Prevalence of malaria infection in Butajira area, south-central Ethiopia

This study was conducted in six kebeles (the smallest administrative units) located about 130 km south of Addis Ababa in the Meskan and Mareko districts of the Gurage Zone in Ethiopia (Figure 1), which is designated as the Butajira area. These six kebeles are the location of the Demographic Surveillance Site (DSS) of the Butajira Rural Health Program (BRHP) [15].

There were 58,335 people living in the BRHP DSS in 2008. Half (50.1%, n = 29,243) of the population were female. Of the total population, 46% lived in the study area. The study area is part of an altitudinal transect between 1,800 and 2,300 masl. The mean annual rainfall in the study area during the last nine years was 945 mm (yearly range 510 mm to 1,329 mm). However, annual rainfall was below average in 2009 and 2010. The main rainy season usually occurs from June to September. The mean temperature was 18.2°C, with average annual minimum and maximum temperatures of 10.0°C and 26.3°C, respectively.

Most people in the area practise subsistence farming. The main cash crops are pepper at low altitude and khat (Catha edulis) in high altitude areas. The main staple food in the highlands is Enset (Ensete ventricosum), and in the lowlands maize, wheat, barley and Teff (Eragrostis tef) are consumed. Animal husbandry is also performed on the farms. The Odamo, Kelakel and Asass rivers flow through the area. The population usually seeks health services at the health posts found in each kebele, three health centres (two at low and one at high altitude) and two hospitals in the area.

Estimation of the sample size for the assessment of malaria prevalence was based on a 4.1% prevalence reported from three Ethiopian regions [16]. Employing assumptions of expected prevalence = 4%, margin of error = 1%, α = 5% (95% confidence level), design effect = 2 and 15% non-response rate, a sample size of 3,393 persons was calculated. Thus, with the assumption of an average family size of 4.5, a total of 3,393 people were recruited from 750 households. The sample size was calculated as n = (1.96)² (0.04) (0.96)/0.01)² = 1,475; total n = (1,472 × 2) + (1,475 × 2 × 0.15) = 2,950 + 443 = 3,393; and 3,393/4.5 = 750.

The study used a community-based prospective design based on repeated cross-sectional surveys. The surveys used a multi-stage random sample design with kebeles as the first-stage unit, villages as the second, and households as the third stage [17].

Six rural kebeles were selected (Hobe, Bati Lejano, Dirama, Shershera Bido, Yeteker and Wurib), with two areas each representing low altitude (1,800-1,899 masl), mid-level (1,900- 1,999 masl) and high altitude (2,000-2,300 masl). The targeted households residing in those kebeles numbered 4,816. Sixteen villages with 750 households were selected randomly from the six kebeles, using probability proportion to size (PPS) sampling. Given that a lower prevalence of malaria was expected in the highlands, more villages and households were selected proportionally. Altitude readings for the households were recorded using a hand-held Global Positioning System (Garmin eTrex^®).

Six cross-sectional surveys were performed on the same households for two consecutive years. The surveys were conducted in October-November 2008 (a month after the main rainy season), January-February 2009 (dry season), June-July 2009 (main rainy season), October- November 2009, January-February 2010, and June 2010. To ensure maximum participation, households with absentees were re-visited once.

Thin and thick blood smears were collected from all family members who consented to the study. Blood specimen processing, examination, and reporting of results were performed using standard guidelines [18]. In brief, the smears were air dried, placed in slide boxes, and examined by a trained microscopist at the field laboratory in Butajira. Thin films were fixed with methanol, and both thin and thick films were stained with 3% Giemsa stain for 30 min. Microscopic examination was done at 1,000× magnification. During the microscopic examination, a slide was regarded as negative after 100 fields had been examined without finding any parasites. Gametocytes were counted against 1,000 leukocytes.

To assure quality of the microscopic examinations, all positive and 10% of the negative slides were re-examined by a second microscopist. In the six surveys, seven of 2,094 slides (0.3%) showed discordant results. The agreement between the first two readers was good (kappa = 0.88). A third reader, blinded to the previous results, re-examined the seven discrepant slides (six vivax malaria and one negative).

