First detection of Anopheles stephensi Liston, 1901 (Diptera: culicidae) in Ethiopia using molecular and morphological approaches

Abstract

Malaria is a major public health concern in Ethiopia. With the increase in malaria cases in the Somali Region of Ethiopia, understanding the distribution and identifying the species of malaria vectors is vital to public health. Here we report the first detection of Anopheles stephensi in Ethiopia, a malaria vector typically found in the Middle East, the Indian subcontinent, and China, but recently found in Djibouti. An entomological investigation was conducted during November to December 2016 in Kebri Dehar town of the Ethiopian Somali Regional State as ancillary work for Anopheles spp. surveillance. Mosquito larvae were collected from water reservoirs. Larvae were reared in the laboratory to the adult stage and identified morphologically. PCR and sequencing of cytochrome oxidase 1 (COI) and internal transcribed spacer 2 (ITS2) loci were performed. Basic Local Alignment Search Tool (BLAST) was used to compare sample sequences to sequences in the NCBI nucleotide database for species identification. To further analyze the relationship between the specimen we collected in Kebri Dehar and other Anopheles samples available in Genbank, phylogenetic analysis was performed using a maximum likelihood approach. Molecular and morphological results confirm specimens were An. stephensi. The closest high scoring hit was for all specimens was for the An. stephensi sequence. Independent phylogenetic analyses of COI and ITS2 sequences revealed in both cases strong bootstrap (100) support for our sequence forming a clade with other An. stephensi sequences to the exclusion of any other species of Anopheles. In conclusion, Anopheles stephensi is present in Kebri Dehar town in Ethiopia. These findings highlight the need for additional research to examine the role of An. stephensi in malaria transmission in Ethiopia.

Geographical distribution of Anopheles stephensi in eastern Ethiopia

“…These data, taken with previous reports of An. stephensi in Kebridehar in 2016(Carter, Yared et al 2018), confirm that An. stephensi is established in this region.…”

“…An. stephensi was detected in Ethiopia for the first time in 2016 in Kebridehar (Somali Region) but it remains unclear how broadly distributed the species is in the rest of the country (Carter et al 2018).…”

“…Understanding the distribution of An. stephensi in Ethiopia is critical to evaluating the threat it poses to malaria control in Ethiopia and the rest of the Horn of Africa (Carter et al 2018). It is important during initial surveillance of a potential new vector to evaluate the accuracy of species identifications.…”

Anopheles stephensias an emerging malaria vector in the Horn of Africa with high susceptibility to EthiopianPlasmodium vivaxandPlasmodium falciparumisolates

“…Maximum-likelihood topologies were constructed using representative reference sequences with published geographical data downloaded from GenBank. Within the Ethiopian population, due to the presence of a hyper-variable microsatellite region, ITS2 sequences (A) were trimmed to create a consensus alignment of 289bp; one polymorphic site separated samples into two genotypes (indicated with filled asterisk together with the previously reported genotype, MH650999, Carter, et al [14] in unfilled asterisk). COI sequences (B) were assembled into a consensus alignment of 687bp; a total of four variable sites were identified, corresponding to four haplotypes (indicated with filled asterisk together with the previously reported genotype, MH651000, Carter, et al [14], unfilled asterisk).…”

“…Recent reports indicate that An. stephensi is spreading in the Horn of Africa (Djibouti [13], Ethiopia [14] and the Republic of Sudan [15]). An.…”