Seroprevalence, trends, and risk factors of hepatitis B and C among family replacement blood donors; a 7-year retrospective study at Sunyani Municipal Hospital, Ghana

ABSTRACT Hepatitis B and C cause chronic infections which develop into liver-related sequelae, like cirrhosis and liver carcinoma. This study determined the seroprevalence, trends, and risk factors of HBV and HCV among family replacement donors. A retrospective review of primary data on blood donors screened between January 2015 and December 2021 was conducted at Sunyani Municipal Hospital. The data were assessed for seroprevalence, trends, and odds ratios using SPSS. Of 6847 donors, the majority were males (88.1% [6033]), ≤24 years (27.4% [1874]), O blood type (69.8% [4776]), and Rh-positive (89.9% [6154]). The seroprevalences of HBV and HCV were 3.2% and 1.9%, respectively, with more males infected with HBV and HCV (3.4% vs 2.0%). Males were 2.842 times (p = .001) and 2.399 times (p = .025) more susceptible than females to HBV and HCV, respectively. In the rainy season, donors were 1.489 times (p = .041) more susceptible to HCV. HBV and HCV seroprevalence declined over the period (slope: −0.5464, p ≤ .001 vs slope: −0.6179, p ≤ .001). Male gender and rainy season were significant determinants of both infections. The seroprevalence of HBV was higher than HCV despite the significant decline in both infections. We, recommend health authorities intensify health education among males and during the rainy season.


Introduction
Worldwide, whole blood and its fractionated products are transfused to save the lives of patients suffering from anemia due to either sudden or insidious blood shortages. [1] Patients with a high risk of transfusion include hemorrhaging pregnant women, patients with anemia due to Plasmodial infections, [2,3] sickle cell disease and other hemoglobin disorders. [3] Although blood transfusion plays a significant therapeutic role in patient care, healthcare providers still rely on the benevolence of human donors for blood, since there is currently no alternative to blood. [4] Consequently, there is the propensity to disseminate diseases through the donation and transfusion of infected blood units.
Blood transfusion is a notable source of spread for transfusion-associated infectious agents like hepatitis B virus (HBV) and hepatitis C virus (HCV), which have similar routes of spread including sexual intercourse and or contact with infected blood. [5] Although effective prophylaxis against HBV exist for several decades, about 2 billion infected individuals have been recorded globally, of which about 248 million are persistently infected. [6] Hepatitis B virus causes chronic infections in over 81 million individuals in Africa. [7] Such persistent infections result in hepatitis-associated sequelae like cirrhosis and carcinoma of the liver. [7] For instance, approximately 60% of liver carcinomas detected in underdeveloped countries are due to either HBV or HCV infection. [8] Conversely, the burden of HCV infection is estimated at over 71 million cases globally, [6] of which 8% occur in Africa. [9] Consequently, infections resulting from both HBV and HCV account for approximately 1.1 million deaths annually due to persistent liver morbidities and carcinoma. [10] In Ghana, blood donors undergo screening to exclude infected donors, however, the screening may be insufficiently done due to limited resources [9] and this may pose a significant health risk to blood recipients. The burden of HBV and HCV in Ghana is fragmented and varies from one place to the other, with some [11,12] showing local variations. Although several studies [11,13,14] conducted in Ghana reported on the prevalence, and trends of HBV and HCV, they failed to determine either the risk factors of these infections or their association with season. Furthermore, a similar study conducted by Walana et al. [14] in the Bono Region focused only on the prevalence of HBV while no attention was accorded to HCV despite its association with severe liver-related diseases.
This study, therefore, determined the seroprevalence, trends, and risk factors of HBV and HCV among apparently healthy family replacement blood donors. This aims to document the epidemiology of HBV and HCV in the Sunyani Municipality since there is currently a paucity of data on the burden of hepatitis infections in this setting.