A training manual was prepared for the data collectors, who were trained in data collection. Data collection was performed using a pre-tested structured questionnaire and format prepared for this purpose. All data collectors were fluent in the local spoken languages. Data collection in the house-to-house survey was guided by DSS enumerators or the supervisors of each study kebele.

The questionnaire was translated from English to the local language (Amharic) and back to English to check for consistency. All family members who consented to participate in the study were included. The principal investigator and a supervisor from Butajira DSS performed the daily field supervision and cross-checking of the completed questionnaires.

Data entry and cleaning were performed using Epi Info version 6 (Centers for Disease Control and Prevention (CDC), Atlanta, Georgia (USA)). The data were analysed using SPSS version 18.0 (SPSS, Inc., 2009, Chicago). Descriptive statistics were applied to examine the characteristics of the sample; 95% confidence intervals were also calculated. Malaria prevalence was computed by dividing the number of people who showed infection with Plasmodia species by the total number of people examined from the study population. Households with missing values were not considered for analysis.

The study received ethical clearance from the Institutional Review Board of the College of Health Sciences of the Addis Ababa University, and from the Ministry of Science and Technology of Ethiopia. The local and regional health authorities were informed. Individual informed consent was obtained from adults, and from the parents or guardians of children aged less than 18 years. Blood specimens were collected aseptically using an alcohol swab and disposable blood lancets by trained staff. All people found to be malaria positive during the survey were treated directly according to the national guideline [19]. Febrile patients were screened for malaria parasites on the spot using malaria RDTs (CareStart^®), and blood slides were also collected for the survey to investigate the nature of the malaria parasite using light microscopy. Malaria positive cases detected using malaria RDTs were treated immediately. Moreover, those found to be malaria positive on Giemsa-stained light microscopy were also treated. Plasmodium vivax positives were treated with chloroquine, 25 mg/kg for three days (10 mg base per kg on days 1 and 2, and 5 mg base per kg on day 3). Artemether-lumefantrine (20 mg artemether plus 120 mg lumefantrine in a fixed dose combination) was administered, based on body weight, two times a day for three days to Plasmodium falciparum-positive patients. Moreover, Coartem^® and chloroquine stocks were kept at the health posts for treatment of cases that occurred between our repeated surveys.

In the six surveys, 19,207 blood slides were collected during house-to-house visits to 738 households in six kebeles, with an average of 3,780 persons included in each survey. This gave a response rate of 94.4% (n = 19,207/20,339). The remaining 5.6% (1,132 of 20,339) had missing data on different household variables and were not included in the analysis. The households for which data were excluded were not different from other households in any aspect. The average family size was 5.1 (range 1-12). The mean age was 20.5 years, and the median age was 15 years. Table 1 shows the age distribution and other study characteristics.

The overall prevalence of malaria was 0.93% [95% CI (0.79-1.07)] (178 malaria cases among 19,207 people). However, the prevalence varied among the villages, with the highest prevalence of 2.8% in Dadesso and Horosso villages (both below 1,850 masl), and the lowest prevalence (0.0%) at Sunke Wenz and Akababi village (2,100-2,180 masl) (data not presented). Malaria varied with altitude, with a prevalence of 1.91% [95% CI (1.55-2.27)] in low, 1.37% [95% CI (0.87-1.87)] in mid-level and 0.36% [95% CI (0.25-0.47)] in high altitude zones. The highest prevalence was found at low altitude between October and November 2009 (Table 2).

Two persons with P. vivax-positive slides, one from the lowlands and one from the highlands, reported that they had travelled outside their villages. Malaria infection varied among age groups, and in a different way at varying altitudes. At mid-level altitudes, malaria infection reached its peak in children aged one to four years, and at low altitude in children aged one to nine years. However, malaria prevalence at higher altitude was low and was similar across all age groups (Figure 2). Table 3 shows that Plasmodium falciparum malaria occurred rarely throughout the survey periods, with relatively more cases during the survey performed in October-November 2009 in the low altitude zone. Plasmodium vivax was found in all survey periods; however, the prevalence of Plasmodium vivax differed with respect to survey period and altitude (Table 4).