Pink rot of potato (Solanum tuberosum), mainly mentioned to be caused by Phytophthora erythroseptica, is an important disease in many potato growing regions of the world, including the USA (Salas et al. 2000), Canada (Peters and Sturz 2001; Al-Mughrabi et al. 2007), China (Xuemei Zhang 2013), and India (Roy et al. 1992).

Pink rot is known to be caused by Phytophthora erythroseptica since 1949 in Nebraska, where Goss (1949) cited the occurrence of the pathogen first in Holland in 1914, then again in 1922, Scotland 1919, England and Wales 1921, Sumatra 1923, Bulgaria 1927, Maine in USA in 1938, Canada 1943, and Tasmania 1946. He also described the disease development, symptoms, and variety resistance, pointing out the high rate of pathogenicity of P. erythroseptica compared the other two pathogens; P. drechsleri and Pythium debaryanum. Pink rot and leak disease are collectively known as watery rot, and have had significant impacts for decades. In many potato growing soils in Canada, sometimes up to 50% of potatoes are impacted (Secor and Gudmestad 1999). Pink rot caused by P. erythroseptica, usually appears at late growing stages on the haulms and tubers when soil moisture content is high. Mainly the disease occurs in the field, it also cause high damage in the storages. Although pink rot is widespread in the water logged fields, the disease has also been observed in the warm, sandy soils with low organic content and pH (Miller and Miller 2008).

Pink rot of potato may be caused by other Phytophthora species including P. drechsleri, and P. cryptogea, as demonstrated through inoculation assays (Mostowfizadeh-Ghalamfarsa et al. 2006). Recently, the Phytophthora cryptogea species complex was re-evaluated, based on morphological and molecular characteristics, and differentiated four species including P. cryptogea sensu stricto, P. pseudocryptogea, P. erythroseptica and P. sp. kelmania, differentiating P. erythroseptica from the other species by its homothallic natural and its position in a strongly supported monophletic clade (Safaiefarahani et al. 2015)

Turkey produced about 4,800,000 tons of potato in 142,883 ha in 2017 (TSI 2017). Phytophthora cryptogea was reported from the eastern Anatolian region of Turkey, from Erzurum, causing similar symptoms of Pink rot (Çakır and Demirci 2012). During the 2018 potato growing season in a field near Konya province of Turkey, about 80% tuber rot was observed during harvest from a 5 ha potato field growing Everest potato variety. Tuber rot was also observed in around 50% of the stored potatoes that appeared healthy within the field.

The rotted tubers were taken to the laboratory and the symptoms were diagnosed. This paper describes the first occurrence of Phytophthora erythroseptica in Turkey, its identification and pathogenicity together with its distinction from P. cryptogea which was reported previously and similar Phytophthora species.

To isolate the pathogen, twenty infected tissue pieces from ten tubers selected from a bulk sample of 30 kg seed lot were surface sterilized with 1% sodium hypochlorite for three min. Then washed with sterile distilled water and plated on amended selective P5ARPNH-agar medium containing (as mg per l); pimaricin 5, ampicillin 250, rifampicin 10, PCNB 50, nystatin 50, hymexazole 50; (Jung et al. 1996), in grated carrot agar (GCA; prepared using 40 g thinly grated carrot and 18 g agar per l) (Akıllı et al. 2012) and incubated in the dark at 22 ± 2 °C for 5–7 days. Pure cultures of the pathogen were obtained by removing mycelial tips of non-septate hyphae, growing on the isolation media and plating on Amended Grated Carrot Agar (AGCA; as μg ml−1); β-sitosterol 30, thiamine hydrochloride 1 and tryptophan 20) (Akıllı et al. 2012), a medium to see sporangia, oospores and other morphological characteristics.

The identity of the pathogen was determined using both morphological and molecular characteristics. Colony growth pattern was determined on PDA, incubated under diurnal day light cycles at 22 ± 2 °C. Formation of sporangia, their shapes and caducity was investigated on the culture discs taken from AGCA and incubated in sterile and non-sterile soil extracts. Oospore formation was examined under a compound microscope on the culture, grown for three weeks on AGCA.

Molecular identification was done by analysing ITS, Cox and β-tubulin regions of the pathogen. DNA isolation of the isolate was performed using Qiagen blood and tissue kit by using small amount of mycelium, scraped from the actively growing, 5–7 days old cultures of PDA. Amplifications of ITS 4 and ITS 6, Cox and β-tubulin regions used for molecular characterization, were done as described by (Camele et al. 2005, Safaiefarahani et al. 2015) and sequence analysis was carried out by the laboratory; BM Laboratuvar sistemleri, Kızılırmak Mahallesi, 1441. cadde, MEVA iş merkezi No8/41, 06510, Ankara. Nucleotide sequences were blasted to NCBI GenBank and the isolates were identified according to the homology with sequences already present in the GenBank. The culture of our P. erythroseptica isolate was deposited at the culture collection of Nazife Tuatay Museum, run by General Directorate of food and Control, Ministry of Agriculture and Forestry under the name NTM-F-117. The sequences of ITS, Cox1 and β-tubulin genes were recorded in GenBank with the accession no’s of MK461967, MK572804 and MK572805 respectively.

