Hepatitis E virus (HEV) is a single-stranded positive-sense non-enveloped RNA virus with a full-length genome of approximately 7.2 kb, which can cause viral hepatitis E (abbreviated as hepatitis E) [1] . Hepatitis E usually presents as asymptomatic or self-limiting recovery, and the symptoms generally subside within 2 to 6 weeks. However, pregnant women, organ transplant patients, the elderly, and immunocompromised individuals have a higher probability of developing severe hepatitis E and a higher mortality rate after infection [1] . The transmission routes of HEV include the digestive tract (fecal-oral), blood transmission, and mother-to-child transmission. In recent years, with an increasing number of cases reporting the transmission of hepatitis E virus through blood transfusion, the issue of HEV transmission through blood transfusion has become a serious public health challenge, attracting widespread attention in the field of blood transfusion [2] - [6] . China is a high-prevalence region of HEV, and currently, HEV has not been included in the blood screening program for blood donors. Therefore, it is urgent to conduct research on the prevalence of HEV in various regions to provide a reference for formulating appropriate recruitment and screening strategies. The Qinzhou region faces challenges in addressing the safety of blood transfusion regarding HEV and lacks detailed data support. In this study, the antigen and antibodies of hepatitis E virus in unpaid blood donors in the Qinzhou region were detected by the enzyme-linked immunosorbent assay (ELISA) method, and nucleic acid testing for HEV RNA was performed on samples positive for HEV-Ag and/or HEV-IgM. The aim is to understand the infection status and distribution of hepatitis E in the Qinzhou region, provide data support for the national blood screening work, and thus, it is of great significance for ensuring blood safety.

In recent years, the number of cases of hepatitis E (HEV) transmitted through blood transfusion has gradually increased globally, which has attracted widespread attention. According to the report of the World Health Organization (WHO), there are approximately 20 million cases of hepatitis E virus infection worldwide every year, resulting in about 3.3 million symptomatic cases of hepatitis E. WHO estimates that in 2015, hepatitis E caused about 44,000 deaths, accounting for 3.3% of the deaths from viral hepatitis [7] . According to the data of the National Disease Prevention and Control Bureau, from 2015 to 2021, a total of 185,946 cases of hepatitis E were reported in the whole country (excluding the Hong Kong and Macau Special Administrative Regions and Taiwan Region), with 103 deaths. The average annual incidence rate was 2 per 100,000, ranking third in both incidence and mortality among viral hepatitis [8] . In the “Technical Operation Procedures of Blood Stations (2019 Edition)” in China, HEV has not been included in the blood screening strategy, and there is a lack of sufficient data support on whether to conduct routine or selective screening of blood donor samples. Many blood collection and supply institutions in China have carried out epidemiological investigations on the HEV infection status of unpaid blood donors, and the results show that there are significant regional differences in HEV infection. According to a Meta-analysis in 2023 on the prevalence of HEV infection markers in unpaid blood donors in China [9] , the prevalence rates of HEV-IgG, HEV-IgM, and HEV RNA in unpaid blood donors in China were 23.0%, 1.13%, and 0.028%, respectively, with significant differences among different cities and regions. This study shows that the positive rates of HEV-IgG, HEV-IgM, and HEV-Ag in unpaid blood donors in the Qinzhou region were 11.59%, 0.73%, and 0.054%, respectively. The positive rate of HEV in the Qinzhou region is only higher than that in Tongzhou, Beijing (1.17%, 0.5%) [10] , but generally lower than that in other domestic regions, such as Wuhan (16.44%, 1.0%, 0.08%) [11] , Guangzhou (20.05%, 0.76%, 0.04%) [12] , Liuzhou, Guangxi (26.03%, 1.67%, 0%) [13] , Jiangsu (19.23%, 1.49%) [14] , Xiamen (23.10%, 0.67%, 0.19%) [15] , Chongqing (49.19%, 1.67%, 0.20%) [16] , Lishui, Zhejiang (24.81%, 1.67%) [17] , Hong Kong (15.17%, 0.8%, 0.02%) [18] , and Qingdao (12.94%, 0.78%, 0) [19] , etc. The research results show that although there is a potential risk of HEV transmission through blood transfusion in the Qinzhou region, the prevalence rate of HEV is at a relatively low level in China, which may be closely related to local factors such as the economy, health conditions, medical environment, and vaccination policies. It is worth noting that Qinzhou City was rated as a “National Hygienic City” in 2021. In recent years, the municipal Party committee and government have attached great importance to consolidating and enhancing the achievements of creating a national hygienic city, continuously strengthening the construction of public health environment infrastructure, accelerating the improvement of urban and rural environmental hygiene, comprehensively improving the environmental hygiene level, enhanced the joint control ability of infectious diseases, and effectively reduced the risk of disease transmission.

