This study was conducted to assess the current P. falciparum genotypic structure in the semi-arid area in North East Ethiopia, using the highly polymorphic (block 3) region of the msp2 gene as a molecular marker. The msp2 marker is recommended for genotyping P. falciparum parasite populations compared with msp-1 and glurp [30]. Plasmodium falciparum isolates from this region were mainly monoclonal with a low MOI and limited genetic diversity. These findings are important for ongoing evaluation of the effect of malaria control strategies, as Ethiopia moves towards malaria elimination.
The 3D7/IC allelic family of msp2 was more prevalent than the FC27 allelic family. This is in agreement with previous reports from Burkina Faso [31], South West Ethiopia [22] and Sudan [32]. However, this finding differs to results from North West Ethiopia [20], and Central Sudan [33], where FC27 was the more prevalent allelic family. These differences could relate to the semi-arid geographic setting and low transmission intensity compared to the hot and humid climate in North West Ethiopia.
Limited genetic diversity of P. falciparum was observed in this study. Similar results have been reported in other areas with low P. falciparum transmission [34] and in regions with declining transmission related to malaria control efforts [35]. In contrast, a high level of genetic diversity was reported in high endemicity settings in Cameroon [15] and Burkina Faso [31].
The current study found that the P. falciparum parasite population in Melka-Werer exhibited a low heterozygosity (He=0.5), consistent with that reported in Mubuga, Rwanda (He=0.49) [36]. In areas with declining local transmission, it is expected that lower parasite diversity (heterozygosity) will be present [37]. Declining diversity and transmission have been associated with improved malaria control interventions [38,39].
The overall mean MOI reported in this study was low (MOI=1.2). This is in agreement to previous studies where low malaria transmission settings are commonly associated with lower MOIs [40,41], and is consistent with reports from semi-desert settings in neighbouring Sudan and Djibouti [32,38]. The low MOI contrasted with a finding from a higher endemic setting in Humera, Ethiopia [19]. The low MOI observed in this study may reflect most positive samples being from adult patients, with previous reports finding a reduction in MOI in adults compared with children [43].
The majority of participants in the current study were older than 10 years, similar to results from an area with a lower intensity of malaria transmission [31] but contrasting to reports from high transmission settings [44]. It is also possible that the age-related malaria risk may have been influenced by implementation of effective malaria control interventions, such as the widespread distribution of long-lasting insecticidal nets (LLINs) and indoor residual spray (IRS), and sustained treatment of malaria patients with artemisinin-based combination therapy (ACT). This is supported by the 2015 malaria indicator survey, which found that the Afar region had the highest percentage of use of LLINs compared to other regions of the country [6].
Age is considered an important factor in the acquisition of immunity against P. falciparum and may have also an effect on MOI [45], although, the influence of age on the MOI is highly affected by malaria transmission intensity [46]. Previous studies have shown an association between age and MOI in areas with intense perennial malaria transmission or hypo-meso-endemic malaria transmission [47,48]. However, the current study found no association between age and MOI. Similar findings have been reported in other countries [49,50].
A higher geometric mean parasite density in individuals with previous exposure to malaria attack was found. In this low endemicity setting, a lower proportion of individuals will have likely had prior immunity, meaning that infected patients will be more likely to become symptomatic at a lower parasitaemia than in high endemicity settings.
It was difficult to correlate transmission levels with genetic diversity and MOI in the current study due to a lack of entomologic inoculation rate (EIR) data from the study area. However, the genetic diversity and the MOI reported in the present study supported a low average microscopy positivity rate (8.5%) (Melka-Werer rural town health office data, 2015, unpublished). A limitation of this study was the small sample size, in part due to the nomadic nature of the local communities. Furthermore, due to resource restrictions, lower discriminatory power agarose gel electrophoresis compared to capillary electrophoresis was used [51]. Further, the limited allelic frequency and genetic diversity observed may have been due to the detection limit of the PCR technique used in the study. Allelic fragment length intervals of less than 20 base pairs may not be clearly distinguished on agarose gel and may lead to misclassification of the genotype. Allele differentiation could be improved by using more discriminatory techniques in future studies, such as DNA sequencing or SNPs.