New Disease Reports (2018) 37, 17. http://dx.doi.org/10.5197/j.2044-0588.2018.037.

017

First report of Coniella granati causing dieback and fruit rot of

pomegranate in Tunisia
H. Jabnoun-Khiareddine 1*, N. Ibrahim 2, R. Aydi Ben Abdallah1 1, M. Mars 3, Z. Kthiri 4 and M. Daami-Remadi 1

1UR13AGR09- Integrated Horticultural Production in the Tunisian Centre-East, Regional Research Centre on Horticulture and Organic Agriculture,
University of Sousse, Chott-Mariem, Tunisia ; 2 Higher Agronomic Institute of Chott-Mariem, University of Sousse, 4042, Chott-Mariem, Tunisia; 3
UR13AGR05- Agrobiodiversity, Higher Agronomic Institute, ChottMariem, IRESA-University of Sousse, Tunisia; 4 National Agronomic Institute of Tunisia,
1082, Tunis, University of Carthage,

Tunisia
*E-mail: jkhayfa@yahoo.fr
Received: 18 Apr 2018. Published: 25 Apr 2018.
Tunisia is one of the main regions for pomegranate (Punica granatum)
cultivation and production. During the spring of 2016, twig necrosis (Fig. 1)
and dieback associated with marginal leaf browning (Fig. 2) and dry fruit
rots were observed on pomegranate cvs. Gabsi and Kalai, in several
orchards in the region of Sousse, on the east coast of Tunisia.
Surfacedisinfected tissues of diseased twigs, leaves and fruits were plated
onto potato dextrose agar (PDA) medium. After seven to fifteen days of
incubation at 25°C, consistent fungal colonies (15 isolates) with white to
pale green aerial mycelia and concentric rings of black pycnidia were
observed. Pycnidia, 75 - 225 μm in diameter, were globose, membranous
and contained hyaline, one-celled and ellipsoid to fusiform conidia,
averaging 8.75 -25 × 2.5 -7.5 μm in size. These morphological features
matched those described earlier for Coniella granati Sacc. (syn. Pilidiella
granati Sacc.) by Alvarez et al. (2016).
Total genomic DNA of one representative isolate was extracted, amplified
by PCR using the universal ITS1/ITS4 primers, sequenced and the
nucleotide sequence obtained had 99% identity with C. granati isolates in
GenBank (Accession Nos. KX507098 and KX833578). Consequently, the
pathogen was ascribed to C. granati and its ITS sequence was deposited in
GenBank (MG256184).
Pathogenicity tests were done with three representative isolates using
disinfected and detached leaves, fruits and branches of pomegranate cv.
Gabsi. A 6 mm agar plug cut from seven-day-old cultures on PDA was
deposited in the centre of the upper side of each leaf (ten leaves per
isolate). Fruits were wounded with a sterile cork borer (3 mm in depth and
6 mm in diameter), and a 6 mm agar plug was placed in each wound (ten
fruits per isolate). Branch segments (15 cm long and 1 to 1.6 cm in
diameter) were wounded (3 mm in diameter and in depth) in the centre
and a 3 mm mycelium plug was inserted into each wound (12 branches per
isolate) and the inoculated area was wrapped with plastic film. All
inoculated and control (inoculated with pathogen-free agar plugs) leaves,
fruits and branches were placed in moistened plastic boxes and maintained
at 25°C for five, nine and thirty days, respectively. Additionally, attached
shoots of one-year-old potted pomegranate cv. Gabsi plants (ten shoots
per isolate) were inoculated as previously described for the detached
branch
test and were grown for sixty days under greenhouse conditions.
Leaves were highly susceptible to C. granati and completely decayed five
days post-inoculation. On fruits, isolates induced soft rot after nine days
and completely decayed within fifteen days (Figs. 3-4). C. granati isolates
were also found to be pathogenic on detached branches and attached
shoots giving rise to brown necrotic lesions that were 2 to 3.5 cm and
reached 11 to 13 cm, respectively. All controls remained symptomless.
Furthermore, the pathogen was isolated from all inoculated tree parts,
thus fulfilling Koch's postulates.
Coniella granati has previously been reported as a pomegranate pathogen
in many regions (Michailides et al., 2011; Mirabolfathy et al., 2012; Chen
et al., 2014; Mincuzzi et al., 2016). To our knowledge, this is the first report
of this pathogen in Tunisia causing leaf necrosis, fruit rot, branch dieback
and shoot blight on pomegranate.
References
Alvarez LV, Groenewald JZ, Crous PW, 2016. Revising the Schizoparmaceae:
Coniella and its synonyms Pilidiella and Schizoparme.
Studies in Mycology 85, 1-34.
http://dx.doi.org/10.1016/j.simyco.2016.09.001
Chen Y, Shao DD, Zhang AF, Yang X, Zhou M, Xu YL, 2014. First report of a
fruit rot and twig blight on pomegranate (Punica granatum) caused by
Pilidiella granati in Anhui province of China. Plant Disease 98, 695.
http://dx.doi.org/10.1094/PDIS-09-13-1012-PDN
Michailides TJ, Puckett R, Reyes H, Morgan DP, 2011. Fruit decay caused by
Pilidiella granati in California. Phytopathology 100, S83.
Mincuzzi A, Garganese F, Ippolito A, Sanzani SM, 2016. First report of
Pilidiella granati causing postharvest fruit rot on pomegranate in
southern Italy. Journal of Plant Pathology 98, 377.
http://dx.doi.org/10.4454/JPP.98I2.045
Mirabolfathy M, Groenewald JZ, Crous PW, 2012. First report of Pilidiella
granati causing dieback and fruit rot of pomegranate (Punica granatum)
in Iran. Plant Disease 96, 461. http://dx.doi.org/10.1094/PDIS-10-11-0887

Figure 1 Figure 2 Figure 3 Figure 4

To cite this report: Jabnoun-Khiareddine H, Ibrahim N, Aydi Ben Abdallah1 R, Mars M, Kthiri Z, Daami-Remadi M, 2018. First report of Coniella granati causing
dieback and fruit rot of pomegranate in Tunisia. New Disease Reports 37, 17. http://dx.doi.org/10.5197/j.2044-0588.2018.037.017
©2018 The Authors This report was published on-line at www.ndrs.org.uk where high quality versions of the figures can be found.

