Screening and identification of DNA aptamers toward Schistosoma japonicum eggs via SELEX.

Schistosomiasis is a major parasitic disease caused by blood flukes of the genus Schistosoma. It affects the liver, mesentery, and urogenital tract of infected individuals. The World Health Organization (WHO) estimates that more than 200 million people are currently infected by this disease worldwide, resulting in a mortality rate exceeding 200,000 each year1. Most human schistosomiasis is caused by Schistosoma haematobium (S. haematobium), Schistosoma mansoni (S. mansoni ) and Schistosoma japonicum (S. japonicum). Among these, S. japonicum is the major human blood fluke that occurs in Asia, particularly in China, the Philippines, Thailand, and Indonesia2. The ovulation rate of S. japonicum is approximately 2200 eggs daily per female adult worm. Schistosoma eggs can be deposited in the host organs, which causes the symptoms of schistosomiasis derived from host immune response to eggs trapped in tissues Egg-released antigens stimulate a granulomatous reaction involving T cells, macrophages, and eosinophils that results in clinical disease, such as granuloma and fibrosis3,4. Schistosoma eggs can be emitted into water with the host feces, which is the beginning of schistosomiasis transmission. The current in vitro diagnostic method involves the demonstration of schistosome eggs in the stool, and it remains the gold standard for the diagnosis of schistosomiasis and the technique of choice for testing candidate drug efficacy. However, low-intensity infections reduce the sensitivity of other tests, such as the Kato-Katz thick smear method5. Although antibody-based detection techniques may be more sensitive, cross-reaction with other helminth infections often results in lower specificity, and these methods are also more sophisticated and expensive6,7. Aptamers are short single-stranded oligonucleotide molecules (RNA or ssDNA) that undergo conformational changes that allow folding into unique tertiary structures able to bind specific cell-surface targets8. Similar to antibodies, aptamers possess high-binding affinity for their targets, typically from picomolar to nanomolar levels, depending on the nature of targets9. Comparable to antibodies, aptamers also recognize their targets with high specificity. For instance, aptamers can discriminate among homologous proteins that contain only a few amino acid changes10. In addition, aptamers exhibit some unique features over antibodies, including low molecular weight, high stability, easy and controllable synthesis and modification for different diagnostic and therapeutic purposes, lack of immunogenicity, rapid tissue penetration, and nontoxicity11. Owing to these molecular properties, aptamers have been widely employed as a novel tool for diagnosis and treatment of disease. Aptamers can be generated from a synthetic random library with 1013–1016 ssDNA or ssRNA molecules by an in vitro iterative selection process called systematic evolution of ligands by exponential enrichment (SELEX)12,13. Over the last 20 years, aptamers have been successfully isolated for a variety of targets, ranging from small molecules, such as metal ions, organic dyes, amino acids, or short peptides, to large proteins or complex targets, including whole cells, viruses, bacteria, or tissues14,15,16. Especially, the selection against complex targets can be carried out without knowing the distinct molecular signature in individual target cells, suggesting that these molecular tools can support improved clinical diagnosis and treatment, as well as speed up the discovery of new biomarkers17. In this study, we employed egg-based systematic evolution of ligands by exponential enrichment (SELEX), resulting in the identification of a panel of ssDNA aptamers specifically binding to eggs derived from S. japonicum. Among these, aptamers LC6 and LC15 exhibited strong binding affinity and specific recognition of S. japonicum eggs. Furthermore, tissue imaging results revealed that aptamer LC15 could recognize S. japonicum eggs laid in liver tissues with a detection ratio of 80.5%. Collectively, therefore, we obtained novel aptamers specifically recognizing S. japonicum eggs, which will facilitate the development of an effective tool for both schistosomiasis diagnosis and drug delivery.