Hepatitis B virus (HBV), a highly contagious infectious disease being as a major threat in developing countries. Hence, this study focuses on assessing the prevalence of HBV genotype in Eritrea. This molecular genotyping of HBV was first of its kind in Eritrea using a PCR-based method, and no data about the genotypes and mutants of HBV in patients with liver disease were previously reported.
Different HBV genotyping methods have been developed including sequencing, INNO-LiPA, restriction fragment polymorphism, multiplex PCR, serotyping, oligonucleotide microarray chips, reverse dot blot, restriction fragment mass polymorphism, invader assay, and real-time PCR. However, the sensitivity, specificity, expense and time requirements differ among these methods [17]. In this study, we focused on six major genotypes (A–F) among our patients using the multiplex nested PCR technique developed by Naito et al. [16]. This method appears to have higher sensitivity for detecting mixed genotypes, and it is simple and cost-effective for large population studies with a high accuracy rate of 93% [17].
The genotypes of HBV were examined in 293 patients, and samples were successfully genotyped in 122 (41.6%) patients. This genotype success rate was comparable with the findings obtained using different genotype methods in various countries, such as United Arab Emirates, Nigeria, Egypt, India, Cote d’Ivoire and Iraq, in which the genotype success rates were 95%, 83.6%, 71.4%, 69.7%, 68.7%, and 60.5%, respectively [18–22]. These findings confirmed the variability of different methods of HBV-DNA detection in relation to HBsAg positivity, and that the variability could be influenced by whether patients have chronic infection [23].
The most important finding in our results was that single-genotype HBV infection (57.38%) was more common than mixed-genotype infection (42.62%). This is in concordance with findings from Taiwan, in which genotype B infection was most common [24]. Our finding of single-genotype predominance also agrees with a study in Egypt using INNO-LiPA, which reported that 87% of patients harboured single-genotype infection, most commonly genotype D [25]. However, these findings contrast with those from Iraq and Nigeria, in which mixed-genotype infections were documented in 75 and 82.6% of patients, respectively [19, 26].
The identification of five HBV genotypes (A–E) in this study corroborates their higher prevalence in certain geographical regions in Africa [2, 4, 27]. Irrespective of the occurrence of mixed or single infection, our study identified that genotype D had the highest prevalence, followed by genotypes C, E, C/D, B, A/E, B/E, and A/D/C. However, in terms of single-genotype infection, genotype D was the most prevalent, followed by genotypes C, E, A, and B. On the other hand, concerning mixed-genotype HBV infection, genotype C/D infection predominated, followed by genotypes C/E, C/D/E, A/D, D/E, A/E, B/E, and A/D/C.
We found that the genotypes of HBV in Eritrea conform to those described in the region. Our results of this study concur with previous findings in neighbouring countries, including the predominance of HBV genotype D in Sudan [12, 13] and Egypt [28]. Globally, mixed-genotype HBV infection has been reported to predominant in different regions [10]. Eritrea’s geographical location in the horn of Africa with close proximity to Asia through the Red Sea could explain the nature of distribution of HBV genotypes in the country. The observation of genotype C infection alone or together with genotype D, especially in people hailing from coastal areas of Eritrea, may be attributable to frequent migration and contact with people from regions such as the Middle East because of commercial activities. In addition, mixed-genotype infection was noted in patients from different geographical areas of Eritrea, in line with observations of infection by genotypes A, D and E, including single and mixed infections, in Sudan [10]. In addition, the observation of mixed-genotype infection in this country could be linked to the significant number of refugees returning from neighbouring and distant countries [29].
The clinical impact of HBV genotype D has not been studied extensively. Emerging evidence suggests that patients with genotype D infection may develop fulminant hepatitis at high frequency [30]. The prevalence of different HBV genotypes in our study subjects provides a basis to compare different parameters in a stratified manner for various genotypes (Table 5). Patients infected with HBV genotype C exhibited a higher viral load (3.63 log IU/ml) than that reported in one study [31] but this was not true when compared with a second report [14]. When assessing HBeAg seropositivity among isolated genotypes, a significantly higher frequency of HBeAg positivity was noted among reported genotypes. In a similar study, Yousif et al. found higher rates of HBeAg positivity in patients with liver disease associated with HBV genotype D [13]. Conversely, Mahgoub et al. observed higher HBeAg positivity rates in blood donors infected with HBV genotype E [32].
The distribution of HBV genotypes within the country was assessed using the location where the patients hailed from. Concerning single-genotype infection, genotype D was predominant in Zoba Maekel (23.6%) and Gash Barka (27.3%). Among mixed HBV genotypes, genotype C/D infection was predominant in Gash Barka (18.2%) and Debub (16%), whereas HBV genotype C/E infection was predominant in Maekel (14.5%) and DKB (25%). Similarly, the genotype distribution among various subgroups in various countries, including Eritrea, was reported by Scotto et al. [33], and Asaad et al. described the genotypic distribution of HBV in the Saudi population [34].
Knowing the predominant HBV genotype in specific areas is important for assessing diagnostic capabilities and vaccine efficacy [1]. Our study narrows the existing gaps in HBV molecular research in Eritrea. All of our isolates were obtained from patients with chronic HBV infection in hospital settings. Generally, patients with liver disease who are identified as reactive for HBV infection during the study were not monitored or evaluated concerning the exact stage of their liver disease such as cirrhosis or hepatocellular carcinoma (HCC). Hence, we suggest that HBV genotyping of patients be studied in correlation with the clinical progression of liver disease to provide a clear clinical picture and molecular epidemiology. Furthermore, future studies must evaluate the clinical relevance, treatment response, and rates of co-infection, which may affect disease outcome.