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Identification and validation of SSR markers for Xanthomonas axonopodis pv. punicae an incitant of bacterial blight of pomegranate

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Abstract

This study reports genome wide characterization and development of first set of microsatellite markers through in silico analysis of eight sequenced Xanthomonas axonopodis pv. punicae strains available in the public database. SSR survey resulted in identification of ~ 4638 perfect SSRs, with mean marker frequency 901 SSRs/Mb and densitiy of 11,006 bp/Mb aross the eight genomes. Frequency distribution graphs revealed hexa-nucleotide repeats were more prominent fowllowed by tri-, tetra-, di- and penta-nucleotides in the analysed genomes. We desinged 2927 SSR primers that are specific to the strain LMG 859 and ePCR confirmed on seven other Xap genomes. This resulted in identification of 542 informative SSRs that are producing single amplicons, from which 66 primers were successfully validated through wet lab experiments on eight Xap isolates of pomegranate. Furthermore, utility of these SSRs were demostrated by analysing molecular diversity among 22 Xap isolates using 20 Xap_SSR primers. SSRs revealed moderate genetic diversity among Xap isolates (61%) and grouped 11 isolates that are repersenting six different states into one cluster. This proved the earlier evidence of wider spread of ST3 type Xap acoss India using Multi locus Sequence Typing (MLST) technique. In summary, Xap_SSR will serve as powerful genomics tools that would helps in monitoring of population dynamics, taxonomy, epidomology and quarantine aspects in bacterial blight pathogen through development of microsatellite based Multilocus Variable number of Tandem repeat analysis (MLVA) in future.

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Data availability

The data which supports our findings were listed in the supplementary files. All the relevant information obtained here will be freely available to any scientist for their research purpose.

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Acknowledgements

The authors are highly grateful to the Indian Council of Agricultural Research (ICAR), New Delhi for extended financial support through ICAR–National Research Centre on Pomegranate, Solapur. Authors are also thankful to Ms. Anitha Alarimar, project assistant working in Plant Pathology Department, ICAR–NRCP, Solapur for extending her help in carrying out this work.

Funding

Funds to conduct current research work were provided by Indian Council of Agricultural Research, New Delhi, India as institutional grants to ICAR–National Research Centre on Pomegranate, Solapur, India.

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Authors and Affiliations

  1. Biotechnology and Plant Pathology, ICAR-National Research Centre on Pomegranate (NRCP), Solapur, 413255, India

    Prakash G. Patil, Jyotsana Sharma, Manjunatha Nanjundappa, N. V. Singh, Shivani M. Jamma, Jeer Vinayaka, Vipul R. Sangnure & R. A. Marathe

  2. State Agriculture Biotechnology, Centre, Centre for Crop & Food Innovation, Murdoch University, Perth, Western Australia

    Abhishek Bohra

  3. Department of Biotechnology and Crop Improvement, University of Horticultural Sciences (UHS), Bagalkot, 587104, India

    Raghavendra Gunnaiah

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  1. Prakash G. Patil
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Patil, P.G., Sharma, J., Nanjundappa, M. et al. Identification and validation of SSR markers for Xanthomonas axonopodis pv. punicae an incitant of bacterial blight of pomegranate. 3 Biotech 12, 153 (2022). https://doi.org/10.1007/s13205-022-03209-z

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