Department of Biological Sciences, Fordham University, Bronx, New York 10458.
Division of Invertebrate Zoology, American Museum of Natural History, Central Park West at 79th Street, New York, New York 10024.
J Parasitol. 2020 Nov 12;106(6):843-853. doi: 10.1645/19-189.
Leech-derived invertebrate DNA (iDNA) has been successfully leveraged to conduct surveys of vertebrate host biodiversity across the Indo Pacific. However, this technique has been limited methodologically, typically only targeting mammalian 16S rDNA, or both 16S and vertebrate 12S rDNA for leech host determination. To improve the taxonomic richness of vertebrate host species in iDNA surveys, we re-analyze datasets from Bangladesh, Cambodia, China, and Madagascar through metabarcoding via next generation sequencing (NGS) of 12S, 16S (2 types, one designed to target mammals and the other, residual eDNA), nicotinamide adenine dinucleotide hydride dehydrogenase 2 (ND2), and cytochrome c oxidase subunit 1 (COI). With our 5 primer sets, we identify 41 unique vertebrate hosts to the species level, among 1,200 leeches analyzed, along with an additional 13 taxa to the family rank. Within our 41 taxa, we note that adding ND2 and COI loci increased species richness detection by 25%. NGS has emerged as more efficient than Sanger sequencing for large scale metabarcoding applications and, with the decline in cost of NGS, our pooled sample multilocus protocol is an attractive option for iDNA biodiversity surveys.
已成功利用水蛭衍生的无脊椎动物 DNA（iDNA）对印度洋太平洋地区脊椎动物宿主生物多样性进行调查。然而，该技术在方法上受到限制，通常仅针对哺乳动物 16S rDNA，或同时针对 16S 和脊椎动物 12S rDNA 进行水蛭宿主鉴定。为了提高 iDNA 调查中脊椎动物宿主物种的分类丰富度，我们通过下一代测序（NGS）的 12S、16S（2 种类型，一种设计用于靶向哺乳动物，另一种是残留的 eDNA）、烟酰胺腺嘌呤二核苷酸脱氢酶 2（ND2）和细胞色素 c 氧化酶亚基 1（COI）对来自孟加拉国、柬埔寨、中国和马达加斯加的数据集进行了重新分析。通过我们的 5 个引物对，我们在分析的 1200 只水蛭中鉴定出 41 种独特的脊椎动物到种水平，另外还有 13 种到科水平。在我们的 41 个分类群中，我们注意到添加 ND2 和 COI 基因座可将物种丰富度检测提高 25%。NGS 已成为比 Sanger 测序更有效的大规模代谢组学应用技术，随着 NGS 成本的降低，我们的组合样本多基因座方案是 iDNA 生物多样性调查的一个有吸引力的选择。
