Improvement of the specific detection of <i>Xanthomonas phaseoli</i> pv. <i>manihotis</i> based on the <i>pthB</i> gene
Resumo
Modifications were made in the PCR conditions aiming to overcome the problem of non-amplification of the Xanthomonas phaseoli pv. manihotis (Xpm) fragment, using the primer pair XV / XK described in the literature. The objective of this study was to propose changes in the primers already described (XV / XK_MOD) and validate the use of these new primers in identifying Xpm. The validation procedure was carried out with the primer pair XV and XK_MOD, using different strains of Xpm, other plant pathogenic and endophytic bacteria genera and one isolate of X. phaseoli pv. passiflorae. As a control, additional reactions were conducted in multiplex with the universal primers for the 16S rRNA gene of the bacteria together with XV / XK and XV / XK_MOD. Using the forward primer (XV) described in the literature together with the modified reverse primer (XK_MOD), it was possible to achieve amplification from DNA extracted from in vitro cultures and from infected tissue, but no amplification was noticed for the primer pair described in the literature, confirming the effectiveness of the proposed modification.
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Referências
Adikini, S., Tripathi, L., Beed, F., Tusiime, G., Magembe, E. M., & Kim, D. J. (2011). Development of a specific molecular tool for detecting Xanthomonas campestris pv. musacearum. Plant Pathology, 60(3), 443-452. DOI: 10.1111/j.1365-3059.2010.02419.x
Adriko, J., Aritua, V., Mortensen, C. N., Tushemereirwe, W. K., Kubiriba, J. M., & Lund, O. S. (2012). Multiplex PCR for specific and robust detection of Xanthomonas campestris pv. musacearum in pure culture and infected plant material. Plant Pathology, 61(3), 489-497. DOI: 10.1111/j.1365-3059.2011.02534.x
Al-Saadi, A., Reddy, J. D., Duan, Y. P., Brunings, A. M., Yuan, Q., & Gabriel, S. W. (2007). All five host-range variants of Xanthomonas citri carry one pthA homolog with 17.5 repeats that determines pathogenicity on citrus, but none determine host-range variation. Molecular Plant-Microbe Interactions, 20(8), 934-943. DOI: 10.1094/MPMI-20-8-0934
Arrieta-Ortiz, M. L., Rodriguez, L. M., Perez-Quintero, A. L., Poulin, L., Diaz, A. C., Rojas, N. A., Bernal, A. (2013). Genomic survey of pathogenicity determinants and VNTR markers in the cassava bacterial pathogen Xanthomonas axonopodis pv. manihotis strain CIO151. PLoS ONE, 8(11), 1-18. DOI: 10.1371/journal.pone.0079704
Berg, T., Tesoriero, L., & Hailstones, D. L. (2005). PCR-based detection of Xanthomonas campestris pathovars in Brassica seed. Plant Pathology, 54(3), 416-427. DOI: 10.1111/j.1365-3059.2005.01186.x
Constantin E. C., Cleenwerck I., Maes B. S., Van Malderghem C., De Vos P., Cottyn B. (2016) Genetic characterization of strains named as Xanthomonas axonopodis pv. Dieffenbachiae leads to a taxonomic revision of the X. axonopodis species complex. Plant Pathology 65(5), 792–806. https://doi.org/10.1111/ppa.12461
Doyle, J. J., & Doyle, J. L. (1987). A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin, 19(1), 11-15.
