Susceptible Cultivars Research Articles

Bitter rot is an emerging disease of apple (Malus domestica) fruit in Ontario in part due to changing weather conditions. The disease was mostly documented in warm and humid regions such as the southern USA, and Central and South America. Thirteen Ontario orchards in the fall of 2019 and 15 in 2020 were scouted for bitter rot based on their previous history of the disease. 100 fruit were collected from ten asymptomatic trees per cultivar and two susceptible cultivars, 'Empire' and 'Ambrosia' were scouted per orchard. If an orchard did not have either one of these cultivars, 'Honeycrisp' or 'Gala' were used. The fruit was stored at 4-5 oC for five months and then left at 22 oC for two weeks and assessed thereafter for bitter rot symptoms. Monoconidial cultures of Colletrotrichum spp. were established from the symptomatic fruit using potato dextrose agar media with antibiotics at 22 oC 14-hour light cycles. The fungal isolates were divided into two groups based on colony morphology observations seven days after culturing. The first group showed light grey, cottony, aerial mycelium on top and a pink to dark red color on the reverse of the plate. The second group showed light to dark grey, cottony mycelium on top and dark green colonies on the reverse of the plate. It is difficult to identify Colletrotricuhum species solely based on morphology, therefore the representative isolates from each group were used for multilocus gene sequencing and species identification. Genomic DNA was extracted using the Qiagen DNeasy Plant Mini Kit according to the manufacturer's protocols. The ITS region was amplified and sequenced using the primers ITS-1F (Grades & Bruns 1993) and ITS-4 (White et al. 1990). Primers T1 and T2 (O'Donnell & Cigelnik 1997) were used to amplify and sequence the 750 bp region of the TUB gene. Primers GDF1 and GDR1 (Guerber et. al. 2003) were used to amplify a 280 bp region of the GADPH gene. Following an NCBI nucleotide blast search, the isolates from group 1 were identified as C. fioriniae.The ITS sequences of group 2 isolates were matched 100% to the Colletotrichum godetiae type strain CBS133.44 and they were 99% matched to the closest species C. Johnstonii CBS128532. The TUB sequences matched 100% identity to 20 sequences belonging to C. godetiae, 99.59 % identity to C. godetiae type strain CBS133.44, and 97.75% to C. Johnstonii CBS128532. The GADPH sequences matched with 100% identity to C. godetiae ON241087.1 or MT816329.1 and 99-99.5% identity to the type strain CBS133.44 and 98.61-99% identity to CBS128532. Based on the blast results the group 2 isolates were identified as C. godetiae and their sequences were submitted to GenBank with ID OP702962 for ITS and OP972240 and OP972241 for GADPH and OP972242 for the TUB gene. Out of 50 isolates collected in this work, 94% belonged to group 1 and 6% belonged to group 2. Koch's postulates were performed on selected isolates by artificial inoculation of 5 healthy detached fruits of the cultivar, 'Empire.' Fruit surfaces were wiped with 70% ethanol, dried, wounded with a sterile needle, and then inoculated with a 10 µl of spore suspension containing 1x10^6 spores /ml. Inoculated fruits were incubated in a humid chamber at 22°C in dark. Symptoms started to appear at 5 days post-inoculation and looked like small brown circular lesions which developed orange spore masses as they grew into larger lesions. The non-inoculated control fruit did not develop lesions. Fungal colonies were established from the spores collected from the inoculated fruit and found to have identical morphological characteristics to the original isolates. C. godetiae has previously been reported to cause bitter rot in apples from various countries in Europe (Baroncelli et al. 2014; Munda, 2014; Wenneker et al. 2016). C. fioriniae has previously been reported as the dominant species causing bitter rot in apples although other species including C. chrysophilum and C. noveboracense have also been reported as causal agents from the Eastern USA (Khodadadi et al. 2020). To the best of our knowledge, this is the first report of Colletotrichum godetiae causing the bitter rot of apples in Ontario, Canada.