Study design and setting
This was a single-center hospital-based retrospective study conducted between January and May 2022, at the Sunyani Municipal Hospital, Ghana. The Sunyani Municipal Hospital is a 105-bed capacity healthcare facility located within an urban and central business district of the Sunyani Municipality. The hospital is one of several-and the second largest public hospital in Sunyani, after the Regional Hospital. The hospital manages both in-patient and outpatients, with services ranging from public health, pharmacy, trauma and surgery, internal medicine, clinical diagnostic laboratory, chest clinic, earnose-and-throat (ENT) clinic, antiretroviral therapy (ART) clinic, dental clinic, eye clinic, medical imaging (including ultrasonography and x-ray), antenatal and post-natal clinic, and a morgue. The hospital has separate inpatient wards for obstetric, post-surgery recovery, pediatrics, and adult female and adult male patients. Sunyani is the administrative capital of the Sunyani Municipality and is located within the western section of the Bono Region. The municipality is inhabited by approximately 123,224 people. [15]

Study population
In this study, we retrospectively reviewed primary laboratory data of prospective family replacement blood donors who visited the blood bank of Sunyani Municipal Hospital in the Bono Region between January 2015 and December 2021. Information on 6847 family replacement blood donors screened within this seven years (2015-2021) and had their data entered in the blood bank's archived records were retrieved using convenient sampling.

Sample size
The sample size was determined using Epi Info, version 7.2.2.2 (Centers for Disease Control and Prevention [CDC], U.S.A). An HBV seroprevalence of 9.6% reported by Lokpo et al., [13] was used as the percentage outcome of unexposed group, 12.0% as percentage outcome in exposed group, 80% power, 95% two-sided confidence level, and ratio (unexposed: exposed) of 1. This yielded a maximum sample size of 5414, which was conveniently increased to 6847 donors to give a good representation of the population.

Data collection
Primary laboratory data on prospective family replacement blood donors which were completely entered in the blood bank registers and legible for data extraction were included in the study. Conversely, records with incomplete and/or illegible donor information were excluded. The eligibility of each donor record was determined by the completeness and correctness of the information. Some of the donor information retrieved included year and month of screening, age of donors in years, gender, ABO and Rh blood types, and results of serological screening for HBV and HCV.

Screening of blood donors
The prospective blood donors were healthy individuals between 17 and 60 years of age. All the donors were assisted to complete a self-exclusion predonation questionnaire to help exclude unsuitable donors with high-risk behavior and morbidities. Anthropometric data including weight and blood pressure were measured. Furthermore, serological screening of infectious agents which included HCV and HBV was performed on serum from each blood donor using rapid lateral-flow immunoassays. The ABON test kits with catalog numbers IHC-401 and IHBsg-401 for HCV and hepatitis B surface antigen (HBsAg) respectively were used (Abbott Laboratories, U.S.A). The assays qualitatively detect the presence of HCV antibodies and HBsAg in blood. In the HCV assay, HCV antibodies present in either whole blood, plasma or serum reacts with antigen-colloidal gold conjugate embedded in the specimen pad of the test kit to produce conjugate-HCV antibody complex. The complex is then captured by immobilized antibody-binding protein A to produce a pink colored band on the test strip. Conversely, in the HBV assay, HBsAg present in the specimen reacts with monoclonal antibody-colloidal gold conjugate embedded in the nitrocellulose membrane. The complex migrates to the test region where it is captured by immobilized anti-HBs antibodies and generates a colored band. For both HBV and HCV assays, the presence of colored control and test bands indicate a positive test. A confirmatory and viral load test of all positive donor specimens were performed using polymerase chain reaction. Donors with positive results were disqualified from donating and immediately referred to see a physician for counseling and management.

Statistical data analysis
The spreadsheet containing the data was checked for entry errors and imported into IBM SPSS Statistics for Windows, Version 23 (Armonk, NY: IBM Corp.) for binning, recoding, and analyses. Microsoft Excel 2016 for Windows (Microsoft Corporation, USA) was used to visualize the data. The only scale variable, age, was presented as median and interquartile range. Also, it was transformed into four categories, using "visual binning." The months in which donors were screened were recoded into rainy-and dry seasons, the two major seasons observed in Ghana. This was done as described by Logal et al. [16] : the rainy season included April-July and September-November, whereas the dry season included December-March and August. All the categorical data were presented as frequencies and proportions, while Chi-square and Fisher's exact tests were used to determine associations between different categories. The determination of age as a risk factor for the viral markers was done using multinomial logistic regression, whereas for gender and donor type binary logistic regression was used. P-value ≤0.05 was considered significant for all statistical analyses.

