Hepatitis D is associated with faster liver disease progression than other viral hepatitis. Although clinical guidelines recommend single-step diagnosis for HBsAg-positive cases, we show that it was not implemented in many hospitals in our geographical area during the retrospective phase of our study. The retrospective phase results showed underdiagnosis of HDV, with only a fifth of hepatitis B patients undergoing anti-HDV testing from 2018 to early 2022. This highlights the need to implement these recommendations to identify undiagnosed infected patients. Similarly, the implementation of reflex testing tripled the detection of chronic hepatitis D patients compared to the previous phase. With the reflex test strategy, the percentage of undiagnosed HDV RNA-positive patients is significantly lower than with clinical practice. This further emphasizes the importance of reflex testing in early HDV infection detection, which can have significant implications for the clinical management of co-infected patients.
Our analysis also underscores the need for accurate prevalence data to formulate effective strategies to identify undiagnosed co-infected patients. Comparing our prevalences with those from another Spanish study that also used reflex testing for HBsAg-positive patients reveals differences (6% vs 8.1% retrospective and 4.7% vs 9.6% prospective)20. In the retrospective phase, our study may have underestimated prevalence as delta determination was only included in 18% of HBsAg-positive patients. Likely, testing was only done for patients with HDV risk factors, justifying the difference.
Additionally, comparing our prospective phase results with those in a recently published article using systematic review and meta-analysis methods to estimate HDV prevalence in 25 countries shows significant variability among European countries23. England, France, and Germany reported anti-HDV prevalences between 2.9% and 3.5%, lower than observed in our study. However, the estimated percentages of HDV RNA-positive cases were higher at 50%, 75%, and 60%, respectively. In the same article, anti-HDV prevalence was reported at 8.3% in Italy and 12.6% in Portugal, with adjusted prevalences of 3.4% and 1.5%, both lower than our study. Data on HDV RNA-positive patients were also significantly higher at 61% and 73%. The prevalences found in this study are higher than those adjusted for risk factors such as population, geographic region, or detection methods23,24. Ignoring these factors can lead to overestimating the actual prevalence and distorting the problem’s magnitude, possibly contributing to the decision not to allocate healthcare resources to effective strategies. Comparing prevalences between countries highlights the importance of understanding HDV epidemiology in each context to improve HBV/HDV co-infection diagnosis.
There is a scarcity of studies analyzing the impact before and after implementing reflex testing on HBV/HDV co-infection detection. Comparing the percentages of HBsAg-positive patients undergoing specific HDV tests in our study with those from another Spanish study shows differences in the previous phase (18% vs 7.6%), but alignment with the later phase (96%)20. This study concluded that reflex HDV testing would increase the number of diagnosed Chronic Hepatitis D cases 9 times over eight years22. The difference from our analysis is primarily due to the higher prevalences used and the longer analysis horizon. Another study that evaluated this strategy’s potential benefits based on a review of published epidemiological data in countries with different prevalence levels, including Spain, concluded that it is effective for identifying undiagnosed individuals, especially in countries with low HBsAg prevalence, such as those in the European Union24. Other studies related to hepatitis C also support the strategy’s effectiveness25.
The economic analysis assessed the cost impact associated with the diagnosis of HDV in HBsAg + patients by incorporating the dual reflex test in southern Spain and does not consider the follow-up of the remaining HBsAg + patients. Given the limited amount of information in published literature, this study contributes epidemiological data and demonstrates the efficiency of its implementation, resulting in a significantly lower cost per detected viremic patient than clinical practice. This is due to avoiding unnecessary visits for result confirmation and duplicate tests. These results align with other HDV and HCV studies22,26.
Moreover, implementing double reflex testing for HDV could change the clinical management of patients in the long term, not only in terms of diagnosis but also in resource planning and public health policies. Double reflex testing would allow early identification of HBV/HDV co-infected patients, giving them a treatment opportunity. Previously, patients were treated with Peg-IFN, a treatment with low response rates and many contraindications, but recently, Bulevirtide, a drug with better tolerability and greater effectiveness, has been approved in many countries11. Early treatment prevents the development of severe liver complications and improves patients’ quality of life27. In addition, a modeling showed that reflex HDV testing would reduce cases of cirrhosis and hepatocellular carcinoma and even liver-related mortality, resulting in savings associated with preventing liver complications22. This underscores the need for strategies that promote diagnosis. Although our study did not analyses the impact beyond diagnosis, increasing the number of HDV-RNA-positive patients detected would improve patient health outcomes. Additionally, identifying and treating the infection helps reduce virus transmission, contributing to preventing new infections. These tests are well-accepted by patients28, which could facilitate detection in hard-to-reach at-risk populations, such as drug users and migrants.
Due to the observational nature of our study, its primary limitation lies in its retrospective design, as well as its “localized” scope, given that the data were collected exclusively from Andalusia. This may influence the prevalence rates and costs included in the analysis. However, sensitivity analyses conducted with the different prevalence found in both phases can provide a broader view of the results. Regarding costs, sensitivity analyses were not performed since the actual costs according to the Andalusian Public Health System were included, aligning with our objective. Lastly, regarding the test methodology, it is noteworthy that currently, the diagnosis and monitoring of treatment response are based on HDV RNA detection and quantification. These tests are not yet standardized for HDV, and results vary between laboratories due to different sensitivities and RNA extraction methodologies, making them non-comparable29,30. Therefore, advancing more sensitive and specific HDV RNA quantification methods is necessary to improve co-infection detection and management. Additionally, longitudinal studies are needed to track the progress and outcomes of patients diagnosed using this strategy. In addition, although our study was conducted in Andalusia rather than across the entire country, and some degree of selection bias cannot be completely ruled out, we believe it is unlikely to significantly affect the generalizability of our findings. Andalusia is the most populous autonomous community in Spain, representing approximately 20% of the national population, which supports the representativeness of the studied population within the broader Spanish context.
In conclusions, the prevalences found indicate a high percentage of HDV-infected patients in southern Andalusia and provide valuable information on the burden of HDV co-infection. The introduction of double reflex testing for HDV in HBsAg-positive patients reduces the underdiagnosis of the infection, significantly increasing the detection of hidden chronic hepatitis delta patients. Moreover, it simplifies infection diagnosis and generates savings for the healthcare system, being an efficient strategy that should be considered for the care of chronic hepatitis B patients.
