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Alternaria alternata causes leaf blight of rosy periwinkle (Catharanthus roseus) in Iraq

  • Adnan A. LahufEmail author
Article

Abstract

Nursery-grown rosy periwinkles (Catharanthus roseus) in Kerbala province, Iraq, showed severe symptoms of leaf blight. The causal agent associated with the diseased plants was identified as the fungus Alternaria alternata. This identification was based on the pathogen’s morphological, molecular and pathogenic characteristics. To the best of the author’s knowledge, leaf blight of rosy periwinkle caused by A. alternata is the first record in Iraq.

Keywords

Alternaria alternata Leaf blight Catharanthus roseus rDNA-ITS Iraq 

Catharanthus roseus, known as rosy periwinkle or Madagascar periwinkle, is an annual or perennial evergreen herbal plant that is native from Madagascar (Stearn 1975). Nowadays, it is cultivated globally, including in Iraq, for its therapeutic and ornamental attractive features. It has an ornamental aesthetic appearance related to its green variegation and prolific flowers in splendid colors (Nejat et al. 2015). The plant is also a source of pharmaceutical compounds such as terpenoid indole alkaloids including vincristine and vinblastine that have valuable antitumor properties (Aslam et al. 2010).

Several phytopathogens have been reported to cause a variety of diseases on periwinkle plants worldwide. The majority of the fungal diseases recorded were root rot and damping-off incited by numerous fungi such as Rhizoctonia solani, Fusarium oxysporum, F. solani, Thielaviopsis sp., Sclerotinia sclerotiorum, Sclerotium rolfsii and Pythium aphanidermatum and foliar blight caused by Phytophthora parasitica, P. tropicalis, Colletotrichum dematium and Botrytis cinerea (Nejat et al. 2015; Ou-Yang and Wu 1998). However, there are no published studies relating to pathogens infecting periwinkle plants in Iraq.

During the growing season 2015/ 2016, severe symptoms of foliar blight were observed on rosy periwinkle in one of the main ornamental nurseries in Kerbala province, Iraq. The symptoms initiated as small circular to irregular light brown spots that gradually enlarged and darkened to cover most of the leaf surface. The diseased leaves ultimately became withered and dry leading to the death of the plant.

Representative symptomatic leaves of rosy periwinkle plants were collected, dissected, surface sterilised (2% sodium hypochlorite solution) and placed on water agar (WA). After two days of incubation at 25 °C in darkness, a hyphal tip of each emerging colony was sub-cultured on potato dextrose agar (PDA). Nine single-hyphal-tipped fungal isolates were recovered from the diseased leaf samples. Morphological examinations of these isolates showed that the colonies grown on PDA were white in the beginning, converting to brownish olivaceous in the later stage of growing (Fig. 1a). The reverse surface of the colony was dark brown to black. Conidiophores were usually short being either ramified or simple, mostly curved measuring 13–15 μm long and 3–6 μm wide. Conidia (Fig. 1b) produced usually in long branched strings in diverse shapes; oval, obclavate, obpyriform or ellipsoidal with average size 20–30 μm long and 15–18 μm wide. They possessed two types of septa; transverse (1–5) and longitudinal (0–2), with a short apical beak shaped either conical or cylindrical and a prominent basal hole. The morphological features of all nine isolates were similar and agreed with those of Alternaria alternata described by Simmons (2007). One representative isolate of the fungus A. alternata was deposited in the microorganisms culture collection of phytopathogens and biological agents, University of Kerbala, Kerbala, Iraq with the accession No. IRAQAA 00025P.
Fig. 1

Morphological characteristics of Alternaria alternata (a) pure colony (b) diverse shapes and sizes of conidia. Scale bar =20 μm