HEV RNA detection is widely regarded as the “gold standard” for the diagnosis of hepatitis E infection. Previous studies have shown that there are significant differences in the positive rate of HEV RNA among blood donors in different cities and regions. For example, it is 0.26% (3/1144) in Nanjing, 0.13% (8/6263) in Xiamen, and 0.002% (1/46,837) in Shangrao. A Meta-analysis in 2023 showed that the pooled prevalence rate of HEV RNA in unpaid blood donors in China was 0.028% [9] . In this study, no HEV RNA positive samples were detected among the 95 HEV-IgM positive samples and 7 HEV-Ag positive samples were sent for inspection. There may be several reasons for this: First, a positive antibody and negative nucleic acid may indicate that the blood donor is in the recovery period of HEV. At this time, the HEV RNA has turned negative, but the antibody still exists in the body due to its long half-life. Some studies have shown that IgM antibodies can persist for five months or even up to three years after the recovery of hepatitis E [20] [21] . Second, a positive antigen and negative nucleic acid may be due to the blood donor being in the early stage of infection with a low viral load. Since hepatitis E is an RNA virus and is easily degraded, HEV RNA cannot be detected in the body. Third, the possibility of false positives of antigen and antibody cannot be excluded. Due to the large mobility of blood donors, some blood donors are not in the local area all year round. This study did not conduct follow-up tests on this group of people. Future research can increase the follow-up monitoring of such people. Fourth, the number of specimens sent for inspection is limited, and the scope of HEV nucleic acid detection is small, so the HEV RNA infection situation in Qinzhou City cannot be comprehensively evaluated.

Many studies suggest that an elevated alanine aminotransferase (ALT) level may be a non-specific marker of hepatitis E virus (HEV) infection. The results of this study show that there was no statistically significant difference in the positive rate of HEV-IgG between the normal ALT group (11.61%) and the elevated ALT group (9.52%) ((P > 0.05)). Similarly, there was no significant difference in the positive rate of HEV-IgM between the normal ALT group (0.73%) and the elevated ALT group (0.79%) ((P > 0.05)), and the difference in the positive rate of HEV-Ag between the normal ALT group (0.054%) and the elevated ALT group (0%) was also not statistically significant ((P > 0.05)). The results of this study are similar to those of You Qingzhu et al. [12] and Zhang Longmu et al. [19] , but differ from other reports [11] [17] . The reasons for these differences may be as follows: First, an elevated ALT level is not a sensitive marker of HEV infection. Second, all samples in this study passed the initial ALT screening before blood donation. Even if the test results show an elevated ALT level in the laboratory, in most cases, the ALT value is only 1 - 2 times the normal value. There are numerous factors affecting ALT within this range, and the clinical significance is limited [11] . The research results indicate that routine ALT testing can screen out some HEV-infected individuals, but there are still HEV-infected individuals among blood donors with normal ALT levels. Therefore, relying solely on ALT testing cannot effectively prevent the transmission of HEV through blood.

To understand the population distribution characteristics and risk factors of HEV infection among unpaid blood donors in the Qinzhou region, this study analyzed factors such as the gender, age, ethnicity, occupation, and educational level of blood donors positive for HEV-IgG/IgM/Ag. The results show that the infection rates of HEV-IgG/IgM among blood donors in this region are higher in males than in females ((P < 0.05)). This may be related to the fact that males have more opportunities for infection, such as greater job mobility, more social interactions, and certain living habits. This result is consistent with the studies of Li Meilin [10] , Liu Qiaomei [13] , Li Wei [16] , etc. However, studies by Yu Qin [11] , You Qingzhu [12] , etc., suggest that gender is not an independent risk factor for HEV infection. The inconsistent conclusions may be due to factors such as differences in the study population, regional public health conditions, reagent variations, sample size, and statistical methods. Age is considered a risk factor for HEV infection in this region. With the increase in age, the positive rates of HEV-IgG/IgM/Ag show an upward trend. The positive rate of IgG in the age group over 46 years old reaches 26.91%, which is consistent with the finding reported by Hu Liping [22] et al. that the incidence of hepatitis E virus in Guangxi is higher in middle-aged and elderly people. This may be because the older a person is, the more life experiences they have, and the greater the risk of exposure to HEV infection. In addition, the older group may have a weakened immune function and reduced resistance to the virus, making them more susceptible to HEV infection.

