Teshome Gebre-Michael

Malaria is a public health problem in Ethiopia and its transmission is generally unstable and seasonal. For the selection of the most appropriate vector control measures, knowledge on the ecology of the vector is necessary at a local level. Therefore, the objectives of this study were to document the species composition, breeding habitat characteristics and occurrence of anopheline larva in Sheni stream and the vectorial role of the prevailing Anopheles in relation to malaria transmission in Addis Zemen, Ethiopia. Immature anophelines were sampled from breeding habitats and characteristics, such as water temperature, turbidity, water current, water pH and other variables, of the habitats were measured from October 2011 to February 2012. Adult anophelines were sampled inside human dwellings using space spray and Center for Disease Control light traps. Artificial pit shelters and clay pots were also used for outdoor adult collections. Anophelines collected were identified using morphological key. The enzyme-linked immunosorbent assay was applied to detect circumsporozoite proteins of Plasmodium and source of blood meals. A total of 6258 Anopheles larvae were collected and identified morphologically. Five anopheline species were found: An. gambiae (s.l.), An. cinereus, An. demeilloni, An. christi and An. pretoriensis. Anopheles gambiae (s.l.) existed in most of the habitats investigated. Only the former three species were captured in the adult collections. Sun-lit Sheni stream, rain pools, hoof prints, drainage and irrigation canals were found to be habitats of larvae. Anopheles gambiae (s.l.) larvae were most abundantly sampled from sand mining and natural sand pools of Sheni stream. Multiple regression analysis showed that clear, permanent and temporary habitats devoid of mats of algae were the best predictors of An. gambiae (s.l.) larval abundance. It is also the responsible malaria vector in the study area and exhibits anthropophilic and endophagic behaviour. The malaria vector An. gambiae (s.l.) was found in Addis Zemen throughout the study period from both adult and larval collections. Sheni stream is the main larval habitat responsible for the occurrence of anopheline larvae during the dry season of the study area when other breeding sites perish.

Leishmania donovani, the causative agent of visceral leishmaniasis (VL), is most probably vectored by Phlebotomus orientalis in north-western Ethiopia. The aim of this study was to determine and compare the nocturnal activity patterns of Ph. orientalis in VL endemic foci of Libo-Kemkem (highland) and Metema (lowland) districts of north-western Ethiopia. Sampling of sand flies was conducted bimonthly from January-May 2012 in the highland and from March-June 2013 in the lowland. Sand flies were sampled using two CDC light traps placed in compounds occupied by both cattle and humans. Sampling of sand flies started at 18:00 h and ended at 06:00 h. Every hour, a cage was replaced by another cage. In total, 9479 nocturnally active sand flies were collected from both study areas. Six Phlebotomus species (Ph. orientalis, Ph. duboscqi, Ph. papatasi, Ph. bergeroti, Ph. rodhaini and Ph. martini) and several Sergentomyia spp. were identified. In both areas, of the six Phlebotomus spp., Ph. orientalis was the preponderate. In the highland, the hourly activity pattern of Ph. orientalis females was higher before midnight with a peak in density between 22:00–23:00 h, whereas in the lowland after midnight between 03:00–04:00 h. The present study showed that Ph. orientalis females exhibited different nocturnal activity patterns with a peak in the early part of the night in the highland and in the latter part of the night in the lowland areas. As the risk of acquiring L. donovani infections vary in the two areas, appropriate control strategies should be developed according to the activity of Ph. orientalis in the respective areas.

Background: The human landing catch (HLC) is the standard reference method for measuring human exposure to mosquito bites. However, HLC is labour-intensive, exposes collectors to infectious mosquito bites and is subjected to collector bias. These necessitate local calibration and application of alternative methods. This study was undertaken to determine the relative sampling efficiency (RSE) of light traps with or without yeast-produced carbon dioxide bait vs. HLC in south-central Ethiopia. Methods: The experiment was conducted for 39 nights in a 3 × 3 Latin square randomized design with Anopheles arabiensis as the target species in the period between July and November 2014 in Edo Kontola village, south-central Ethiopia. Center for Disease Control and Prevention light trap catches (LTC) and yeast-generated carbon dioxide-baited light trap catches (CB-LTC) were each evaluated against HLC. The total nightly mosquito catches for each Anopheles species in either method was compared with HLC by Pearson correlation and simple linear regression analysis on log-transformed [log₁₀(x + 1)] values. To test if the RSE of each alternative method was affected by mosquito density, the ratio of the number of mosquitoes in each method to the number of mosquitoes in HLC was plotted against the average mosquito abundance. Results: Overall, 7606 Anopheles females were collected by the three sampling methods. Among these 5228 (68.7%) were Anopheles ziemanni, 1153 (15.2%) An. arabiensis, 883 (11.6%) Anopheles funestus s.l., and 342 (4.5%) Anopheles pharoensis. HLC yielded 3392 (44.6%), CB-LTC 2150 (28.3%), and LTC 2064 (27.1%) Anopheles females. The RSEs of LTC and HLC for An. arabiensis were significantly correlated (p < 0.001) and density independent (p = 0.65). However, for outdoor collection of the same species, RSEs of LTC and CB-LTC were density dependent (p < 0.001). It was estimated that on average, indoor LTC and CB-LTC each caught 0.35 and 0.44 times that of indoor HLC for An. arabiensis respectively. Conclusions: Results showed that HLC was the most efficient method for sampling An. arabiensis. LTC can be used for large-scale indoor An. arabiensis surveillance and monitoring when it is difficult to use HLC. CB-LTC does not substantially improve sampling of this major vector compared to LTC in this setting. Trial registration PACTR201411000882128 (retrospectively registered 8 September, 2014)

Indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs) are the key malaria vector control interventions in Ethiopia. The success of these interventions rely on their efficacy to repel or kill indoor feeding and resting mosquitoes. This study was undertaken to monitor human-biting patterns of Anopheles species in south-central Ethiopia. Human-biting patterns of anophelines were monitored for 40 nights in three houses using human landing catches (HLC) both indoors and outdoors between July and November 2014, in Edo Kontola village, south-central Ethiopia. This time coincides with the major malaria transmission season in Ethiopia, which is usually between September and November. Adult mosquitoes were collected from 19:00 to 06:00 h and identified to species. Comparisons of HLC data were done using incidence rate ratio (IRR) calculated by negative binomial regression. The nocturnal biting activities of each Anopheles species was expressed as mean number of mosquitoes landing per person per hour. To assess malaria infections in Anopheles mosquitoes the presence of Plasmodium falciparum and P. vivax circumsporozoite proteins (CSP) were determined by enzyme-linked immunosorbent assay (ELISA). Altogether 3,408 adult female anophelines were collected, 2,610 (76.6 %) outdoors and 798 (23.4 %) indoors. Anopheles zeimanni was the predominant species (66.5 %) followed by An. arabiensis (24.8 %), An. pharoensis (6.8 %) and An. funestus (s.l.) (1.8 %). The overall mean anopheline density was 3.3 times higher outdoors than indoors (65.3 vs 19.9/person/night, IRR: 3.3, 95 % CI: 1.1–5.1, P = 0.001). The mean density of An. zeimanni, An. pharoensis and An. funestus (s.l.) collected outdoors was significantly higher than indoors for each species (P < 0.05). However, the mean An. arabiensis density outdoors was similar to that indoors (11.8 vs 9.4/person/night, IRR: 1.3, 95 % CI: 0.8–1.9, P = 0.335). The mean hourly human-biting density of An. arabiensis was greater outdoors than indoors and peaked between 21:00 and 22:00 h. However, An. arabiensis parous population showed high indoor man biting activities during bedtimes (22:00 to 05:00 h) when the local people were indoor and potentially protected by IRS and LLINs. All mosquito samples tested for CSP antigen were found negative to malaria parasites. Results show much greater mosquito human-biting activities occurring outdoors than indoors and during early parts of the night, implying higher outdoor malaria transmission potential in the area. However, high bedtime (22:00 to 05:00 h) indoor biting activities of parous An. arabiensis suggest high potential intervention impact of IRS and LLINs on indoor malaria transmission.

This study was part of the work to prepare for a cluster-randomized controlled trial to evaluate the effect of combining indoor residual spraying and long-lasting insecticidal nets on malaria incidence. A pilot study was done to estimate the variations of malaria incidence among villages, combined with entomological collections and an assessment of susceptibility to insecticides in malaria vectors. A cohort of 5309 residents from four kebeles (the lowest government administrative unit) in 996 households was followed from August to December 2013 in south-central Ethiopia. Blood samples were collected by a finger prick for a microscopic examination of malaria infections. A multilevel mixed effect model was applied to measure the predictors of malaria episode. Adult mosquitoes were collected using light traps set indoors close to a sleeping person, pyrethrum spray sheet catches and artificial outdoor pit shelters. Enzyme-linked immunosorbent assays were used to detect the sources of mosquito blood meals, while mosquito longevity was estimated based on parity. The World Health Organization’s tube bioassay test was used to assess the insecticide susceptibility status of malaria vectors to pyrethroids and carbamates. The average incidence of malaria episode was 4.6 per 10,000 person weeks of observation. The age group from 5 to 14 years (IRR = 2.7; 95 % CI 1.1–6.6) and kebeles near a lake or river (IRR = 14.2, 95 % CI 3.1–64) were significantly associated with malaria episode. Only 271 (27.3 %) of the households owned insecticide-treated nets. Of 232 adult Anopheles mosquitoes collected, Anopheles arabiensis (71.1 %) was the predominant species. The average longevity of An. arabiensis was 14 days (range: 7–25 human blood index days). The overall human blood index (0.69) for An. arabiensis was higher than the bovine blood index (0.38). Statistically significant differences in Anopheline mosquitoes abundance were observed between the kebeles (P = 0.001). Anopheles arabiensis was susceptible to propoxur, but resistant to pyrethroids. However, An. pharoensis was susceptible to all pyrethroids and carbamates tested. This study showed a high variation in malaria incidence and Anopheles between kebeles. The observed susceptibility of the malaria vectors to propoxur warrants using this insecticide for indoor residual spraying, and the results from this study will be used as a baseline for the trial.

