传统测序与高通量测序在稚鱼食性分析中的比较

对鱼类早期生长阶段的摄食研究有助于了解其饵料来源及其在食物网中的功能地位,而全面准确地获取其食物种类信息是关键,高通量测序技术的发展给动物食性研究带来了前所未有的机遇和挑战.本研究以大亚湾人工码头海域金钱鱼稚鱼为对象,以18S rDNA为靶标,分别使用传统Sanger测序和Illumina Solexa高通量测序对其食物组成进行分析,比较两种方法在稚鱼摄食研究中的适用性.结果表明: 金钱鱼稚鱼为杂食性,食物多样性高,纤毛虫和苔藓动物是最优势的食物类群.使用传统测序方法共获得67条有效食物序列,分属于8个类群,涵盖23个生物种类;使用高通量测序方法共获得17000多条有效食物序列,分属于9个类群,涵盖35个生物种类.两种方法检测到的食物类群基本相同,但高通量测序方法在反映食物多样性和覆盖范围上更具优势,且灵敏度更高,检测出传统测序方法未发现的甲藻和褐藻种类,说明高通量测序技术可以较全面而准确地覆盖稚鱼的食物谱.高通量测序获取的大量数据,可在一定程度上提供半定量信息,克服传统测序在定量研究方面的不足.高通量测序技术在稚鱼摄食研究上优势更明显,食谱覆盖更广,检测灵敏度更高,显著提升了数据与结果的可信度,可为海洋生物摄食生态学研究提供强有力的支撑.

Comparison between traditional sequencing and high-throughput sequencing on the dietary analysis of juvenile fish.

Feeding analysis of fish at early growth stages is helpful to understand its food source and functional role. Comprehensively and accurately obtaining the information of food species is critical for this endeavor. The development of high-throughput sequencing (HTS) technology has brought unprecedented opportunities and challenges to dietary analysis of animals. Molecular techniques have been widely used for analyzing diet composition of different animals, among which HTS has been successfully used on marine fish. But the accuracy and suitability of HTS have not been confirmed and compared with traditional Sanger sequencing on the dietary analysis of juvenile fish yet. In this study, a comprehensive comparison was performed between traditional Sanger sequencing and HTS on the diet analysis of Scatophagus argus juveniles, an important commercial fish in South and Southeast Asia. The juvenile fish were collected near a wharf in Daya Bay, China, and analyzed both by traditional Sanger sequencing and HTS methods targeting 18S rDNA gene. Results showed that the diet composition was complicated and dominated by ciliates and bryozoans, indicating an omnivorous feeding of juveniles. Sixty-seven effective sequences were obtained and 21 food species were identified by Sanger sequencing, while 35 food species from 17000 effective sequencings were identified by HTS. The food groups obtained by both techniques were similar. HTS had advantages in dietary coverage due to the large amount of data. HTS detected preys of dinoflagellates and brown algae which were not found in Sanger sequencing. In addition, HTS could provide semi-quantitative information to some extent based on a large amount of data, overcoming the deficiency of Sanger sequencing in quantitative research. Our results suggested that HTS is much more powerful than Sanger sequencing on diet analysis of juvenile fish, which can provide more details and comprehensive information about the feeding of juvenile fish.