New Disease Reports is a peer-reviewed on-line journal published by the British Society for Plant Pathology,
for more information visit http://www.ndrs.org.uk/ Page 17

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Laporan pertama Coniella granati yang menyebabkan mati pucuk dan busuk buah
Delima di Tunisia.

Oleh : H. Jabnoun-Khiareddine, N. Ibrahim, R. Aydi Ben Abdallah, M. Mars, Z. Kthiri,

dan M. Daami-Remadi

Tunisia adalah salah satu daerah utama untuk penanaman dan produksi buah delima
(Punica granatum). Selama musim semi 2016, ranting mengalami kematian jaringan
(Gambar. 1) dan mati pucuk yang berhubungan dengan pencoklatan ujung daun (Gambar.
2) dan busuk buah kering diamati pada kultivar delima Gabsi dan Kalai, di beberapa
kebun di wilayah Sousse, di pantai timur Tunisia.
Cara penulis menemukan penyakit Coniella granati
Permukaan jaringan yang didesinfeksi dari ranting-ranting yang sakit, daun-daunan
dan buah-buahan disalut ke dalam media agar kentang dekstrosa (PDA). Setelah tujuh
sampai lima belas hari inkubasi pada 25°C, koloni cendawan yang telah murni (15 isolat)
dengan miselia udara berwarna putih hingga hijau pucat dan cincin bulat dari pycnidia
hitam yang diamati. Pycnidia berdiameter 75 - 225 μm, berbentuk bulat, bermembran dan
mengandung hialin, bersel satu dan bentuk elips dan konidia berbentuk torpedo, ukuran
rata-rata 8,75 -25 × 2,5 -7,5 μm. Cir-ciri morfologis ini cocok dengan yang dijelaskan
sebelumnya untuk Coniella granati Sacc. (syn. Pilidiella granati Sacc.) oleh Alvarez et
al. (2016).
Total genom DNA dari satu isolat yang mewakili diekstraksi, diamplifikasi oleh
PCR menggunakan primer ITS1 / ITS4 universal, diurutkan dan sekuens nukleotida yang
diperoleh memiliki 99% identitas dengan isolat C. granati dalam Bank Gen (Accession
No. KX507098 dan KX833578). Oleh karena itu, patogen dianggap berasal dari C.
granati dan urutan ITS disimpan dalam Bank Gen (MG256184).
Tes patogenisitas dilakukan dengan menggunakan tiga isolat representatif daun,
buah dan cabang delima kultivar Gabsi yang telah didesinfeksi dan dilepas. Potongan agar
6 mm dari kultur berumur tujuh hari pada media PDA disimpan di tengah sisi atas setiap
daun (sepuluh daun per isolat). Buah-buahan dilubangi menggunakan cork borer
(kedalaman 3 mm dan diameter 6 mm), dan agar 6 mm ditempatkan di setiap luka
(sepuluh buah per isolat). Segmen cabang (panjang 15 cm dan diameter 1 hingga 1,6 cm)
dilubangi (diameter dan kedalama 3 mm) di tengah dan miselium 3 mm dimasukkan ke
setiap lubang (12 cabang per isolat) dan area yang diinokulasi dibungkus menggunakan
plastic wrapping. Semua inokulasi dan kontrol (diinokulasi dengan media agar bebas
patogen) daun, buah-buahan dan cabang ditempatkan dalam kotak plastik yang dibasahi
dan dipertahankan pada 25 ° C masing-masing selama lima, sembilan dan tiga puluh hari.
Selain itu, pucuk delima kultivar Gabsi berumur satu tahun yang telah ditanam (sepuluh
tunas per isolat) diinokulasi seperti yang dijelaskan sebelumnya untuk uji cabang terpisah
dan ditanam selama enam puluh hari dalam kondisi rumah kaca.
Hasil
Daun sangat rentan terhadap C. granati dan benar-benar membusuk lima hari
setelah inokulasi. Pada buah-buahan, isolat mulai menginduksi busuk lunak setelah
sembilan hari dan benar-benar membusuk dalam waktu lima belas hari (Gambar 3-4).
Isolat C. granati juga ditemukan bersifat patogenik pada cabang-cabang terpisah dan
pucuk-pucuk yang dilekatkan sehingga menimbulkan lesi nekrotik berwarna coklat yang
masing-masing berukuran 2 hingga 3,5 cm dan mencapai 11 hingga 13 cm. Semua kontrol
tetap tanpa gejala. Selanjutnya, patogen diisolasi dari semua bagian pohon yang
diinokulasi, sehingga memenuhi kaidah postulat Koch.
Coniella granati sebelumnya telah dilaporkan sebagai patogen delima di banyak
daerah (Michailides et al., 2011; Mirabolfathy et al., 2012; Chen et al., 2014; Mincuzzi et
al., 2016). Sepengetahuan penulis, ini adalah laporan pertama dari patogen ini di Tunisia
yang menyebabkan nekrosis daun, pembusukan buah, mati pucuk dan cabang serta hawar
pucuk pada buah delima.