ABSTRACT
Objectives
Hepatitis C virus (HCV) infection is a public health issue of great importance. HCV genotyping helps in monitoring prognosis, selecting appropriate antiviral drugs, monitoring side effects, and deciding on treatment duration. The aim of this study was to determine the HCV genotypes in our hospital to reveal their distribution over time and to contribute to epidemiological data by evaluating the relationship between HCV genotypes and viral load.
Materials and Methods
Serum samples from 144 patients diagnosed with chronic hepatitis C between January 01, 2019 and December 31, 2023 were included in this study. HCV-RNA loads were determined using a Bosphore quantification kit (Anatolia, Turkey) via a Montania 4896 thermal cycler (Anatolia, Turkey). HCV genotypes were detected using the Bio-Rad CFX96 system with the Diagnotech HCV genotyping quantitative polymerase chain reaction kit targeting the 5'NS5B region.
Results
The most frequently detected genotypes in our hospital were genotypes 1b (34.7%, genotype 3 with 32.6% and genotype 1 with 15.3%). The lower mean age of genotype 3 patients was statistically significant compared with the ages of patients with genotypes 4 and genotype 1b (p<0.001). It was found to be statistically significant that the median viral load of patients with genotype 1 was lower than that of patients with genotype 1b and genotype 3 (p=0.046). The higher frequency of genotype 4 among foreign nationals was statistically significant (p=0.034).
Conclusion
HCV genotypes vary between regions based on geographical location, migration, socioeconomic level, and drug use. Therefore, molecular studies on this issue are important for appropriate antiviral drug treatment and duration.
Introduction
Hepatitis C virus (HCV) infection may cause acute and chronic HCV infections and lead to complications, such as liver failure, cirrhosis, and hepatocellular carcinoma, with high mortality and morbidity (1). HCV infection is a major health problem worldwide, and approximately 85% of acute HCV infections progress to chronic HCV infection. The World Health Organization reports that approximately 58 million people worldwide are affected by chronic HCV infection, and approximately 1,5 million people become infected with HCV each year (2, 3). In Turkey, approximately 1 million people are infected with HCV.
HCV is usually transmitted through sexual intercourse, intravenous drug use, blood transfusion, and surgical and dental procedures (4). The most important characteristic of HCV infection is that it can cause chronic HCV infection and become resistant to antiviral drugs as a result of mutations in host cells (5). Different HCV genotypes exist due to differences in nucleotide sequences among regions of the HCV genome (6). The nucleotide sequences of HCV genotypes differ by 30-35%, and there is at least a 15% difference between the nucleotide sequences of the subgroups of HCV genotypes (5, 7). The main factors affecting HCV genotype distribution are socioeconomic level, geographical location, migration, and intravenous drug use.
The most common HCV genotype worldwide is genotype 1 with a rate of 46-49%, followed by genotype 3 at 22% (8, 9). When the distribution in the world is analyzed; genotype 1 is more common in North and South America, genotype 2 in East Asia, genotype 3 in Asia and Europe, genotype 4 in the Middle East and North Africa, genotype 5 in South Africa, and genotype 6 in Southeast Asia (10). HCV genotyping helps in monitoring prognosis, selecting appropriate antiviral drugs, monitoring side effects, and deciding on the treatment duration (11).
The aim of this study was to determine the HCV genotypes in our hospital, monitor their distribution over the years, and contribute to the epidemiological data by revealing the relationship between HCV genotypes and viral load.
Materials and Methods
Serum samples taken from 144 patients diagnosed with chronic HCV infection between January 01, 2019 and December 31, 2023 were included in this study. HCV-RNA levels were determined using a Bosphore quantification kit (Anatolia, Turkey) via a Montania 4896 thermal cycler (Anatolia, Turkey). Results are expressed as International units per milliliter (IU/mL). HCV genotypes were detected on a Bio-Rad CFX96 PCR thermal cycler (California, USA) using a Diagnotech HCV genotyping qPCR kit (Diagnotech, Turkey) targeting the 5'NS5B region.
This retrospective study was approved by the Ethics Committee on Non-Medicine and Non-Medical Device Research of Necmettin Erbakan University (decision number: 2024/4963, date: 17.05.2024).
Statistical Analysis
The data obtained were analyzed using Statistical Package for Social Sciences 21.0 package program. In descriptive analyses, frequency data were presented as number (n) and percentage (%), whereas numerical data were presented as median (minimum-maximum). The conformity of numerical data to the normal distribution was analyzed by visual (histogram) and analytical methods (Kolmogorov-Smirnov test). For numerical variables that were found not to conform to normal distribution, the Kruskal-Wallis test was used to compare more than two independent groups. Post-hoc Mann-Whitney U test and Bonferroni correction were performed for pairwise comparisons between groups with statistically significant differences. The chi-square (χ2) test and Fisher’s exact test were used to compare categorical variables (statistical significance p<0.05).
