Discovery of novel Schistosoma japonicum antigens using a targeted protein microarray approach

Abstract

BackgroundNovel vaccine candidates against Schistosoma japonicum are required, and antigens present in the vulnerable larval developmental stage are attractive targets. Post-genomic technologies are now available which can contribute to such antigen discovery.MethodsA schistosome-specific protein microarray was probed using the local antibody response against migrating larvae. Antigens were assessed for their novelty and predicted larval expression and host-exposed features. One antigen was further characterised and its sequence and structure were analysed in silico. Real-time polymerase chain reaction was used to analyse transcript expression throughout development, and immunoblotting and enzyme-linked immunosorbent assays employed to determine antigen recognition by antibody samples.ResultsSeveral known and novel antigens were discovered, two of which showed up-regulated transcription in schistosomula. One novel antigen, termed S. japonicum Ly-6-like protein 1 (Sj-L6L-1), was further characterised and shown to share structural and sequence features with the Ly-6 protein family. It was found to be present in the worm tegument and expressed in both the larval and adult worms, but was found to be antigenic only in the lungs that the larvae migrate to and traverse.ConclusionsThis study represents a novel approach to vaccine antigen discovery and may contribute to schistosome vaccine development against this important group of human and veterinary pathogens.

Specific humoral response of hosts with variable schistosomiasis susceptibility

“…Our previous work 26 has shown that ASC probes from the skin‐ and lung‐draining LNs of the rat were induced by the passage of migrating schistosome larval stages. In addition, our earlier preliminary protein microarray study detected some possible antigens recognized in the skin and lung immune compartments 27 . We have extended this work and attempted to identify parasite protein antigens that are unique to these important sites of parasite elimination and immune priming.…”

“…The time points in TC1 were based on observations that immune resistance in the rat has been shown to be highest between 4 and 8 weeks after re‐infection 19 , 60 . The time points in TC2 were selected to capture the local tissue response after schistosome larval migration through the skin and lung after the first and fourth day post infection, respectively, 19 and after allowing a further 5 days for an optimal LN response 26 , 27 , 63 . The cercarial challenge load for TC1 was determined from previous studies 8 , 19 and from preliminary experiments for TC2; 26 specifically, the higher cercarial challenge for the second infection in TC2 ensured that skin‐ and lung‐draining ASCs were adequately stimulated.…”

“…The protein microarrays were probed with mouse sera and rat BAL fluid, sera and ASC probes and IgG reactivity measured. As a consequence of variations in aims, different samples were used and a more rigorous probing and analysis strategy was applied compared with our preliminary protein microarray ASC study 27 . BAL fluid and ASC probes had lower antibody titers than sera with reactivity being at least 100‐fold lower for the ASC probes 26 , 27 and 50‐fold lower for BAL fluid (data not presented), and they were therefore used at lower dilutions (1:2 and 1:10, for ASC probes and BAL fluid, respectively, compared with 1:100 for sera).…”

“…Protein microarray feature data were pre‐processed and analyzed as described 27 , 32 but with some modifications. The raw SI was generated after local background correction of protein microarray features.…”

“…In this study, we describe the use of a schistosome‐specific protein microarray we have developed 23 , 25 to: (1) compare the specific antibody response between resistant and susceptible animal hosts; (2) examine the longitudinal serum antibody response in the resistant rat host during successive infections; (3) define the tissue‐specific local antibody response using antibody‐secreting cell (ASC) probes 26 , 27 and (4) discover new potential vaccine and diagnostic antigens. To achieve these objectives, time courses of rats and mice experimentally infected with S. japonicum were completed to collect antibody samples for microarray probing and to characterize the parasitological resistance of these two animal models, particularly the striking resistance to reinfection intrinsic to the rat.…”