BackgroundNon-invasive sampling has opened avenues for the genetic study of elusive species, which has contributed significantly to their conservation. Where field based identity of non-invasive sample is ambiguous (e.g. carnivore scats), it is essential to establish identity of the species through molecular approaches. A cost effective procedure to ascertain species identity is to use species specific primers (SSP) for PCR amplification and subsequent resolution through agarose gel electrophoresis. However, SSPs if ill designed can often cross amplify non-target sympatric species. Herein we report the problem of cross amplification with currently published SSPs, which have been used in several recent scientific articles on tigers (Panthera tigris) and leopards (Panthera pardus) in India. Since these papers form pioneering research on which future work will be based, an early rectification is required so as to not propagate this error further.ResultsWe conclusively show cross amplification of three of the four SSPs, in sympatric non-target species like tiger SSP amplifying leopard and striped hyena (Hyaena hyaena), and leopard SSP amplifying tiger, lion (Panthera leo persica) and clouded leopard (Neofelis nebulosa), with the same product size. We develop and test a non-cross-amplifying leopard specific primer pair within the mitochondrial cytochrome b region. We also standardize a duplex PCR method to screen tiger and leopard samples simultaneously in one PCR reaction to reduce cost and time.ConclusionsThese findings suggest the importance of an often overlooked preliminary protocol of conclusive identification of species from non-invasive samples. The cross amplification of published primers in conspecifics suggests the need to revisit inferences drawn by earlier work.Electronic supplementary materialThe online version of this article (doi:10.1186/s12863-016-0344-y) contains supplementary material, which is available to authorized users.

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Recent genetic analysis has shown that the extinct Caspian Tiger (P. t. virgata) and the living Amur Tigers (P. t. altaica) of the Russian Far East are actually taxonomically synonymous and that Caspian and Amur groups historically formed a single population, only becoming separated within the last 200 years by human agency. A major conservation implication of this finding is that tigers of Amur stock might be reintroduced, not only back into the Koreas and China as is now proposed, but also through vast areas of Central Asia where the Caspian tiger once lived. However, under the current tiger conservation framework the 12 "Caspian Tiger States" are not fully involved in conservation planning. Equal recognition as "Tiger Range States" should be given to the countries where the Caspian tiger once lived and their involvement in tiger conservation planning encouraged. Today, preliminary ecological surveys show that some sparsely populated areas of Central Asia preserve natural habitat suitable for tigers. In depth assessments should be completed in these and other areas of the Caspian range to evaluate the possibility of tiger reintroductions. Because tigers are a charismatic umbrella species, both ecologically and politically, reintroduction to these landscapes would provide an effective conservation framework for the protection of many species in addition to tigers. And for today's Amur Tigers this added range will provide a buffer against further loss of genetic diversity, one which will maintain that diversity in the face of selective pressures that can only be experienced in the wild.

Unfortunately, after publication of this article [1], it was noticed that the LSP Forward primer listed in Additional file 2 was incorrect. The corrected supplementary material can be found here.