Early detection of Phytophthora infestans in potato using carbohydrate binding module 1 protein (CBD1) based antiserum
Tóm tắt
Phytophthora infestans is a global challenging pathogen among the biotic stresses which affect healthy and sustainable potato production. Desirable and successful management strategies depend on understanding of its biology, disease cycle and more importantly rapid and accurate detection of the pathogen during early stage of infection. Here, we report production of recombinant antibodies against whole cellulose binding domain (CBD-1) protein of P. infestans and a synthetic signal peptide of CBD1 and their effectiveness in detection of P. infestans both in infected potato leaf and tuber. The recombinant whole CBD1 protein and synthetic signal peptide of CBD1 protein was used to immunize the rabbits and blood serum was collected, purified and measured by SDS–PAGE. Among the two antibodies, synthetic peptide based antibody showed high specificity for mycelia, sporangia and zoospores. It also showed strong signal intensity for P. infestans infected leaf and tuber tissues. The data indicated that the produced synthetic peptide antiserum was efficient and accurate in discrimination of negative and positive samples in ELISA. Reaction of antiserum with early stage of infected leaf and tuber tissue confirmed the expression of the CBD protein, which in turn confirmed the presence of the pathogen in the plant tissues. Therefore it will enable to detect the pathogen in the potato seed before planting as well as in the standing crop in the field thus, would be helpful to ensure production of healthy seed potatoes and for effective schedule for management practices.
Tài liệu tham khảo
Abdel-Salam AM, Abdel-Kader HS, El-Saghir SM, Hussein MH (2004) Purification, serology and molecular detection of Egyptian isolates of banana bunchy top babuvirus and faba bean necrotic yellows nanovirus. Arab J Biotech 7:141–155
Abdel-Salam AM, Abdel-Kader HS, El-Saghir SM (2005) Biological, serological and molecular detection of banana streak badnavirus in vegetatively propagated banana plants in Egypt. Egypt J Virol 2:255–268
Appel R, Adler N, Habermeyer J (2001) A method for the artificial inoculation of potato tubers with Phytophthora infestans and polymerase chain reaction assay of latently infected sprouts and stems. J Phytopathol 149:287–292
Erwin DC, Ribeiro OK (1996) Phytophthora disease worldwide. APS, St. Paul
Erwin DC, Bartnicki-Garcia S, Tsao PH (1983) Phytophthora: its biology, taxonomy, ecology and pathology. APS, St. Paul
Fry WE, Birch PRJ, Judelson HS, Grunwald NJ, Danies G, Everts KL, Grevens AJ, Gugino BK, Johnson DA, Johnson SB, McGrath MT, Myers KL, Ristaino JB, Roberts PD, Secor G, Smart CD (2015) Five reasons to consider Phytophthora infestans a re-emerging pathogen. Phytopathology 105:966–981
Gaulin E, Jauneau A, Villalba F, Rickauer M, Esquerre-Tugaye MT (2002) The CBEL glycoprotein of Phytophthora parasitica var-nicotianae is involved in cell wall deposition and adhesion to cellulosic substrates. J Cell Sci 115:4565–4575
Hardham AR (2001) The cell biology behind Phytophthora pathogenicity. Aust Plant Pathol 30:91–98
Hardham AR (2007) Cell biology of plant–oomycete interactions. Cell Microbiol 9:31–39
Hardham AR, Mitchell HJ (1998) Use of molecular cytology to study the structure and biology of phytopathogenic and mycorrhizal fungi. Fungal Genet Biol 24:252–284
Harrison JG, Barker HR, Lowe R, Rees EA (1990) Estimation of amount of Phytophthora infestans mycelium in leaf tissue by enzyme-linked immunosorbent assay. Plant Pathol 39:274–277
Hussain T, Sharma S, Singh BP, Jeevalatha A, Sagar V, Sharma NN, Kaushik SK, Chakrabarti SK, Firoz A (2013) Detection of latent infection of Phytophthora infestans in potato seed tubers. Potato J 40:142–148
Jones RW, Ospina-Giraldo M (2011) Novel cellulose-binding-domain protein in Phytophthora is cell wall localized. PLoS One 6:1–6
Kynerova B, Kratka J, Zemanova A, Podana M (1998) The diagnosis of Phytophthora nicotianae var. nicotianae and Phytophthora infestans by polyclonal antibodies detection of the pathogen in plants. Ochrana Rostin 34:15–19
Picard K, Tirilly Y, Benhamou N (2000) Cytological effects of cellulases in the parasitism of Phytophthora parasitica by Pythium oligandrum. Appl Environ Microbiol 66:4305–4314
Sharma S, Singh BP, Sumit S, Patil VU (2016) Phenotypic and genotypic characterization of Phytophthora infestans population of Himachal Pradesh. Indian Phytopathol 69(4):391–395
Skottrup PM, Nicolaisen M, Justesen AF (2006) Rapid determination of Phytophthora infestans sporangia using a surface plasmon resonance immune sensor. J Microbiol Methods 68:507–515
Trout CL, Ristaino JB, Madritch M, Wangsomboonde T (1997) Rapid detection of Phytophthora infestans in late blight-infected potato and tomato using PCR. Plant Dis 81:1042–1048