The Australian ginger industry is small (less than 1 % contribution to world production), but it is a cash crop for around 30 full time growers, who produce approximately AUD 15.6 million at farm gate value per year (Camacho and Brescia 2009). The annual production of the whole Australian ginger industry is estimated at about 8,000 tonnes with almost half of this delivered to the domestic fresh market and the rest for the processing industry (Camacho and Brescia 2009). In 2007, farmers unexpectedly experienced severe losses (up to 100 % in some ginger fields) due to Pythium Soft Rot (PSR). In 2009, Pythium myriotylum was reported as a causal agent responsible for the outbreak in Australia (Stirling et al. 2009). However, from our recent research, it is proposed that there is probably more than one Pythium sp. associated with PSR of ginger in Australian fields. In 2012, ginger with PSR symptoms caused by Pythium spp. was sampled from a representative selection of farms at Yandina, on the Sunshine Coast, Queensland, Australia. Along with nine Pythium spp. isolated from PSR ginger and soil around the ginger rhizome, ten isolates of a Pythium-like organism were also obtained from PSR ginger sampled from two farms (farm 1 at 26° 32′S, 152°56′E and farm 2 at 26° 33′S, 152°56′E) at Yandina. Isolations had been undertaken by excising sections (5 mm2) of rhizome with PSR symptoms, quickly surface decontaminating (around 10 s) with 25 % bleach containing 1 % HOCl, then washing twice with sterilized distilled water, blotted dry with autoclaved paper towel. The sections were then transferred on to Petri plates with corn meal agar with an amendment of 50 μg/ml Penicillin, 50 μg/ml Polymyxin, and 25 μg/ml Pirmaricin (CMA + 3P). The Petri plates were incubated in the dark overnight at 27 °C, then sections taken from the growing edge of the colonies were transferred onto 1.5 % water agar, incubated again under the same conditions for another night, after which hyphal tips of each of the isolates were excised out under an inverted compound microscope (Leica) and placed onto full strength potato dextrose agar (PDA Difco).

All ten Pythium-like isolates were considered to be heterothallic as attempts to produce sexual structures on dual cultures were not successful. Using the keys of Jee et al. (2000) and Huang et al. (2013), these isolates were then identified as Pythiogeton ramosum based on the typical features of sickle-shape appressoria, terminal, subspherical, ellipsoidal, utriform, or bursiform sporangia with apical beaks (Fig. 1) which measured from 20–214 × 18–48 μm. Colonies on PDA and CMA media did not show any special pattern, mycelia were quite sparse on media and rarely aerial (Fig. 1). The hyphae were aseptate, hyaline, smooth, 2–5 (−7) μm wide. The hyphae also had many shorter branches which were usually bearing appressoria and growing at a right angle with the main hyphae. On solid culture media, old sporangia formed vacuoles, which had the potential to be misidentified as oospores. Two isolates of this species description were deposited with Queensland Plant Pathology Herbarium with BRIP numbers (BRIP59948 and BRIP59949). For confirmation of species identification, ITS regions were amplified and analysed. Pure DNA from the mycelial mats growing in potato broth was obtained using CTAB protocol (Doyle and Doyle 1990). A PCR reaction was carried out in a Eppendorf Mastercycler® using the two universal primers ITS1 and ITS4 (White et al. 1990) to allow amplification (approximately 860 bp) of ITS the region including the 5.8S rRNA encoding region. The Genbank database was searched (BLAST) for similar sequences resulting in 100 % homology with a Py. ramosum isolate JQ610190.1 from Tawain (Huang et al. 2013). The sequences were also deposited with Genbank under KF151204 and KF151205.

Fig. 1
figure 1

Py. ramosum growing on PDA showing an indistinctive pattern (left), terminal bursiform sporangia with different sizes and shapes of globe and elongate (right) (Bar = 20 μm)

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Pythiogeton ramosum was first isolated from plant debris by Minden (1916) and was later commonly recovered from materials under anaerobic conditions (Czeczuga et al. 2005; El-Hissy et al. 1994; Huang et al. 2013; Nascimento et al. 2012). Py. ramosum is not well studied and was believed to be a saprophyte and until recently its pathogenicity on living plants had not been confirmed (Ann et al. 2006; Huang et al. 2013). In this study we initially attempted to carry out some in-vitro pathogenicity tests for our isolates to check if they were pathogenic on tested materials.

Pathogenicity tests were conducted in-vitro on ginger (Zingiber officinale), carrot (Daucus carota), radish (Raphanus sativus), sweet potato (Ipomoea batatas) and potato (Solanum tuberosum) rhizome/tubers. The experiment was based on descriptions of Le et al. (2010) to test the pathogenicity and extent of rotting from isolates (BRIP59948 and BRIP59949) inoculated on excised rhizome/tuber sections (1 × 1 × 5.5 cm) at 27 °C. All tested isolates of Py. ramosum were shown to be non-pathogenic on excised ginger and radish pieces, but they were pathogenic on carrot, sweet potato and potato tubers. The tested tubers were decayed, soft and brown in colour and Py. ramosum was reisolated after placing decayed tissue onto CMA+3P. Additionally, pre-emerging damping off tests were carried out in Petri plates according to descriptions of Zhang and Yang (2000) and disease indices (scale from 0 to 1) were calculated to evaluate aggressive levels of Py. ramosum on some vegetable crops including bean (Phaseolus vulgaris), capsicum (Capsicum annuum), and cauliflower (Brassica oleracea var.botrytis). Disease indices recorded from 0.13 to 0.50 indicated that Py. ramosum was able to attack and cause pre-emerging damping off on these crop species (Fig. 2). The results from this study allowed us to conclude that Py. ramosum is pathogenic on several hosts and this is the first report of the pathogenicity of Py. ramosum on either excised rhizome/tuber sections or seeds/seedlings of the tested materials. Although Py. ramosum was isolated directly from soft rot ginger, it was non-pathogenic on ginger at 27 °C. Therefore, more pathogenicity tests including on living plants at different temperatures will be warranted.

Fig. 2
figure 2

Cauliflower seedlings following inoculation with Py. ramosum (left) showing necrosis, brown rot in comparison to white and healthy seedlings in the uninoculated controls (right)

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