To confirm this identification, one of the representative isolates was selected for amplifying and sequencing of the ribosomal DNA internal transcribed spacer region (rDNA- ITS) utilising the ITS1 and ITS4 primers (White et al. 1990). The sequence obtained (GenBank Accession No. MF099863) was highly similar to those of A. alternata isolates recorded in GenBank database. Accordingly, the phylogenetic analysis using MEGA7 with neighbour-joining method showed that it respectively assembled with numerous reference of A. alternata strains (Fig. 2).
Fig. 2

Phylogenetic analysis of the fungus isolated from the diseased rosy periwinkle plant (MF099863.1; indicated with a black dot) with different strains of A. alternata depending on the alignment of the nucleotide sequences using MEGA7.0 applying neighbour-joining tree process. The out-group of this tree was Fusarium solani strain AY097318.1

Pathogenicity of all nine A. alternata isolates were assessed using the detached leaf assay (Akhtar et al. 2011). Healthy looking leaves of rosy periwinkle were surface sterilised using 70% ethanol and placed into transparent plastic boxes on moist tissues. A drop of conidial suspension (50-μl of 2000 conidia/ml concentration), harvested from 7 day old pure cultures of each isolate, were placed on the sterilised leaves while others were left without inoculation for control. All boxes were closed firmly, incubated at 25 °C in darkness and monitored daily until blight symptoms were seen, typically 10–14 days after inoculation (Fig. 3). However, no symptoms were seen on leaves of the control. To accomplish Koch’s postulates, pieces of diseased leaves collected from the edge of blight lesion and healthy tissue were surface sterilised and placed on PDA as mentioned above. The fungus was re-isolated consistently from all inoculated leaves with morphological characterisations identical to those described above.
Fig. 3

Detached leaf assay with pathogenic A. alternata inoculation, (left) symptoms of leaf blight on rosy periwinkles leaf (right) control leaf

The fungal pathogen isolated from symptomatic leaves of rosy periwinkle was characterised to species level based on its morphological and molecular characteristics, which were similar to those of A. alternata.

Alternaria alternata has been reported as a fungus causing various diseases mainly leaf spot and blight on a wide range of plant hosts. It was, for example, reported as the cause of leaf spot of Musa spp., Actinidia deliciosa, Rumex vesicarius and Juglans regia (Parkunan et al. 2013; Corazza et al. 1999; Sankar et al. 2012; Belisario et al. 1999) and blight of Zinnia acerosa, Artemisia annua, Incarvillea emodi and Zanthoxylum piperitum (Colbaugh et al. 2001; Samanta et al. 2009; Shanmugam et al. 2011; Yang et al. 2013). Additionally, A. alternata was reported as an endophytic fungus on Catharanthus roseus (Kharwar et al. 2008; Momsia and Momsia 2013; Palem et al. 2015). However, there is no previous report relating this pathogen to disease on rosy periwinkle plants in Iraq. Hence, this is the first report of A. alternata causing leaf blight of rosy periwinkle.

Notes

Acknowledgements

The author would like to acknowledge Dr. Jennie Brierley for proofreading and the Plant protection Department/Agriculture College/University of Kerbala for using of the facilities.