Ethical consideration
Ethical approval was sought from the Committee on Human Research, Publication and Ethics (CHRPE) at the School of Medicine and Dentistry, Kwame Nkrumah University of Science and Technology (Reference: CHRPE/ AP/342/22). Also, permission was sought from the Administration of the Sunyani Municipal Hospital before conducting the study. However, due to the retrospective design of the study, consent from the donors was not required.   Table 2).   (Table 3). Table 4 shows the sociodemographic risk factors of HBV and HCV among the blood donors. Male blood donors were 2.842 times (CI: 1.500-5.385, p = .001) and 2.399 times (CI: 1.116-5.157, p = .025) more susceptible than females to HBV and HCV, respectively. The risk of the blood donors being infected with either HBV or HCV did not significantly differ across different age groups.  Blood donors screened during the rainy season were 1.489 times (CI: 1.017-2.180, p = .041) more susceptible to HCV. The risks of the donors being infected with HBV, and HCV were significantly different between gender, whereas for season the risks of being infected with HCV was significant. The age of blood donors, however, was not a significant risk factor for HBV and HCV in this study (Table 4).

Discussion
Presently, human blood donors remain the only source of blood for transfusion. There is, therefore, the propensity for infectious agents like viral hepatitis B and C from infected blood donors to threaten the blood supply chain. This retrospective study reviewed records of 6847 family replacement donors at the Sunyani Municipal Hospital, to determine the seroprevalence, risk factors, and trends of HBV and HCV among family replacement donors. The blood donors in this study were predominantly males (88.1%), with a male to female ratio of 7:1. This was consistent with findings of other studies in Ghana [9] and Mali. [17] Nkansah et al. [9] inferred that the high turnout of male donors is attributed to the fact that a lot of males meet the standards for blood donor selection, and further used this to justify the increased population of younger donors. However, the basis for the reduced level of female participation in blood donation is suggested to be the result of peculiarities associated with the female gender; according to some studies, obstetric factors like menstruation, [18][19][20] childbearing, [18][19][20][21] and lactation [18,20,21] lead to increased deferral of females, making males more suitable candidates. [19] Conversely, a study by Degefa et al. [22] in Ethiopia showed more female than male donors, and although this contrasting finding was not justified, the probable cause may be due to the recruitment of more voluntary donors in their study. The seroprevalences of HBV and HCV among the family replacement donors in this study (3.2% vs 1.9%) were lower compared to the findings of other studies conducted in other parts of Ghana. [11,13,14] Also, the observation of an increased HBV seroprevalence than HCV in the current study corroborates similar patterns (9.6% vs 4.4%) reported by Walana et al. [12] in the Bono Region, Ghana; (6.94% vs 1.84%) in the Volta Region [13] ; and (4.7% vs 0.7%) in Ethiopia. [23] The comparison of the lower seroprevalences of the infectious markers observed in this study to the findings reported in other parts of Ghana [11,14] suggests the existence of local variations in the burden of hepatitis. This corroborates data from other West African states, including Burkina Faso. [24,25] The probable cause of such variations in the burden of infectious agents has been attributed to differences in the geographical distribution of the disease, immigration of the population, [26,27] the accuracy of the assays employed, and the donor recruitment methods used. [26] Furthermore, Attaullah et al. [28] posit that although blood donors are used as a proxy to extrapolate the burden of infectious agents to the general population, they may not give a true reflection of the burden in the general population since the seroprevalence may be miscalculated owing to varied population traits. This may have accounted for the comparably low seroprevalences reported in our study. Conversely, in the study by Nkansah et al., [9] the burden of HCV was rather increased compared to HBV (11.7% vs 10.3%). This is because the frequency of HCV is much affected by the geographical settings of inhabitants [5] ; whereas Nkansah and colleagues conducted their study in a rural district, ours was in the city. Therefore, the exposures, population risks, and other determinants that influence the transmission of these infectious agents may be different.