Gonzalez, C., Restrepo, S., Tohme, J., & Verdier, V. (2002). Characterization of pathogenic and nonpathogenic strains of Xanthomonas axonopodis pv. manihotis by PCR-based DNA fingerprinting techniques. FEMS Microbiology Letters, 215(1), 3-31. DOI: 10.1111/j.1574-6968.2002.tb11365.x
López, C. E., & Bernal, A. J. (2012). Cassava Bacterial Blight: Using Genomics for the Elucidation and Management of an Old Problem. Tropical Plant Biology, 5(1), 117-126. DOI: 10.1007/s12042-011-9092-3
Mansfield, J., Genin, S., Magori, S., Citovsky, V., Sriariyanum, M., Ronald, P., & Foster, G. S. (2012). Top 10 plant pathogenic bacteria in molecular plant pathology. Molecular Plant Pathology, 13(6), 614-629. DOI: 10.1111/j.1364-3703.2012.00804.x
Miller, S. A., Beed, F. D., & Harmon, C. L. (2009). Plant disease diagnostic capabilities and networks. Annual Review of Phytopathology, 47, 15-38. DOI: 10.1146/annurev-phyto-080508-081743
Munoz-Bodnar, A., Perez-Quintero, A. L., Gomez-Cano, F., Gil, J., Michelmore, R., Bernal, A., Szurek, B., & Lopez, C. (2014). RNAseq analysis of cassava reveals similar plant responses upon infection with pathogenic and non-pathogenic strains of Xanthomonas axonopodis pv. manihotis. Plant Cell Reports, 33(11), 1901-1912. DOI: 10.1007/s00299-014-1667-7
Ogunjobi, A. A., Fagade, O. E., & Dixon, A. G. O. (2010). Comparative analysis of genetic variation among Xanthomonas axonopodis pv. manihotis isolated from the western states of Nigeria using RAPD and AFLP. Indian Journal of Microbiology, 50(2), 132-138. DOI: 10.1007/s12088-010-0037-6
Parkes, E. Y., Fregene, M., Dixon, A., Boakye-Peprah, B., & Labuschange, M. T. (2013). Combining ability of cassava genotypes for cassava mosaic disease and cassava bacterial blight, yield and its related components in two ecological zones in Ghana. Euphytica, 194(1), 13-24. DOI: 10.1007/s10681-013-0936-9
Robene-Soustrade, I., Legrand, D., Gagnvein, L., Chiroleu, F. M., Laurent, A., & Pruvost, O. (2010). Multiplex nested PCR for detection of Xanthomonas axonopodis pv. Allii from onion seeds. Applied and Environmental Microbiology, 76(9), 2697–703. DOI: 10.1128/AEM.02697-09
Simenc, J., & Potocnik, U. (2011). Rapid differentiation of bacterial species by high resolution melting curve analysis. Applied Biochemistry and Microbiology, 47(3), 256–263.
Sousa, D. R. T., Santos, C. S. S., Wank, B., Silva Júnior, R. M., Cruz, K. S., Monte, R. L., … Souza, J. V. B. (2015). PCR-RFLP as a useful tool for diagnosis of invasive mycoses in a healthcare facility in the North of Brazil. Electronic Journal of Biotechnology, 18(3), 231-235. DOI: 10.1016/j.ejbt.2015.03.012
Teló, E. P., Machado, A. B. M. P., Schmitt, V. M., & Chesky, M. (2007). Determinação do limite mínimo de detecção da técnica de pcr “semi-nested” para Neisseria meningitidis, Haemophilus influenzae e Streptococcus pneumoniae. Revista Brasileira de Análises Clínicas, 39(3), 197-200.
Trindade, L. C. D., Marques, E., Lopes, D. B., & Ferreira, M. Á. D. S. V. (2007). Development of a molecular method for detection and identification of Xanthomonas campestris pv. viticola. Summa Phytopathologica, 33(1), 16-23. DOI: 10.1590/S0100-54052007000100002
Trujillo, C. A., Ochoa, J. C., Mideros, M. F. Restrepo, S., López C., Bernal A. (2014) A complex population structure of the cassava pathogen Xanthomonas axonopodis pv. manihotis in recent years in the Caribbean Region of Colombia. Microbial Ecology, 68(1), 155-167. https://doi.org/10.1007/s00248-014-0411-8
Turner, S., Pryer, K. M., Miao, V. P., & Palmer, J. D. (1999). Investigating deep phylogenetic relationships among cyanobacteria and plastids by small subunit rRNA sequence analysis. Journal of Eukaryotic Microbiology, 46(6), 327-338. DOI: 10.1111/j.1550-7408.1999.tb04612.x
Verdier, V., Mosquera, G., & Assigbétsé, K. (1998). Detection of the cassava bacterial blight pathogen, Xanthomonas axonopodis pv. manihotis, by polymerase chain reaction. Plant Disease, 82(1), 79-83, 1998.
Wensing, A., Gernold, M., & Geider, K. (2011). Detection of Erwinia species from the apple and pear flora by mass spectroscopy of whole cells and with novel PCR primers. Journal of Applied Microbiology, 112(1), 147-158. DOI: 10.1111/j.1365-2672.2011.05165.x
Wonni, I., Ouedraogo, L., Dao, S., Tekete, C., Koita, O., Taghouti, G. M., … Verdier, V. (2015). First Report of Cassava Bacterial Blight Caused by Xanthomonas axonopodis pv. manihotis in Burkina Faso. Plant Disease, 99(4), 551-551. DOI: 10.1094/PDIS-03-14-0302-PDN
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