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Identification of two cryptic species in the Anopheles (Cellia) annularis complex using ribosomal DNA PCR-RFLP

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Abstract

Anopheles (Cellia) annularis Van der Wulp is a complex of two sibling species provisionally designated as species A and B and can only be differentiated on the basis of the paracentric inversion in the ovarian polytene chromosomes. To analyze the distribution of these two species and to develop a molecular method for the identification of these two cryptic species, we sequenced the ribosomal DNA internal transcribed spacer 2 (ITS2) and domain 3 (D3) of A. annularis specimens collected from Sonapur (Assam), Jabalpur (Madhya Pradesh), Ranchi (Jharkhand), and Ghaziabad (Uttar Pradesh). We did not find any sequence variation among the specimens collected from Assam, Madhya Pradesh, and Jharkhand states, whereas two types of sequences were obtained from the specimens collected from the state of Uttar Pradesh, which correspond to species A and B of the A. annularis complex. Species A was more prevalent among the all four regions studied. The ITS2 sequence of species A showed unique restriction sites for MvaI and Eco24I, while species B displayed HinfI and NruI sites. Similarly, the D3 sequence of species A showed unique restriction site for Alw26I, while species B showed a unique KpnI site. In this study, we report for the first time the development of ribosomal DNA polymerase chain reaction-restriction fragment length polymorphism methods for identifying these two cryptic species of the Annularis complex.

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Acknowledgments

The authors would like to thank Prof. A. P. Dash [Director, National Institute of Malaria Research (ICMR), Delhi] for his support and Ms. Sumiti Vinayak for the discussion and for proofreading this manuscript. MTA acknowledges the Council of Scientific and Industrial Research for the Senior Research Fellowship. Financial support for the work granted by the Indian Council of Medical Research and the Department of Biotechnology, Government of India, is acknowledged. We are also grateful for the bioinformatics facilities of the Biotechnology Information System (BTIS). The authors also declare that the experiments performed in this manuscript comply with the current laws of India.

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Authors and Affiliations

  1. Department of Biotechnology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, 110029, India

    Mohammad Tauqeer Alam & Yagya D. Sharma

  2. National Institute of Malaria Research (Field Station), Car Nicobar, Andaman and Nicobar Islands, India

    Manoj K. Das

  3. National Institute of Malaria Research (Field Station), Sonapur, Kamrup, Assam, 782402, India

    Vas Dev

  4. Regional Medical Research Centre for Tribes, Jabalpur, Madhya Pradesh, India

    Musharraf A. Ansari

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  1. Mohammad Tauqeer Alam
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  2. Manoj K. Das
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  3. Vas Dev
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  4. Musharraf A. Ansari
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  5. Yagya D. Sharma
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Correspondence to Yagya D. Sharma.

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Alam, M.T., Das, M.K., Dev, V. et al. Identification of two cryptic species in the Anopheles (Cellia) annularis complex using ribosomal DNA PCR-RFLP. Parasitol Res 100, 943–948 (2007). https://doi.org/10.1007/s00436-006-0375-9

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  • DOI: https://doi.org/10.1007/s00436-006-0375-9

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