Pathogenicity of the isolate was determined by placing 5 mm diameter culture disks of the P. erythroseptica grown on PDA into the wells drilled on surface sterilized potato tubers. The inoculated tubers were kept at 22 ± 2 °C in dark. Symptom development was observed after ten days from inoculation.

Eleven non septate mycelial fungal growth from twenty seed pieces were obtained and all of them showed similar morphological characteristics with homothallic nature, for this reason only one isolated was used for further studies.

The isolate produced typical symptoms of pink rot caused by P. erythroseptica as watery black lesions on the skin; dark brown watery leak on tubers. When cut into two, salmon colour on the flesh after 15–20 min when exposed to air (Fig. 1 a,b,c). Phytophthora cryptogea also produced salmon colour watery lesions when exposed to air (Fig. 1d).

Fig. 1
figure 1

Symptoms on the potato tubers produced by Phytophthora erythroseptica and P. cryptogea, a) Dark, watery blemishes, b) Watery leak on the newly cut surface infected by Phytophthora erythroseptica, c, d) Salmon coloured flesh exposed to air, infected by P. erythroseptica and P. cryptogea respectively

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The two Phytophthora spp., the one obtained from this study and other obtained previously are members of Phytophthora clade 8 and are mostly morphologically similar to each other having a petalloid colony growth on PDA (Fig.2a), non-papillate, non-caducous sporangia (Gallegly and Hong 2008). Sporangia of P. erythroseptica, on the other hand are variable, from ellipsoid to ovoid with both rounded and tapered base, the ellipsoidal sporangia being frequently constricted in the middle (Fig.2b). The major morphological difference between the two Phytophthora spp. was the homothallic nature of P. erythroseptica, having plerotic oospores mostly amphigynous but sometimes with paragynous antheridia (Fig.2c). Sporangia of P. cryptogea are also non-papillate, non-caducous, ovoid to obpyriform (Fig.2d), with variable sizes.

Fig. 2
figure 2

Some characteristics of Phytophthora erythroseptica and P. cryptogea causing pink rot on potato. a) Petalloid growth of P. erythroseptica on PDA, b) Sporangia of P. erythroseptica constricted at the middle, c) Aplerotic oospore of P. erythroseptica, with paragynous antheridia, d) Ovoid, obpyriform sporangium of P. cryptogea

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Molecular identification of Phytophthora species was outlined in Table 1.

Table 1 Molecular identification of Phytophthora species isolated from potato in Turkey
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Although more than one Phytophthora species were described to cause pink rot on potato, the most frequently occurring species is P. erythroseptica (Goss 1949). The other species, reported to cause pink rot symptoms are; P. arecae, P. cactorum, P. cinnamomi, P. cryptogea, P. drechsleri, P. erythroseptica, P. megasperma, P. pseudocryptogea, and P. sp. kelmania (Erwin and Ribeiro 2005; Mostowfizadeh-Ghalamfarsa et al. 2006; Safaiefarahani et al. 2015). For this reason, pink rot of potato should not be attributed to only one species, as to P. erythroseptica.

Although they have many similar morphological and cultural characteristics; the two species of Phytophthora causing pink rot on potato; P. erythroseptica and P. cryptogea, can be differentiated by its homothallic nature of P. erythroseptica produced on sterol containing media. The formerly reported species, P. cryptogea, should be interpreted with caution when the identification is based on sequences of ITS region, since the sequence of our isolate matched 99% to many isolates of P. cryptogea, 100% to three isolates of P. cryptogea. As stated by Safaiefarahani et al. (2015), when molecular characterisation of the Phytophthora species causing pink rot of potato is taken into consideration more gene sequences should be analysed. Differentiation of the two species can also be done by examining their morphological characteristics. Occurrence of oospores of P. erythroseptica on the growth media, such as CMA or GCA, having β sterols is an easy approach, though there are some differences on the sporangial shapes. This distinction of the oospores was also stated by Safaiefarahani et al. (2015) who investigated this Phytophthora complex in detail.

Phytophthora cryptogea was first reported by Çakır and Demirci (2014) on potato in Erzurum province of Turkey and with this study another species, P. erythroseptica was found another region of Turkey, from Konya. Their first occurrences in Turkey implies their import with seed potatoes, which is a common practice in Turkey. Phytophthora erythroseptica might be more important for Turkey since, it might remain in soil for a long time with its oospores and be distributed to other regions.