Description of the study area and population
The study was conducted in Adami Tulu District, which is part of the East Showa Zone of the Oromia Regional State in between October 2006 and May 2007. Geographically the area is located between 3820 and 38.55 and 735and 805. The District covers an area of 1403.3km2 (Adami Tulu District agricultural development office report, 2006/07; unpublished report). Ecologically, Adami Tulu is found in the Central Rift Valley of Ethiopia and it is located at 160km away from Addis Ababa to the South Central part and the significant part of the main rift valley lake in the area is Lake Zeway. The relief of the area is characterized by plain and flat stretched land.
A study was undertaken for five months during the major and minor malaria transmission seasons in Aneno Shisho, Kamo Gerbi and Jela Aluto kebeles in the Adami Tulu District. The distance between Aneno Shisho and Kamo Gerbi was 14km, and between Aneno Shisho and Jela Aluto 12km (Figure 1). These kebeles were mainly selected due to the fact that there was a major malaria epidemic in the area since in 1992 (the area, with altitudes ranging from 1,600-1,700 metres, is epidemic prone) [11]. The other reason is about four decades before the time of this study, detailed entomological studies had been conducted in the area and both Anopheles gambiae s.l. and Anopheles pharoensis are known to exist in the area, hence these could be used as baseline information for undergoing this study.
Aneno Shisho has an average altitude of 1,660m above sea level (a.s.l), Kamo Gerbi and Jela Aluto have average altitude of 1,680ma.s.l. and 1,675ma.s.l., respectively. Among the three kebeles; Aneno Shisho received DDT and ITNs (population, 4179), Kamo Gerbi received ITNs only (population, 4100), and Jela Aluto received neither of the DDT and ITNs (population, 4286). Their total population was 12,565, according to a census taken in 1994 (Central Statistical Authority). At each of the three kebeles, the major occupation of the population was dependent on agriculture and livestock herding. Most houses in the three kebeles were circular tukuls with thatched conical roofs and mud or thatch walls. There are a few rectangular houses with the roofs of corrugated metal sheets. Cattles were usually kept in outdoor enclosure made of a loose framework of pests and twigs near the human dwellings and it was observed as the breeding site for the mosquitoes.
The residual treatment applied in Aneno Shisho were DDT 75% water dispersible powder (WDP) used in a spray pump with capacity of eight liters of liquid to obtain the target dosage of 2 gm/m2. Spraying of DDT has been carried out in Aneno Shisho beginning from July 2005 and the second round spraying has been carried out in January 2006. Also, DDT has been used in July 2006 and January 2007, as its residual life span is six months. In Aneno Shisho and Kamo Gerbi an average of two LLINs (PermaNets) per household were first supplied free of cost in January 2005 by Adami Tulu health center. The PermaNet supplied to the inhabitants of Aneno Shisho and Kamo Gerbi was ready-impregnated. It is made of polyester netting material (mesh 25 holes/cm2 with deltamethrin incorporated with 55mg ai/m2) in a resin coating of the fibers [12], which remain effective for three years under normal use.
The area received an annual rainfall of about 4,232.3mm and mean annual rainfall was 705.3mm for six consecutive years (2000-2005). In the year from 2000-2005, mean monthly maximum temperature ranges from 26.3C to 29C, whereas mean monthly minimum temperature ranges from 12.1C to 16.4C (Source: National Meteorological Services Agency of Ethiopia, unpublished data).
The area, like most part of Ethiopia, has two periods of rainfall, June-September, known as the heavy rains and March-April, the small rains. Malaria is the principal cause of morbidity in the area affecting all age groups. The malaria control center of Adami Tulu District is responsible for most case management of uncomplicated malaria, vector-control activities and malaria epidemic control in the area. The major control measures at the time of the survey consisted of indoor residual spraying such as DDT, insecticide-treated nets, anti-malarial drug administration distribution to confirmed cases, but sometimes for presumptive treatment also.
Indoor residual spraying (IRS)
Indoor residual insecticides with DDT have been applied for nearly five decades in Aneno Shisho kebele of Adami Tulu District (Ato Haile Gebre - head of malaria control center of Adami Tulu District, Personal Communication). Residual spraying of DDT has been used in this area in the form of DDT 75% water dispersible powder (WDP). To obtain target dosage of 2 grams of insecticide per m2, 535 grams of DDT 75% WDP are used in a spray pump with capacity of eight liters of liquid and is sprayed from stirrup pumps or hand-compression sprayers. Dichloro-diphenyl-trichloroethane spraying in Aneno Shisho at a dosage of 2g/m2 was carried out twice yearly on the walls and roofs of all houses and domestic animal shelters. The first spraying operations indoors are carried out in June and July, just before the major transmission season and the second round spray are also carried out in January and February, just before the minor transmission season of malaria. This application was carried out since DDT was the residual treatment that remains effective up to six months.