Results
Of the patients whose results were included in the study, 66.7% were male and 95.8% were Turkish nationals. Of the samples, 34.7% were obtained in 2019, 20.1% in 2021, 18.1% in 2022, 16% in 2020, and 11.1% in 2023. The most frequently detected genotypes were type 1b (34.7%), type 3 (32.6%) and type 1 (15.3%) (Table 1).
The median age of the patients included in the study was determined as 40.00 (29.00-71.00). The median viral load of the patients was determined as 190000 (12500-1519366.50) (Table 2).
When the median ages of the patients were compared according to the detected HCV genotypes, a statistically significant difference was observed (p<0.001). In the post-hoc analyses, the difference was attributed to the median age of patients infected with type 3 being lower than that of patients infected with type 4 and type 1b (Table 3).
A statistically significant difference was observed when comparing the median viral loads of patients according to the HCV genotypes detected in patients (p=0.046).
In the post-hoc analyses, the difference was attributed to the median viral loads of patients with type 1 being lower than those of patients with type 1b and type 3 (Table 4).
There was a statistically significant difference between the years in which the disease was detected according to HCV genotypes (p<0.001). In post-hoc analyses, a lesser number of cases were found in individuals with type 1 genotype in 2019, whereas a greater number of cases were found in 2022 and 2023. In post-hoc analyses, the number of individuals infected with the type 1 genotype was lower in 2019 but was higher in 2022 and 2023. A statistically significant difference was found between the nationalities of the patients based on the HCV genotypes, a statistically significant difference was found (p=0.034). In post-hoc analyses, the rate of infection with genotype 4 was lower in Turkish patients, whereas the rate was higher in foreign patients (Table 5).
Discussion
Hepatitis C infection is a significant public health issue due to its high rate of chronicity, potential for severe liver diseases, various modes of transmission, and the absence of effective vaccines (12). Diagnosis of HCV infection is based on the detection of HCV-specific antibodies using enzyme-linked immunosorbent assay (ELISA). If the ELISA result is positive; HCV viral core antigen or viral genomic RNA (HCV-RNA) must be tested to confirm the diagnosis. In 80-90% of patients, anti-HCV antibodies become positive six to twelve weeks after exposure (13). The identification of HCV genotypes is important for adjusting the dosage of antiviral agents, determining the duration of treatment, monitoring treatment response, and predicting patient prognosis (7, 14). Various studies were conducted to investigate the frequency of HCV genotypes in our region and country (Table 6). It has been reported that the majority of HCV infection in Turkey is caused by genotype 1, and its prevalence varies between 57.1% and 97.1% 15). The most common subtype in Turkey is genotype 1b, with a prevalence of 52.7%-97.4% (16, 17).
In our study, as in previous data, the most frequent genotype was genotype 1 at a rate of 54.9%, and the most common subtype was genotype 1b at a rate of 34.7%. When the relationship between age and genotype was analyzed, the lower age of genotype 3 patients compared with genotype 1b patients was statistically significant. According to the data in the literature, the ages of patients infected with genotype 1 were statistically significantly higher compared with those infected with other genotypes. (18, 19). In various studies conducted in Turkey, no statistically significant relationship was found between HCV-RNA load and genotype (15, 20).
In our study, the low viral loads of patients infected with genotype 1 were statistically significant compared with those infected with genotypes 1b and 3. We believe that the coronavirus disease 2019 pandemic may lead to variations in the viral loads of infections caused by different HCV genotypes, and the increase in strains that cannot be subtypes may also be related to the pandemic. Different results have been reported in studies investigating genotype distribution and gender association. While some studies have found that genotype 1b is more commonly observed in women and genotype 3 in men, other studies have established that there is no statistical significance between genotype and gender (4, 21, 22). In a study examining the HCV genotype distribution in Syria between 2004 and 2006, the most frequently detected genotype was 4, with a rate of 59% (23). In various studies conducted in Turkey, the frequency of genotype 4 has been reported as 0-11%. In these studies, it was stated that the frequency of genotype 4 was rapidly increasing in our country because of migration as a result of the war in Syria and that its surveillance was important because genotype 4 was more resistant to treatment (23, 24). In our study, we detected genotype 4 at a rate of 3.5%, similar to the literature data.
Treatment may fail in patients infected with mixed genotypes. In various studies conducted in our country, the prevalence of mixed genotype has been reported as 0-8.6% (7, 25, 26).
In our study, the mixed genotype rate was 4.2%, which was in line with the literature. In a study conducted in our hospital in 2019, the most common genotype was genotype 1b at a rate of 58.9%, and it was observed that there was no change in the most dominant genotype in our hospital (18).
Study Limitations
The retrospective nature of our study is one of the limitations due to the inability to identify risk factors and modes of transmission.
Conclusion
In conclusion, the detection of HCV genotypes is important in determining appropriate antiviral therapy, its duration, and monitoring prognosis. Because the genotype distribution in Turkey varies by region, more molecular epidemiological studies on HCV genotypes are required.
