Transcriptional Changes in Schistosoma mansoni during Early Schistosomula Development and in the Presence of Erythrocytes Geoffrey N. Gobert 1 , Mai H. Tran 1 , Luke Moertel 1 , Jason Mulvenna 1 , Malcolm K. Jones 2 , Donald P. McManus 1 , Alex Loukas 1 * 1 Division of Infectious Diseases, Queensland Institute of Medical Research, Herston, Queensland, Australia, 2 School of Veterinary Sciences, The University of Queensland, Brisbane, Queensland, Australia Abstract Background: Schistosomes cause more mortality and morbidity than any other human helminth, but control primarily relies on a single drug that kills adult worms. The newly transformed schistosomulum stage is susceptible to the immune response and is a target for vaccine development and rational drug design. Methodology/Principal Findings: To identify genes which are up-regulated during the maturation of Schistosoma mansoni schistosomula in vitro, we cultured newly transformed parasites for 3 h or 5 days with and without erythrocytes and compared their transcriptional profiles using cDNA microarrays. The most apparent changes were in the up-regulation of genes between 3 h and 5 day schistosomula involved in blood feeding, tegument and cytoskeletal development, cell adhesion, and stress responses. The most highly up-regulated genes included a tegument tetraspanin Sm-tsp-3 (1,600-fold up-regulation), a protein kinase, a novel serine protease and serine protease inhibitor, and intestinal proteases belonging to distinct mechanistic classes. The inclusion of erythrocytes in the culture medium resulted in a general but less pronounced increase in transcriptional activity, with the highest up-regulation of genes involved in iron metabolism, proteolysis, and transport of fatty acids and sugars. Conclusions: We have identified the genes that are up-regulated during the first 5 days of schistosomula development in vitro. Using a combination of gene silencing techniques and murine protection studies, some of these highly up-regulated transcripts can be targeted for future development of new vaccines and drugs. Citation: Gobert GN, Tran MH, Moertel L, Mulvenna J, Jones MK, et al. (2010) Transcriptional Changes in Schistosoma mansoni during Early Schistosomula Development and in the Presence of Erythrocytes. PLoS Negl Trop Dis 4(2): e600. doi:10.1371/journal.pntd.0000600 Editor: Elodie Ghedin, University of Pittsburgh, United States of America Received October 2, 2009; Accepted December 17, 2009; Published February 9, 2010 Copyright: ß 2010 Gobert et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by grants 496600 and 290247 from the National Health and Medical Research Council, Australia (NHMRC). AL is supported by a senior research fellowship from NHMRC. MT was supported by a U.S./Australia Fulbright award. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: Alex.Loukas@qimr.edu.au Introduction The schistosome tegument, an unique double lipid bilayered syncitium that covers the external surface of the intra-mammalian developmental stages, represents the point of interaction between the parasite and mammalian host tissues. This structure is pivotal for parasite survival within the host and is therefore a primary target of anthelmintic drugs [1] and vaccines [2,3]. In similar fashion, the intestine, or gastrodermis of schistosomes is a source of secreted proteins and another point of interaction with host tissues (i.e. blood). The Schistosoma mansoni genome sequence has recently been reported [4] and the secreted proteome (secretome) has also been characterised with a major focus on the proteins present in the tegument and excretory/secretory (ES) products [4,5,6,7,8,9]. While the schistosome gastrodermal proteome has not yet been explored, we recently described tissue-specific gene profiling for adult S. japonicum and characterised the transcriptome of gastro- dermal cells using a combination of laser microdissection microscopy followed by cDNA microarray analysis [10]. Despite the progress made in characterising the mRNA and protein compositions of cells at the host-parasite interface, it is only now with the recent application of gene silencing technologies to the study of schistosomes, that we are understanding the functions of these proteins and how they enable schistosomes to exist as parasites [11]. In terms of vaccine development, the newly transformed schistosomulum is widely viewed as the most susceptible stage to antibody-mediated damage [2,3,12,13,14]. After cercariae trans- form into schistosomula, parasites undergo changes in their surface protein composition [15]. The schistosomula surface is dynamic, with some proteins appearing and others disappearing [16] as the parasites mature during their migration to the lungs. Once the parasites reach the lungs they are refractory to antibody-mediated damage [17] and cloak themselves in host blood group antigens [18] and other proteins involved in immune responses [19,20]. Obtaining sufficient quantities of schistosomula directly from lung tissue for most research purposes is time consuming and involves working with mammalian hosts. As a result, many www.plosntds.org 1 February 2010 | Volume 4 | Issue 2 | e600