In terms of ethnicity, this study found that the HEV infection rates among blood donors of certain ethnic groups are relatively high, which may be related to the living habits, dietary structures, and hygiene conditions of different ethnic groups. For example, some ethnic groups may be more inclined to eat raw or undercooked foods, which may increase the risk of HEV transmission. In terms of occupation, the research results show that certain occupations (such as farmers and fishermen) are significantly associated with the risk of HEV infection. This may be related to the fact that people in these occupations are more frequently exposed to potential virus sources, such as through contact with contaminated water sources or consumption of contaminated foods.

The analysis results in terms of educational level showed that the HEV positive rates among blood donors with lower educational levels are relatively high. This may be related to the lack of health awareness, hygiene habits, and preventive measures among people with low educational levels. Improving public health awareness and the level of health education, especially among high-risk groups, may help reduce the transmission of HEV.

The limitations of this study are that the sample size is relatively limited and only covers blood donors in Qinzhou City, which may not fully reflect the HEV infection situation in the entire region. In addition, the research design is a cross-sectional survey, and the long-term health conditions of infected individuals cannot be tracked. Future research can consider conducting long-term longitudinal studies to more comprehensively understand the epidemiological characteristics and influencing factors of HEV infection.

In conclusion, although the overall infection rate of HEV among unpaid blood donors in the Qinzhou region is low, the study reveals the characteristics of HEV infection in terms of gender, age, ethnicity, and occupation in this region, as well as the potential risks of HEV-infected individuals with normal ALT levels. Therefore, it is necessary to strengthen the screening and monitoring of high-risk groups, improve public awareness, and prevent HEV infection to reduce the risk of the virus’s transmission through blood.

There is a potential risk of hepatitis E virus (HEV) transmission through blood transfusion in the Qinzhou region. Although the overall prevalence of HEV in this region is relatively low in China, there are still high-risk groups, and the current blood screening strategy has not effectively prevented the blood-borne transmission of HEV. The study found that male blood donors over 45 years old have a relatively high infection rate, and the infection rates of ethnic minority blood donors such as the Miao and Dong ethnic groups are higher than that of the Han ethnic group. To ensure blood safety and reduce the risk of HEV transmission through blood, the following effective measures are recommended: First, strengthen publicity and education, expand the publicity targets, especially health education for middle-aged and elderly people, to enhance public health awareness, prevent “disease from entering through the mouth”, and cut off the transmission routes. Second, when recruiting blood donors, priority should be given to recruiting blood donors with a low risk of HEV infection. Finally, for the high-risk HEV-infected population in this region, it is recommended to selectively increase the detection of HEV infection markers, and conduct appropriate epidemiological investigations and etiological monitoring of hepatitis E among the low-educated and middle-aged and elderly groups, so as to understand the infection status, transmission routes, and risk factors of the high-risk population, and provide a basis for scientific prevention and control measures and blood safety.

This study has several limitations. First, the selection of samples may affect the generalizability of the results, especially in some specific populations, and may not fully reflect the HEV infection situation. Second, this study relies mainly on the cross-sectional survey method, which may not fully capture the dynamic changes of HEV infection. In addition, the time and geographical limitations of data collection may also affect the accuracy of the results, resulting in the failure to fully identify the infection trends of some high-risk groups. Future research should consider introducing more variables and a wider range of samples to improve the reliability and applicability of the research results.

During the conduct and completion of this study, we received support and assistance from many people. Here, we would like to especially thank the medical institutions and blood donors in the Qinzhou region for their valuable data and support. We also thank every member of the research team for their hard work in data collection, analysis, and discussion. In addition, we are grateful to the institutions that funded this study and all the participants for their active cooperation, which has ensured the smooth progress of this study. Finally, we thank our family and friends for their understanding and support during the research period.

Scientific Research Project of the Health Commission of Guangxi Zhuang Autonomous Region (NO.: Z-N20221921).

*First author.

^#Corresponding author.