Introduction
Viral hepatitis poses a serious threat to public health. In 2015, viral hepatitis was responsible for 1.34 million fatalities, which is more than HIV-related mortality and on par with TB deaths. It is estimated that 1.75 million new cases of HCV infection occurred in 2015, bringing the overall number of persons living with the HCV to 71 million.1 The Hepatitis C virus (HCV) is a linear, single-stranded, positive-sense RNA virus belongs to the Flaviviridae family.2 The transmission of HCV can occur via the parental route, as well as through perinatal, sexual, and needle-sharing routes, percutaneous exposure to infected blood products, and organ transplants from infected donors.3
Chronic HCV infection establishes itself when the body fails to eliminate the virus after an initial acute phase. This persistent infection can silently progress for years, often without noticeable symptoms, until significant liver damage occurs. According to estimates, 6 to 12 million people are living with chronic hepatitis C. Chronic HCV infection contributes to 12–32% of HCC and 12–20% of cirrhosis cases.4
The World Health Organization (WHO) estimates that roughly 50 million people globally live with chronic HCV, with approximately 1.0 million new infections occurring annually.5
The burden of this disease is substantial, impacting healthcare systems and individual well-being. With the availability of directly acting antiviral, chronic HCV case can be effectively cured. Sero-positivity does not reflect upon the active infection or chronicity of the HCV infection. Detection of viral RNA is required for demonstration of HCV active infection or chronicity.
This study focuses on assessing the prevalence of active infection among HCV antibody reactive (seropositive) individuals majority of whom are asymptomatic. Understanding the hidden burden of active diseases or chronic infections can provide significant insight that not only helps patients in receiving timely treatment but also influences public health strategies that aim to eliminate HCV infection by 2030.5
The prevalence of active infection in seropositive individuals for HCV has not been extensively studied in our region. Most of the studies have focussed on assessing the seroprevalence of HCV only. Hence this study will also function as a reference for future researchers.
Material and Methods
This retrospective study was conducted at Model diagnostic centre, Microbiology Department, GMERS Medical College, Sola, Ahmedabad, from January 2020 to March-2024.
Sample size
The sample size consisted of plasma samples from 763 HCV seropositive individuals received from various districts of Gujarat that were reactive for anti HCV Antibody.
Sample type
3-5 ml of whole blood was collected in a sterile EDTA vacutainer. Plasma separation was done by centrifugation within 6 hours of collection of whole blood. Plasma was stored at -20℃ until testing.
Methods
Plasma samples were tested with quantitative real time RTPCR assay which allowed detection of RNA. Results were available as number of copies and then were converted to internationally accepted quantification in IU/ml. RNA extraction was done using column based kit QiaAmp viral RNA Mini Kit (QIAGEN) and on Automated RNA extraction system (Qiasymphony. RTPCR kit of Altona, Altostar HCV RTPCR 1.5 kit and Artus® HCV RG were used. RNA detection by RTPCR was considered a case of active infection.
Result
Out of 763 seropositive samples tested, HCV RNA was detected in 361samples with a prevalence rate of 47.31% and in 368 samples HCV RNA was not detected. In 34 samples, results were inconclusive and a repeat sample was requested anyhow, fresh repeat samples were not received. (Table 1)
There was higher positivity in males than in females. 63.99% samples from male cases and 38.78% samples from female cases were positive. (Table 2)
The age-wise distribution of the patients is shown in table 3. The majority of the cases belong to age group between 41 to 60 years (41.55%) followed by 21-40 years (38.78%). In extremes of age groups is more than 60 years and between 0-20 years positivity was less ie 14.96% and 4.7% respectively. (Table 3)
hows the region wise distribution of active infection status. Samples were received from 18 different cities of Gujarat falling under various zones like North, South, Central and West Gujarat. North Gujarat region showed prevalence of 47.15% while western Gujarat, central Gujarat and south Gujarat showed a prevalence of 48.20%, 40.20% and 79.16% respectively. Within the regions, diversity was seen. In majority of the cities, positivity ranged from 40-50%. However some regions likeMehsana, Patan, Surat and Valsad showed high positivity of 75 %, 76.92% and 81.81% respectively. However positivity was very low in some regions like Bharuch and Gandhinagar.