Long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS) are the main malaria prevention interventions in Ethiopia. There is conflicting evidence that the combined application of both interventions is better than either LLINs or IRS used alone. This trial aims to investigate whether the combination of LLINs (PermaNet 2.0, Vestergaard Frandsen, Lausanne, Switzerland) with IRS using propoxur will enhance the protective benefits and cost-effectiveness of the interventions against malaria and its effect on mosquito behavior, as compared to each intervention alone. This 2 x 2 factorial cluster randomized controlled trial is being carried out in the Adami Tullu district in south-central Ethiopia for about 116 weeks from September 2014 to December 2016. The trial is based on four arms: LLINs + IRS, LLINs alone, IRS alone and control. Villages (or clusters) will be the unit of randomization. The sample size includes 44 clusters per arm, with each cluster comprised of approximately 35 households (about 175 people). Prior to intervention, all households in the LLINs + IRS and LLINs alone arms will be provided with LLINs free of charge. Households in the LLINs + IRS and IRS alone arms will be sprayed with carbamate propoxur once a year just before the main malaria transmission season throughout the investigation. The primary outcome of this trial will be a malaria incidence based on the results of the rapid diagnostic tests in patients with a fever or history of fever attending health posts by passive case detection. Community-based surveys will be conducted each year to assess anemia among children 5–59 months old. In addition, community-based malaria prevalence surveys will be conducted each year on a representative sample of households during the main transmission season. The cost-effectiveness of the interventions and entomological studies will be simultaneously conducted. Analysis will be based on an intention-to-treat principle. This trial aims to provide evidence on the combined use of LLINs and IRS for malaria prevention by answering the following research questions: Can the combined use of LLINs and IRS significantly reduce the incidence of malaria compared with the use of either LLINs or IRS alone? And is the reduced incidence justifiable compared to the added costs? Will the combined use of LLINs and IRS reduce vector density, infection, longevity and the entomological inoculation rate? These data are crucial in order to maximize the impact of vector control interventions on the morbidity and mortality of malaria. PACTR201411000882128 (8 September 2014).

Increased understanding of the feeding behaviours of malaria vectors is important to determine the frequency of human-vector contact and to implement effective vector control interventions. Here we assess the relative feeding preferences of Anopheles mosquitoes in relation to cattle and human host abundance in southwest Ethiopia. We collected female Anopheles mosquitoes bi-weekly using Centers for Disease Control and prevention (CDC) light traps, pyrethrum spray catches (PSCs) and by aspirating from artificial pit shelters, and determined mosquito blood meal origins using a direct enzyme-linked immunosorbent assay (ELISA). Both Anopheles arabiensis Patton and An. marshalli (Theobald) showed preference of bovine blood meal over humans regardless of higher human population sizes. The relative feeding preference of An. arabiensis on bovine blood meal was 4.7 times higher than that of human blood. Anopheles marshalli was 6 times more likely to feed on bovine blood meal than humans. The HBI of An. arabiensis and An. marshalli significantly varied between the collection methods, whereas the bovine feeding patterns was not substantially influenced by collection methods. Even though the highest HBI of An. arabiensis and An. marshalli was from indoor CDC traps collections, a substantial number of An. arabiensis (65 %) and An. marshalli (63 %) had contact with cattle. Anopheles arabiensis (44 %) and An. marshalli (41 %) had clearly taken bovine blood meals outdoors, but they rested indoors. Anopheles mosquitoes are zoophagic and mainly feed on bovine blood meals than humans. Hence, it is important to consider treatment of cattle with appropriate insecticide to control the zoophagic malaria vectors in southwest Ethiopia. Systemic insecticides like ivermectin and its member eprinomectin could be investigated to control the pyrethroid insecticides resistant vectors.

Blood-feeding behavior studies are important for estimating the efficiency of pathogen transmission and assessing the relative human disease risk. However, in Ethiopia and other parts of East Africa there are large remaining gaps in identifying the feeding habits of Phlebotomus orientalis, the vector of Leishmania donovani. The aim of the study was to determine the blood feeding patterns of P. orientalis in Tahtay Adiyabo district, northern Ethiopia. For bloodmeal analysis, sandflies were collected from three different villages of Tahtay Adiyabo district using CDC light traps, sticky traps, and pyrethrum spray catches. Bloodmeal of engorged female sandflies was identified using cytochrome (cyt) b-PCR and reverse-line blotting (RLB) and enzyme linked immunosorbent assay (ELISA) assays. Most (637/641) of the females analyzed were P. orientalis. Successful identification of the host from bloodmeals was achieved in 83.03 and 92.1 % using cyt b PCR-RLB and ELISA, respectively. Bloodmeal analysis of P. orientalis females revealed that they have a range of hosts with predominant preference to bovines followed by donkey, human, goat, sheep, dog, and camel. Results obtained from bloodmeal analyses demonstrate that the feeding preference of P. orientalis is mainly zoophilic, which could vary depending on the availability of hosts.