Transcriptomic analysis of reduced sensitivity to praziquantel in Schistosoma mansoni. Transcriptomic analysis of reduced sensitivity to praziquantel in Schistosoma mansoni

Schistosomiasis is a chronic neglected tropical disease caused by digenetic parasitic flatworms of the genus Schistosoma. The disease is estimated to affect over 206 million people, the majority of whom live in Africa where Schistosoma mansoni and Schistosoma japonicum are the major causative agents. While a number of drugs have been used to treat schistosomiasis, praziquantel (PZQ) is the only one that is widely available, relatively cheap, and easy to use. The reliance on a single drug for the treatment of such a prevalent disease is a cause for concern due to the potential for resistance to render PZQ ineffective. In this study we examine the transcriptome of three generations of a laboratory strain of S. mansoni (PR1) whose susceptibility to PZQ has been diminished across 9 passages through exposure to increasing sub-lethal doses of the drug. Miracidial susceptibility was significantly reduced after exposure to 2 x 50 mg/Kg PZQ during the first passage. Susceptibility of worms in vivo was first assessed during passage 5 when mice infected with PZQ-selected schistosomes were dosed with a lethal dose of 3 x 300 mg/Kg PZQ resulting in only a 10% reduction in worm number compared to control treatment. The emergence of reduced sensitivity was marked by a shift in sex ratio from a predominantly male to female population, a reduction in the length of females and ultimately the loss of the PZQ-selected line after passage 9, perhaps due to a selection induced fitness cost. Analysis of differentially regulated transcripts did not suggest that any particular gene product or pathway was associated with drug resistance suggesting either a loss of function mutation to a single gene or an epistatic interaction of multiple gene products. Overall design: Examination of lab selected PZQ resistant Schistosoma mansoni parasites, in quadruplicate, through 3 passages using Illumina NextSeq 500.

血吸虫病（Schistosomiasis）是由血吸虫属（Schistosoma）双生涯寄生扁形虫引发的慢性被忽视热带病。据估算，该病影响全球超2.06亿人口，其中多数分布于非洲，曼氏血吸虫（Schistosoma mansoni）与日本血吸虫（Schistosoma japonicum）是当地主要致病病原体。
目前虽有多种药物可用于治疗血吸虫病，但吡喹酮（praziquantel，PZQ）是唯一广泛可及、价格相对低廉且使用便捷的治疗药物。针对如此高发的疾病仅依赖单一药物治疗存在潜在隐患：耐药性的出现可能使吡喹酮丧失药效。
本研究对曼氏血吸虫实验室株PR1的三代转录组（transcriptome）展开分析：该菌株经9次连续暴露于递增亚致死剂量吡喹酮的传代培养后，对吡喹酮的易感性已显著降低。首次传代阶段暴露于2×50mg/kg吡喹酮后，毛蚴的药物易感性即出现显著下降。体内虫体的药物易感性首次在第5代传代时进行评估：感染经吡喹酮筛选的血吸虫的小鼠被给予致死剂量3×300mg/kg吡喹酮后，虫体数量仅较对照组降低10%。敏感性降低的特征包括种群性别比例从以雄性为主转向以雌性为主、雌性虫体长度缩短，最终该吡喹酮筛选株在第9代传代后丢失，这一现象可能源于筛选诱导的适合度代价。
对差异调控转录本的分析未发现特定基因产物或通路与耐药性相关，提示耐药性的产生可能源于单基因功能丧失突变，或是多个基因产物的上位互作（epistatic interaction）。
总体实验设计：采用Illumina NextSeq 500测序平台，对经吡喹酮筛选的曼氏血吸虫耐药株的3个传代样本进行四重复转录组测序分析。