To elucidate the mechanisms of antischistosoma resistance, drug-resistant Schistosoma mansoni laboratory isolates are essential. We developed a new method for inducing resistance to praziquantel (PZQ) The current strategy for schistosomiasis control is based on large-scale treatments of populations aimed at reducing disease morbidity (WHO 2002). Currently, praziquantel (PZQ) is the drug of choice (Utzinger & Keiser 2004, Fenwick & Webster 2006, with the main advantages of its use being oral administration, single dose, low toxicity and low cost (Fenwick et al. 2003, Utzinger & Keiser 2004. Despite the advantages of PZQ, there is concern about the development of Schistosoma mansoni resistance to PZQ, both under laboratory and field conditions (Abdul-Ghani et al. 2009). In the laboratory, induction of resistance is based on the treatment of mice infected with S. mansoni, initially using sub-curative doses of PZQ. Afterwards, the dosage is increased for at least seven passages in mice/snails to complete the life cycle of the parasite (Ismail et al. 1994, Fallon et al. 1995.The complete mechanism of action of PZQ is still unclear (Doenhoff et al. 2008). Obtaining resistant strains is important for the evaluation of such mechanisms as well as for the development of alternative drugs for schistosomiasis treatment and control. Studies show that PZQ is effective not only in adult worms, but also in the intramolluscan phase of the parasite (Coelho et al. 1988. We report a novel meth- od for the induction (or selection) of S. mansoni worms resistant to PZQ using successive treatments of infected Biomphalaria glabrata snails. SUBJECTS, MATERIALS AND METHODSParasites and hosts -The S. mansoni (LE strain) life cycle was maintained using B. glabrata (Barreiro de Cima strain) snails as intermediate hosts and Swiss mice as definitive hosts, according to Pellegrino and Katz (1968) and Souza et al. (1995).Perfusion of adult worms from infected mice -Two methods were used. The methodology described by Pellegrino and Siqueira (1956) used a needle attached to a Brewer's automatic pipetter to inject saline solution under pressure into the descendent aorta. Afterwards, saline was injected into the hepatic hilum of mice after sectioning the portal vein, allowing the perfusion of the portal system and mesenteric veins. Worms were recovered and counted. To recover the worms using the methodology described by Smithers and Terry (1965) the portal vein of the mice was sectioned and the culture medium was gently injected into the base of the left ventricle of the infected mice's hearts. It is not possible to recover all the worms using this methodology with a lower pressure injection, but the integrity of the parasite's tegument is preserved. Therefore, this methodology is ideal for the recovery of worms when the goal is to cultivate or evaluate other parameters such as tegumental integrity and/or excretory activity.Induction of resistance to PZQ in the intramolluscan phase -Two-hundred B. glabrata were individually infected with 10 S. man...

Long noncoding RNAs (lncRNAs) are transcripts generally longer than 200 nucleotides with no or poor protein coding potential, and most of their functions are also poorly characterized. Recently, an increasing number of studies have shown that lncRNAs can be involved in various critical biological processes such as organism development or cancer progression. Little, however, is known about their effects in helminths parasites, such as Schistosoma mansoni. Here, we present a computational pipeline to identify and characterize lncRNAs from RNA-seq data with high confidence from S. mansoni adult worms. Through the utilization of different criteria such as genome localization, exon number, gene length, and stability, we identified 170 new putative lncRNAs. All novel S. mansoni lncRNAs have no conserved synteny including human and mouse. These closest protein coding genes were enriched in 10 significant Gene Ontology terms related to metabolism, transport, and biosynthesis. Fifteen putative lncRNAs showed differential expression, and three displayed sex-specific differential expressions in praziquantel sensitive and resistant adult worm couples. Together, our method can predict a set of novel lncRNAs from the RNA-seq data. Some lncRNAs are shown to be differentially expressed suggesting that those novel lncRNAs can be given high priority in further functional studies focused on praziquantel resistance.

Introduction: Schistosomiasis, caused by Schistosoma mansoni, is a public health concern in Brazil. However, the most popular diagnostic method, the Kato-Katz technique, exhibits low sensitivity in low-endemicity areas. We aimed to compare the performance of an immunological assay, the point-of-care circulating cathodic antigen (POC-CCA®) test, with that of two parasitological techniques in a low-endemicity population. Methods: Our study included 141 individuals living in Estreito de Miralta, Minas Gerais, Brazil. Fecal samples were obtained from all participants and analyzed for schistosomiasis using two parasitological techniques: the Kato-Katz technique and the saline gradient technique. Additionally, POC-CCA® strips were utilized for testing urine samples. The results obtained by the different techniques were compared. Results: Analysis of two or 24 slides using the Kato-Katz technique resulted in a positivity rate of 10.6% (15/141) or 19.1% (27/141), respectively. The saline gradient technique yielded a positivity rate of 17.0% (24/141). The prevalence according to both parasitological techniques was 24.1% (34/141). The POC-CCA® test yielded a positivity rate of 22.7% (32/141); however, the positivity rate was merely 2.1% if trace results were considered negative. The agreements observed between POC-CCA® and the parasitological techniques were good (Kappa indexes > 0.64). The POC-CCA® test was more sensitive than the two-slide Kato-Katz technique (p < 0.05) in detecting cases of S. mansoni infection when trace results were considered positive. Conclusions: These findings reinforce the importance of using multiple diagnostic techniques in low-endemicity areas for effective control of disease.

Current schistosomiasis control strategies are largely based on chemotherapeutic agents and a limited number of drugs are available today. Praziquantel (PZQ) is the only drug currently used in schistosomiasis control programs. Unfortunately, this drug shows poor efficacy in patients during the earliest infection phases. The effects of PZQ appear to operate on the voltage-operated Ca2+channels, which are located on the external Schistosoma mansoni membrane. Because some Ca2+channels have dihydropyridine drug class (a class that includes nifedipine) sensitivity, an in vitro analysis using a calcium channel antagonist (clinically used for cardiovascular hypertension) was performed to determine the antischistosomal effects of nifedipine on schistosomula and adult worm cultures. Nifedipine demonstrated antischistosomal activity against schistosomula and significantly reduced viability at all of the concentrations used alone or in combination with PZQ. In contrast, PZQ did not show significant efficacy when used alone. Adult worms were also affected by nifedipine after a 24 h incubation and exhibited impaired motility, several lesions on the tegument and intense contractility. These data support the idea of Ca2+channels subunits as drug targets and favour alternative therapeutic schemes when drug resistance has been reported. In this paper, strong arguments encouraging drug research are presented, with a focus on exploring schistosomal Ca2+channels.

The use of chemotherapy on a mass scale in endemic areas may lead to the appearance of resistant isolates through the mechanism of selective drug pressure. Studies have demonstrated that praziquantel (PZQ) is able to inhibit the excretory activity and to cause tegumental damage in Schistosoma mansoni adult worms. The use of the probe resorufin to evaluate excretory activity, as well as the probe Hoechst 33258 to detect tegumental damage in adult worms, may represent a method to identify resistant (or less susceptible) isolates. The purpose of the present work was to compare the changes caused by PZQ in the function of the excretory system and in the integrity of the tegument of adult worms from the LE isolate (susceptible to PZQ) and the LE-PZQ isolate (less susceptible to PZQ). Worms from the isolate LE-PZQ showed less severe tegumental lesions, in both in vitro and in vivo experiments, detected by labelling with Hoechst 33258 and continued to have a functional excretory system as shown by labelling with resorufin in vitro.