The seroprevalences of HBV and HCV were significantly associated with gender, with more male than female donors infected with both infections. This corroborates the findings of another study conducted in the middle belt of Ghana. [9] Furthermore, male blood donors were nearly 3 times (OR: 2.842, p = .001) more susceptible to HBV, and 2 times (OR: 2.399, p = .025) more susceptible to HCV. This is because females are less vulnerable to viruses, and can produce adequate, acute, and long-lasting first-line and adaptive immunity. [29] This is further linked to the increased synthesis and differentiation of innate cells like macrophages, dendritic-and monocytic cells which enable females to produce acute inflammatory reactions [29] in response to the viral infection. Furthermore, the increased estrogen levels in females enable them to produce increased CD4+ immune cells that stimulate a pronounced activation of immune T cells. [29] The findings of this study suggest an association between HCV seroprevalence and season, with increased seroprevalence in the rainy season (p = .044). The donors were more susceptible to HCV (OR: 1.489, p = .041) in the rainy season than in the dry season, whereas, HBV was rather increased in the dry season, although not significant. However, the reason for this occurrence is not understood, and since both infections have a similar mode of spread, [30] it is unclear why there exist varied patterns for both infections in different seasons. However, Hernández-Alvarez et al. [31] suggest a nexus between the production of vitamin D and exposure to sunshine, which is affected by a change in season. Therefore, it is plausible that the increased susceptibility of the donors to HCV could be due to the relatively reduced access to sunlight during the rainy season, a situation that could result in inadequate vitamin D synthesis, which may have further led to compromised immunity [31] in the donors.
The burden of both HBV and HCV was increased in blood type B donors, while HBV was increased among the Rh-negative blood donors and HCV was rather increased among Rh-positive donors. Although the significance of the ABO blood group system has been reported widely, and its association with several disease states have been highlighted in several studies, [32][33][34][35] there were no significant associations between hepatitis and the blood group systems in this study. This corroborates the conclusions of a similar study conducted in India. [35] Although the yearly burden of HBV and HCV was inconsistently decreasing, general declining trends were observed for both infections over the 7 years using the seroprevalences obtained in 2015 as the baseline. This corroborates the findings of similar studies conducted in China [36] and the Ashanti Region, Ghana. [9] This observation suggests a paradigm shift toward either improved lifestyle choices, responsible sexual behavior, and increased health education, or in the case of HBV it may be the cause of increased awareness and vaccinations over the years.
This study had some limitations: firstly, due to the retrospective nature of the study, we were unable to ascertain donors' HBV vaccination status and its effect on the overall declining trend of the infections. Secondly, the donor records did not indicate whether the donors were first-time or repeat donors; Ji et al. [36] suggest that there exists an increased prevalence of infectious markers among first-time than repeat donors. Therefore, this information would have better explained the cause of the declining trends of the infections. Thirdly, the retrospective nature of the study permitted the evaluation of only a few sociodemographic risk factors for HBV and HCV. Furthermore, the use of only rapid immunoassays did not permit the determination of occult viral infections, which may have led to underestimation of the infections. Additionally, the study design did not allow for the estimation of serum vitamin D levels in the donors, which could have explained the variability in seasonal seropositivity to HCV.
Our findings suggest that the seroprevalences of HBV and HCV were comparably low. Gender was significantly associated with both HBV and HCV, while season was significantly associated with only HCV. Whereas the male gender was a significant determinant of both infections, the rainy season was a determinant of HCV. Furthermore, the seroprevalence of HBV was higher than HCV, despite the declining trends of both infections recorded over the years. We recommend that health authorities intensify health education, especially during the rainy season and among males. Local variations in the seroprevalence of these infections in Ghana calls for upgrade and standardization in the screening tests, including using molecular techniques to detect occult infections among blood donors across Ghanaian blood centers.

Disclosure statement
No potential conflict of interest was reported by the author(s).

Funding
The work did not receive any specific grant from funding agencies in public, commercial, or non-profit-sectors.