Over the last decade, field assessments of the yellow rust differential lines for resistance genes Yr10 and Yr24 and race analysis in the Middle East have demonstrated efficient yellow rust control by Yr10 and Yr24 (=Yr26). Yellow rust samples collected during 2018-21 in Central West Asia & North and sub-Saharan Africa underwent race analysis at the Regional Cereal Rust Research Center in Izmir, Türkiye. The infected leaf segments were subjected to rehydration at 20°C for three hours. Subsequently, the leaf segments were rubbed on the first leaves of seedlings of susceptible cultivar Morocco. Inoculated seedlings were placed at 10°C in dark conditions with 95% humidity for 24 hrs, then moved to a growth chamber with a 16-hr light (220 µmolm-2s-1) cycle at 15°C and an eight-hour dark period at 12°C. Urediniospores were collected 15 days post-inoculation. A set of yellow rust differential lines including Morocco, Avocet 'S', Avocet 'R', Yr1/6* Avocet 'S', Kalyansona (Yr2), Vilmorin 23 (Yr3), Hybrid 46 (Yr4), Yr6/6* Avocet 'S', Yr7/6* Avocet 'S', Yr8/6* Avocet 'S', Yr9/6* Avocet 'S', Yr10/6* Avocet 'S', Moro (Yr10+), Yr17/6*Avocet 'S', Yr24/6* Avocet 'S', TP1295 (Yr25), Yr27/6* Avocet 'S', YrSp/6* Avocet 'S', Spalding Prolific (YrSP), Strubes Dickkopf (YrSD), Tres/6*Avocet'S', Cham 1, and Ambition was used in race analysis. A mixture of 2 mL Soltrol® and 0.5 mg fresh urediniospores was used to inoculate 10-day-old seedlings of the 23 differential varieties. Pre-inoculation, incubation, and post-inoculation conditions were the same as above. Seedling infection types (ITs) were recorded 15 days post-inoculation on a scale of 0 to 9 (McNeal et al. 1971), where ITs 0 to 6 are classified as low infection types (LITs= avirulent) and ITs 7 to 9 categorized as high infection types (HITs= virulent). HITs of 7 to 9 were observed for the first time on Yr10/6* Avocet 'S', Yr24/6* Avocet 'S', as well as on Moro (Yr10+) for 25 sample of the total 50 isolates from Lebanon and Türkiye in 2018. During the race analysis in 2019 to 2021, virulence for Yr10 and Yr24 was identified among tested samples from Egypt, Lebanon, Jordan, Syria, and Türkiye, indicating the expansion of virulence for Yr10 and Yr24 into new regions. HITs were observed for the durum wheat cultivar Cham 1 and wheat cultivar Ambition in all races. Virulence for YrA, Yr2, Yr6, Yr7, Yr8, Yr17, and 32 was common within the Yr10 and Yr24 virulent races, and virulence for YrSp and Yr27 were observed in low frequency. Molecular genotyping of 209 isolates, including the Yr10 virulent races, was performed using 19 microsatellite markers (Ali et al. 2017; Rodriguez-Algaba et al. 2017) and aligned with the Puccinia striiformis nomenclature system of the Global Rust Reference Center (GRRC). The results showed that 66 isolates were identical to the genotyping lineage "ME2018" identified in Egypt in 2018 by GRRC. This genetic lineage has now been designated as PstS17 (Hovmøller et al. 2023). The durum wheat cultivars have always been resistant to yellow rust in the Middle East. Seedling tests of 50 durum advanced lines from CIMMYT's International Durum Wheat Yield Nursery showed LITs in 45 accessions (90%) against an avirulent race for Yr10 and Yr24 (PstS2), but only 12% remained resistant while tested with a PstS17 (virulent for Yr10 and Yr24). This observation provides compelling evidence of the Yr10 and/or Yr24 presence within tested durum wheat germplasm. Continued monitoring of virulence and resistance of wheat germplasm to yellow rust is critical for successful breeding for rust resistance.

Susceptible Cultivars Research Articles

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