children for their participation, and parents, teachers, and principals for their support in the conduct of the study. We also wish to thank the state Governments of Chhattisgarh, Telangana, and Tripura for their support in different stages of the survey and the Children's Investment Fund Foundation for providing funding for these surveys. The funder had no role in study design, and decision to publish the manuscript.
Financial support: This work was supported by the Children's Investment Fund Foundation: CIFF.
Disclosure: The Institutional Ethics Committee of the Indian Council of Medical Research-National Institute of Epidemiology, Chennai, approved study protocols for each state. Written informed consent from parents of all students assenting to participate in the study was obtained prior to the interviews.