Table 1
Prevalence of active Hepatitis C Virus Infection
|
|
Number of Cases |
Percentage |
|
Total number of Seropositive Cases |
763 |
|
|
HCV RNA detected |
361 |
47.31% |
|
HCV RNA not detected |
368 |
48.23% |
|
Inconclusive |
34 |
4.46% |
Table 2
Gender wise distribution of cases with active infection ( n=361)
|
|
Number of Cases |
Percentage |
|
Male |
231 |
63.99% |
|
Female |
140 |
38.78% |
Table 3
Age group wise distribution of cases with active HCV virus infection (n=361)
|
Age Group (in yrs.) |
Number of Cases |
Percentage of cases |
|
0-20 yr. |
17 |
4.7% |
|
21-40 yr. |
140 |
38.78% |
|
41-60 yr. |
150 |
41.55% |
|
>60 yr. |
54 |
14.96% |
Table 4
Region wise distribution of cases with active infection ( n=361)
Discussion
Hepatitis C virus infection is ongoing challenge in India and might have long-term complications showing major contribution of liver illness in different parts of India. Serological tests like ELISA or rapid test that detect antibodies to HCV are used for screening purposes. Apart from this, during the early stage of acute HCV infection, there is a window period when these tests may not detect the antibodies, resulting in a false negative result. False negative results are common in patients with chronic renal failure receiving haemodialysis, individuals with HIV coinfection, patients receiving chemotherapy, steroid treatment, immunosuppressive medication and new-borns. 6, 7, 8
Sero-positivity does not reflect upon the active infection in acute stage or chronicity in patient. In such cases, RTPCR testing for HCV RNA is considered the standard method for detecting the infection as well as monitoring the treatment. Hence antibody detection helps only for screening purpose of hepatitis C virus infection. RTPCR should be done in such cases to diagnose active hepatitis C virus infection. 8
In the present study from the 763 HCV seropositive positive patients, 47.31% patients had shown HCV RNA confirming the Hepatitis C virus active infection and 48.23% patients were negative for HCV RNA. This is at par with established data of chronicity for HCV that ranges from 50-80% after acute infection. The similar findings is shown by a study by Qamar Z et al. 9 from Buner district in Pakistan showing 34.1% positivity. Another studies by Vannavada Sudha Rani et al from Telangana in dialysis patients and by M S Agular in Venezuela showed positivity in 41.6% and 41% respectively. 10, 11 However a very high positivity was demonstrated from different regions of India like in Kolkata and Secundarabad demonstrating 85.75%, 60.86% respectively. 12, 13 High positivity for HCV RNA is demonstrated across globe. Various studies across globe showed high positivity of 59.6%, 62.5%, 79.5%, 74%, 75.9% in Rwanda, Khyber Pakhtunkhwa Province of Pakistan, Egypt, United States and Southern Italy respectively. 14, 15, 16, 17, 18
Though overall burden of active infection is 47.31%, four different regions of Gujarat have shown variation in presence of active HCV infection that ranges from 40% to 79%. South Gujarat showed highest prevalence of 79.16%. This might be due to relatively lesser number of samples received and tested. Western Gujarat showed a burden of active infection in 48.2% cases however Jamnagar district in this region had a higher burden of 64.71% than other districts in western region. In Central Gujarat, highest cases were seen in Vadodara at 40.20% while Bharuch demonstrated a very low rate of 14.3%.district. North Gujarat had a prevalence of 47.15% with maximum burden in Patan (75%), this again could be due to small sample size. Mehsana, Ahmedabad and Gandhinagar had a prevalence rate of 75%, 51.41% and 28.3% respectively.
There is significant proportion of active infection in the study and regional variation is clearly visible in the study.This emphasizes the role of confirmation and assessment of active infection after serodiagnosis. Region wise distribution of disease helps to plan resource allocation, targeted intervention, monitoring trends and better management of patient.
Among 361 active infection cases 63.99% were males and 38.78% were females. Similar data is revealed by studies from Kolkata 12 64.21% male and 35.79% female and Khyber Pakhtunkhwa Province, Pakistan 15 57% male and 43% female. Study by Deepjyoti K et al. 19 also showed that males were more affected than females. In contrast, another study from Qamar Z et al. 9 from Buner district in Pakistan concluded that prevalence was more in female (51.93%) than male (48.06%). This also suggests the geographical variation and local practices.
In the present study, people in age group 41-60 years showed highest burden of active hepatitis C virus infection followed by age group 21-40 years, >60 years and 15-20 years. Studies from Rwanda 14 showed high prevalence in >55 years. Similar finding were reported by study Deepjyoti K et al. 19 which showed higher prevalence in 40-60 years of age group.Study by Alter MJ in U.S et al. 17 HCV infection showed a high prevalence in individuals aged 20-40 years. In Mardan, Pakistan 20 showed high prevalence 41-50 years in female and 31-40 in male. It might be due to greater exposure to various HCV risk factors and also due to late detection in adulthood as disease runs a silent course for many decades.
Conclusion
ELISA are commonly used for initial HCV screening because they're fast and cost-effective. However, these tests detect antibodies against HCV, which indicate past or present exposure, but not necessarily active infection. Significant number of seropositive individuals are not able to clear virus, have active infection despite appearance of antibodies and progress to chronicity and complication of disease. HCV RNA Testing by RTPCR directly detects the presence of viral RNA in the blood, signifying an active infection. It is a sensitive and specific test for the diagnosis of hepatitis C virus infection both in acute and chronic stage. Presence of HCV RNA makes the individual eligible for the treatment. It’s an early predictor of liver damage risk in seropositive individuals. Early treatment with direct acting antiviral agent (DAA) is very effective as compared to previous regime and thus helps to reduce chronicity, malignancy and associated mortality and morbidity of hepatitis C virus infection. The sustained viral response after a successful treatment as informed by RTPCR demonstrates clearance of virus. Hence routine testing with RNA RTPCR following sero-positivity is the need of the hour and a must do diagnostic test.
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