The Species Dilemma of Northeast Indian Mahseer (Actinopterygii: Cyprinidae): DNA Barcoding in Clarifying the Riddle

BackgroundThe taxonomic validity of Northeast Indian endemic Mahseer species, Tor progeneius and Neolissochilus hexastichus, has been argued repeatedly. This is mainly due to disagreements in recognizing the species based on morphological characters. Consequently, both the species have been concealed for many decades. DNA barcoding has become a promising and an independent technique for accurate species level identification. Therefore, utilization of such technique in association with the traditional morphotaxonomic description can resolve the species dilemma of this important group of sport fishes. Methodology/Principal FindingsAltogether, 28 mahseer specimens including paratypes were studied from different locations in Northeast India, and 24 morphometric characters were measured invariably. The Principal Component Analysis with morphometric data revealed five distinct groups of sample that were taxonomically categorized into 4 species, viz., Tor putitora, T. progeneius, Neolissochilus hexagonolepis and N. hexastichus. Analysis with a dataset of 76 DNA barcode sequences of different mahseer species exhibited that the queries of T. putitora and N. hexagonolepis clustered cohesively with the respective conspecific database sequences maintaining 0.8% maximum K2P divergence. The closest congeneric divergence was 3 times higher than the mean conspecific divergence and was considered as barcode gap. The maximum divergence among the samples of T. progeneius and T. putitora was 0.8% that was much below the barcode gap, indicating them being synonymous. The query sequences of N. hexastichus invariably formed a discrete and a congeneric clade with the database sequences and maintained the interspecific divergence that supported its distinct species status. Notably, N. hexastichus was encountered in a single site and seemed to be under threat. ConclusionThis study substantiated the identification of N. hexastichus to be a true species, and tentatively regarded T. progeneius to be a synonym of T. putitora. It would guide the conservationists to initiate priority conservation of N. hexastichus and T. putitora.

研究背景：印度东北部特有玛希尔鱼（Mahseer）物种Tor progeneius与Neolissochilus hexastichus的分类学有效性长期存在争议。该争议主要源于基于形态特征开展物种界定的分歧，导致二者的分类地位曾被混淆长达数十年。DNA条形码（DNA barcoding）已成为一种可靠且独立的精准物种鉴定技术，将该技术与传统形态分类学描述相结合，可解决这一重要游钓鱼类类群的物种界定难题。
研究方法与主要结果：本研究共采集印度东北部不同产区的28尾玛希尔鱼标本（包含副模标本），并统一测定了24项形态测量性状。基于形态测量数据的主成分分析（Principal Component Analysis, PCA）将样本划分为5个显著分化的类群，经分类学归类后对应4个物种，分别为：Tor putitora、T. progeneius、Neolissochilus hexagonolepis与N. hexastichus。通过对涵盖76条不同玛希尔鱼物种DNA条形码序列的数据集进行分析，结果显示：T. putitora与N. hexagonolepis的查询序列与各自的同种参考数据库序列紧密聚类，最大Kimura双参数（K2P）遗传距离为0.8%。同属内最大遗传距离为种内平均遗传距离的3倍，该差值被视为条形码间隙（barcode gap）。T. progeneius与T. putitora样本间的最大遗传距离为0.8%，远低于条形码间隙阈值，表明二者为同物异名。N. hexastichus的查询序列始终与参考数据库中的同源序列形成独立单系支，其种间遗传距离符合其作为独立有效物种的分类地位。值得注意的是，N. hexastichus仅在单个采样点被发现，其种群似乎正面临生存威胁。
研究结论：本研究证实了N. hexastichus为独立有效物种，并初步认定T. progeneius为T. putitora的同物异名。该研究结果可为保护工作者优先开展N. hexastichus与T. putitora的物种保护提供科学指导。