Despite the widespread use of chemotherapy and other control strategies over the past 50 years, transmission rates for schistosomiasis have changed little. Regardless of the approach used, future control efforts will require a more complete understanding of fundamental parasite biology. Schistosomes undergo complex development involving an alteration of parasite generations within a mammalian and freshwater molluscan host in the completion of its lifecycle. Little is known about factors controlling schistosome development, but understanding these processes may facilitate the discovery of new control methods. Therefore, our goal in this study is to determine global developmentally-regulated and stage-specific gene expression in Schistosoma mansoni using Serial Analysis of Gene Expression (SAGE). We present a preliminary analysis of genes expressed during development and sexual differentiation in the mammalian host and during early larval development in the snail host. A number of novel, differentially expressed genes have been identified, both within and between the different developmental stages found in the mammalian and snail hosts.

SUMMARY Infection of the snail, Biomphalaria glabrata, by the free-swimming miracidial stage of the human blood fluke, Schistosoma mansoni, and its subsequent development to the parasitic sporocyst stage is critical to establishment of viable infections and continued human transmission. We performed a genome-wide expression analysis of the S. mansoni miracidia and developing sporocyst using Long Serial Analysis of Gene Expression (LongSAGE). Five cDNA libraries were constructed from miracidia and in vitro cultured 6- and 20-day-old sporocysts maintained in sporocyst medium (SM) or in SM conditioned by previous cultivation with cells of the B. glabrata embryonic (Bge) cell line. We generated 21 440 SAGE tags and mapped 13 381 to the S. mansoni gene predictions (v4.0e) either by estimating theoretical 3′ UTR lengths or using existing 3′ EST sequence data. Overall, 432 transcripts were found to be differentially expressed amongst all 5 libraries. In total, 172 tags were differentially expressed between miracidia and 6-day conditioned sporocysts and 152 were differentially expressed between miracidia and 6-day unconditioned sporocysts. In addition, 53 and 45 tags, respectively, were differentially expressed in 6-day and 20-day cultured sporocysts, due to the effects of exposure to Bge cell-conditioned medium.

Keywords Schistosoma mansoni; Nitric oxide (NO); Serial analysis of gene expression (SAGE); Extracellular superoxide dismutase (EC-SOD)Nitric oxide (NO)-related pathways potentially play at least two critical roles in schistosomes, the causative agents of schistosomiasis. First, these pathways may represent essential signaling cascades required for normal parasite physiology and survival. Second, NO-related pathways may also play an important role in parasite-host interactions. Several reports have demonstrated that platyhelminths have nitric oxide synthase (NOS) activity and that NO is likely acting as a signaling molecule in these organisms [1][2][3][4]. Furthermore, the host NO pathway may be involved in host defense against schistosome infection, though its precise role in vivo is not clear [5][6][7].Here we examine changes in parasite gene expression in response to exposure to exogenous NO in vitro. Prior studies have provided evidence of NO-related pathways in adult schistosomes [1,2,8]. However, the physiological role and downstream targets of NO have not been elucidated in adult worms. To assay NO-dependent changes in gene expression, we have used Long-SAGE (serial analysis of gene expression) [9], which provides both the identity of expressed genes and the relative levels of their expression.In Long-SAGE, a short 21 bp sequence tag from the most polyA proximal NlaIII restriction site of an mRNA molecule is used to uniquely identify the source gene from within the genome. Short sequence tags are sampled from all NlaIII-positive transcripts in a mRNA sample and are linked together to form long concatenated molecules that are cloned and sequenced. Quantification of all tags provides a relative measure of gene expression (i.e., mRNA abundance). Using SAGE, we have identified genes which respond to NO by changing expression levels, and we also show by RT-PCR that RNA encoding extracellular superoxide dismutase (EC-SOD, also referred to as signal peptide-SOD [10][11] Publisher's Disclaimer:This article was originally published in a journal published by Elsevier, and the attached copy is provided by Elsevier for the author's benefit and for the benefit of the author's institution, for non-commercial research and educational use including without limitation use in instruction at your institution, sending it to specific colleagues that you know, and providing a copy to your institution's administrator. All other uses, reproduction and distribution, including without limitation commercial reprints, selling or licensing copies or access, or posting on open internet sites, your personal or institution's website or repository, are prohibited. For exceptions, permission may be sought for such use through Elsevier's permissions site at: http://www.elsevier.com/ locate/permissionusematerial NIH Public Access After correcting for sequencing error, we sampled 26,072 SAGE tags for the control library and 26,815 tags for the SNP-exposed library. We used log-likelihood statistics (R > 1.5) to reduce the effects of sampling...

Abstract.-Of all Pacific salmonids, Chinook salmon Oncorhynchus tshawytscha display the greatest variability in return times to freshwater. The molecular mechanisms of these differential return times have not been well described. Current methods, such as long serial analysis of gene expression (LongSAGE) and microarrays, allow gene expression to be analyzed for thousands of genes simultaneously. To investigate whether differential gene expression is observed between falland spring-run Chinook salmon from California's Central Valley, LongSAGE libraries were constructed. Three libraries containing between 25,512 and 29,372 sequenced tags (21 base pairs/tag) were generated using messenger RNA from the brains of adult Chinook salmon returning in fall and spring and from one oceancaught Chinook salmon. Tags were annotated to genes using complementary DNA libraries from Atlantic salmon Salmo salar and rainbow trout O. mykiss. Differentially expressed genes, as estimated by differences in the number of sequence tags, were found in all pairwise comparisons of libraries (freshwater versus saltwater¼ 40 genes; fall versus spring¼ 11 genes; and spawning versus nonspawning ¼ 51 genes). The gene for ependymin, an extracellular glycoprotein involved in behavioral plasticity in fish, exhibited the most differential expression among the three groupings. Reverse transcription polymerase chain reaction analysis verified the differential expression of ependymin between the fall-and spring-run samples. These LongSAGE libraries, the first reported for Chinook salmon, provide a window of the transcriptional changes during Chinook salmon return migration to freshwater and spawning and increase the amount of expressed sequence data.