In this study, phenotypic analysis of G6PD enzyme activity indicated that 3.6% (18/498) of the study participants had <30% enzyme activity of the AMM which is stated as G6PD deficiency and 56 female patients were with intermediate G6PD enzyme activity (30-80%) of AMM assumed to have heterozygous gene mutation.
The G6PDd was relatively higher in females (4.8%) than males (2.8%). This result contradicts the assumption that females are less affected with G6PDd due to genetic preferences of the X-chromosomes (2). The prevalence of G6PDd found in this study was slightly lower than finding from a previous study conducted in seven sites of Ethiopia, which reported 5.4% (10/184) among males and 5.2% (7/136) among females (21). This study also revealed lower G6PDd compared to the study report from other African countries, with a G6PDd of 13% (36/278) (32). The current finding is lower than 7.3% (33/449) reported from Gambella Hospital, Southwest Ethiopia, with 8.6% in males and 6.3% in females (33). A nationally conducted study in Ethiopia discovered 8.9% of G6PDd (19). The G6PDd in this study was higher than Jimma (34) and other community-based study in other parts of Ethiopia that showed relatively low levels of G6PD deficiency (22).
G6PD enzyme activity varies among different nations, regions and even between local ethnic groups. The prevalence of G6PDd was slightly higher among Nuers (14.3%) and Anuak (12.0%) compared to the highlanders with no deficiency in Gambella region (33). Malaria infection might impose G6PD enzyme genetic mutation due to natural selection mechanisms which results in relative incremental of G6PDd in malaria endemic areas compared to non-endemic areas (22). Metehara is one of the seasonal malaria endemic areas in Ethiopia that we observed higher G6PDd prevalence than the community based studies in different settings and lower than similar studies in stable endemic areas like Gambella (33).
Malaria status showed marginal but not statistically significant association with G6PDd, while sex and history of malaria infection were significantly associated with G6PDd status. The prevalence of G6PDd among females was three times higher than in males and it was four times higher among previously malaria infected patients than those never infected individual. In a study report from Gambella Hospital, Ethiopia the prevalence of G6PDd was higher among males than females (33). Likewise, another study conducted in seven study sites of Ethiopia found no significant association of G6PDd with sex and malaria infection (21). Conversely, the incidence of G6PDd among malaria smear positive patients was significantly higher than the malaria negative patients (33).
This variation in associations of various factors with G6PDd prevalence might be due to the difference in ethnic variation of the study participants, endemicity of malaria infection, the sample size and sampling techniques of the studies, the laboratory methods used for the analysis of samples in the study and the time of the study.
Based on molecular sequencing, mutations were detected in G267+119C/T, A376T and ChrX: 154535443 with G267+119C/T being most prevalent. Although previous studies in Ethiopia reported the presence of A376G, G202A, C563T, G1116A and 485+37, these mutations were not identified in this study. Contrary to this study, 13% of the study participants showed G6PDA- (G202A) genotype in Brazaville, Republic of Congo (12), 12.5% (39/311) depicted G6PDA+ (A376G) in Eritrea (35). Several studies in different localities of Ethiopia showed that G6PDA+ (A376G) was the only mutation observed in 8.9% of sampled population (19), and 23.26% (20/86) G6PDA+ mutations were detected in Southwestern Ethiopia (20). The G202A mutation was also detected in 3.5% individuals (22) and in another study, G6PDA+ mutation was detected in 6.1% (21/344) of individuals, G267+119C/T and G1116A mutations found in 1.2% (4) and 1.2% (4) individuals, respectively (21). Whereas no mutation was detected in previous study conducted in Shewa Robit (21) and out of 34 low enzyme activity samples genotyped only one G6PDA+ and one G445A mutations were identified in Oromia region (34).
The A376T mutation represents the exchange of adenine by thyamine, which generates the amino acid replacement of 126 Asn with Tyr. While this mutation is new to Ethiopia, it was previously found in Mexico and termed as San Luis Potosi (36). There were six samples with absence of mutations despite phenotypically showing low enzyme activities. In other study one sample with low G6PD enzyme activity had no G6PD mutation (21). On the other hand, the mutations observed in A376G, G267+119C/T, and G1116A were not associated with low G6PD activity (21). The lack of association of G6PD enzyme activities with respective genotypes advocates for the need of further verification with large sample size. A codon that has not been sequenced in this study may have contributed to the phenotypic expression of low enzyme activity.
Generally, previous studies that had been conducted in different parts of Ethiopia on the distribution of G6PDd showed that few variants like G6PDA+, at North, south, West, and East of the country were identified (19, 21). G267+119C/T and G1116A were detected in the southern parts of the county (20, 21) and one mutation at position 445G◊A was identified in Jimma (34). This study also identified the pre-existing genotypes as well as the new genotype, A376T mutation. The occurrence of new genetic variants in such a small-scale study suggests the need of a large scale G6PDd epidemiologic study across the country to characterize the full array of G6PD genetic variants in Ethiopia.