References

  1. Akhtar KP, Sarwar N, Saleem MY, Asghar M (2011) Convolvulus arvensis, a new host for Alternaria solani causing early blight of Solanum lycopersicum in Pakistan. Aust Plant Dis Notes 6(1):84–86.  https://doi.org/10.1007/s13314-011-0029-3 CrossRefGoogle Scholar
  2. Aslam J, Khan SH, Siddiqui ZH, Fatima Z, Maqsood M, Bhat MA, Nasim SA, Ilah A, Ahmad IZ, Khan SA, Mujib A, Sharma MP (2010) Catharanthus roseus (L.) G. Don. An important drug: it’s applications and production. Pharmacie Globale (IJCP) 4:1–16Google Scholar
  3. Belisario A, Forti E, Corazza L, van Kesteren HA (1999) First report of Alternaria alternata causing leaf spot on English walnut. Plant Dis 83(7):696.  https://doi.org/10.1094/PDIS.1999.83.7.696A CrossRefGoogle Scholar
  4. Colbaugh PF, Mackay WA, George S (2001) Alternaria alternata flower blight of Zinnia acerosa in Texas. Plant Dis 85(2):228.  https://doi.org/10.1094/PDIS.2001.85.2.228C CrossRefGoogle Scholar
  5. Corazza L, Luongo L, Parisi M (1999) First report of leaf spot caused by Alternaria alternata on kiwifruit in Italy. Plant Dis 83(5):487.  https://doi.org/10.1094/PDIS.1999.83.5.487D CrossRefGoogle Scholar
  6. Kharwar RN, Verma VC, Strobel G, Ezra D (2008) The endophytic fungal complex of Catharanthus roseus (L.) G. Don. Curr Sci 95:228–233Google Scholar
  7. Momsia P, Momsia T (2013) Isolation, frequency distribution and diversity of novel fungal endophytes inhabiting leaves of Catharanthus roseus. Int J Life Sc Bt Pharm Res 2(4):83–87Google Scholar
  8. Nejat N, Valdiani A, Cahill D, TanY MM, Abiri R (2015) Ornamental exterior versus therapeutic interior of Madagascar periwinkle (Catharanthus roseus): the two faces of a versatile herb. Sci World J.  https://doi.org/10.1155/2015/982412
  9. Ou-Yang W, Wu WS (1998) Survey of periwinkle diseases in Taiwan. Plant Pathol Bull 7:147–149Google Scholar
  10. Palem PC, Kuriakose GC, Jayabaskaran C (2015) An endophytic fungus, Talaromyces radicus, isolated from Catharanthus roseus, produces vincristine and vinblastine, which induce apoptotic cell death. PLoS One 10:e0144476CrossRefGoogle Scholar
  11. Parkunan V, Li S, Fonsah EG, Ji P (2013) First report of Alternaria leaf spot of banana caused by Alternaria alternata in the United States. Plant Dis 97(8):1116.  https://doi.org/10.1094/PDIS-01-13-0007-PDN CrossRefPubMedGoogle Scholar
  12. Samanta JN, Solanki BD, Mandal K (2009) First report of sweet wormwood leaf blight disease in India. Aust Plant Dis Notes 4(1):78–79.  https://doi.org/10.1071/DN09033 CrossRefGoogle Scholar
  13. Sankar R, Devamma MN, Giridhar D (2012) First report of Alternaria alternata causing leaf spot on Rumex vesicarius in India. Aust Plant Dis Notes 7(1):17–18.  https://doi.org/10.1007/s13314-011-0036-4 CrossRefGoogle Scholar
  14. Shanmugam V, Dhyani D, Ananthapadmanaban D (2011) First report of Alternaria sp. causing blight on Incarvillea emodi. Aust Plant Dis Notes 6(1):33–35.  https://doi.org/10.1007/s13314-011-0012-z CrossRefGoogle Scholar
  15. Simmons EG (2007) Alternaria: an identification manual. CBS Fungal biodiversity centre, UtrechtGoogle Scholar
  16. Stearn WT (1975) A synopsis of the genus Catharanthus (Apocynaceae). In: Taylor WL, Fransworth NR (eds) Catharanthus alkaloids. Marcel Dekker, New York, pp 9–44Google Scholar
  17. White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Gelfand MA, Sninsky DH, White TJ (eds) PCR Protocols. a guide to methods and applications. Academic Press, San Diego, pp 315–322Google Scholar
  18. Yang WX, Liu F, Zhang N, Ren XD, Liu DQ (2013) First report of Alternaria alternata causing blight on Zanthoxylum piperitum in China. Plant Dis 97(6):840.  https://doi.org/10.1094/PDIS-10-12-0928-PDN CrossRefPubMedGoogle Scholar

Copyright information

© Australasian Plant Pathology Society Inc. 2019

Authors and Affiliations

  1. 1.Plant protection Department, Agriculture CollegeUniversity of KerbalaKerbalaIraq

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