Insecticide-treated bed nets (ITNs)
Promoting the use of insecticide-treated netting materials is one of the principal technical approaches of the Roll Back Malaria (RBM) partnership in the control of malaria. Therefore, in addition to DDT the use of ITNs is the range of malaria control strategies available. In January 2005, an average of two PemaNets per household has been delivered for Aneno Shisho and Kamo Gerbi kebeles free of charge by the Adami Tulu District health center. The nets provided are long-lasting insecticide-treated nets (LLINs) or PermaNets as they eliminate the need for net re-impregnation of conventional nets and is recommended by WHOPES (WHOs pesticide Evaluation Scheme at the Division of Control of Tropical Diseases) for Ethiopia for malaria prevention. This PermaNet is manufactured by Vestergaard Frandsen of Denmark that are treated with a higher dose of wash-resistant deltamethrin, which is said to remain effective for up to 20 washes, or for a period of three years under normal use.
Sampling technique and sample size determination
The survey was conducted based on similar geographical areas and considering the flight range of Anopheles. Three kebeles (Aneno Shisho, Kamo Gerbi, and Jela Aluto) were selected. Among the three kebeles; Aneno Shisho received ITNs and IRS, Kamo Gerbi received ITNs only, and Jela Aluto received neither of the two malaria control measures.
To determine the required sample size for the prevalence study, a malaria prevalence of 39% is assumed. This is based on the result of unpublished report of Adami Tulu Health Centre records from 1999-2006. A sampling error (d) of 0.04 is considered to get a reasonable estimate. The sample size (n) was determined by using the formula for estimating single proportion, where level of significance is 0.05. Accordingly, the minimum sample size was determined to be 571. However, a total of 218 households were selected systematically from the three kebeles, by taking every 5th household from a random start based on their registration list. Two-step sampling was performed to get the required sample size as the study units are the individuals of the selected households. During the first and second survey three individuals per household were sampled for the three kebeles to sample 654 individuals.
Parasitological survey
A blood sample was taken from all selected members of households by experienced laboratory technician by pricking the finger with disposable blood lancet, and thick and thin blood smears were prepared on the same slide and identification numbers marked on the thin films. The thin films were fixed using 100% methanol and then all slides were stained with 3% Giemsa for 20 minutes. Parasite positivity was then determined from thick smear and species identification was carried out from thin smear slide preparations. Each sample was studied by two qualified laboratory technicians in Health Centre and confirmatory examination was carried out by the third technician in the Biomedical Laboratory of Biology Department, Addis Ababa University. During this survey was carried in the seasons of October/November 2006 and April 2007 a verbal consent was obtained for taking blood film from adult participants in sampled households and appropriate anti-malarial treatment was given to positive cases.
Adult mosquito collection
Adult female mosquito collections were made using aspirators and CDC light-trap from indoors and outdoors. Indoor resting mosquito collections from human dwellings were conducted monthly in the three kebeles for five consecutive morning hours (6:00-8:30) between October-December 2006 and April-May 2007. The collections were made by the experienced entomology technician from the local malaria control center and by the investigator of the study in using flash lights and an aspirator and aspirating Anopheline mosquitoes from walls, ceilings and other objects. Mosquitoes were also collected by aspirator from outdoor resting sites (cattle sheds, pit shelters and tree holes). Two dry cell battery-operated CDC light-traps were set indoors in the three sites and operated from 18:00 to 06:00 hours to collect adult mosquitoes. The collected mosquitoes were placed in paper cups and delivered to the temporary examination post in the field. This survey was done by a house-to-house visit with a view to collecting blood samples, Anopheles mosquito collection and gathering demographic information from the members of the households selected with the help of a questionnaire.
Mosquito identification and sporozoite detection
Identification of all adult mosquitoes collected was undertaken using a key [13]. Dried mosquitoes were kept in vials containing silica jell desiccant and ELISA were developed to detect P. falciparum and Plasmodium vivax circumsporozoite proteins in malaria-infected mosquitoes [14]. Anopheline mosquito collection and identification was undertaken by an investigator of the study with an entomology technician from Adami Tulu malaria control center. The detection of sporozoite rates was conducted by the investigator of the study under supervision of a senior entomology technician from the Institute of Ethiopian Health and Nutrition Research.
Knowledge, attitude and practice (KAP) surveys
Malaria control data were collected from February to April, 2007 at the three kebeles of Adami Tulu District using semi-structured interview administered in the local language (Afaan Oromo). A total of 218 household heads as informants were selected systematically from their registration list of each kebele by taking every 5th household from a random start. Of these, informants from Jela Aluto (73), Kamo Gerbi (72), and Aneno Shisho (73) were selected. The semi-structured interview guide is comprised of open-ended and closed questions, and data were obtained on informants knowledge, attitudes and practices regarding the mode of malaria transmission and its prevention and control in the local population.
Ethical considerations
At the beginning of the study, the purpose of the investigation was explained to administrative staff at District and Kebele levels and requested to cooperate. Similarly, from each of household heads, who was selected as representative sample was also asked to consent verbally and participated in the study. Blood smear was obtained with finger prick using disposable blood lancet and cotton immersed in 75% alcohol. During the study period, any kebele inhabitant found sick of malaria was given treatment in collaboration with Adami Tulu District Health Post freely. Approval to conduct the study was granted by the ethical committee in the Department of Biology, Addis Ababa University.
Data analysis
Data collected on parasitological and KAP surveys were managed and analysed using a statistical computer program SPSS version 13.0. The comparisons between kebeles in malaria prevalence, mosquito density and mode of malaria transmission were carried out using the Pearsons chi-square test. A P- value <0.05 was considered to be statistically significant.