First Report of Pseudomonas savastanoi Causing Bacterial Leaf Spot of Mandevilla sanderi in Slovenia

Abstract

Foliar necrotic spots with narrow chlorotic halos were observed on different cultivars of Brazilian Jasmine (Mandevilla sanderi) during spring 2010 in several commercial greenhouses in Slovenia. Up to 70% were symptomatic and were unmarketable. No galls were observed on the stems of symptomatic plants. Circular, flat, granulated colonies with entire margins were isolated from symptomatic leaves of two plants from different greenhouses on King's B medium (KB). The isolates were negative for levan, oxidase, pectinolytic and arginine dihydrolase activity. They caused a hypersensitive reaction on tomato but not on tobacco cv. White Burley. Isolates were weakly fluorescent on KB under UV light. One isolate per sample (NIB Z 1413 and 1415) was further characterized. Partial sequences of 16S rDNA (1; GenBank KM603318 of 722 bp, KM603319 of 686 bp) grouped the isolates within genomospecies 2 of Pseudomonas. Repetitive polymerase chain reaction (PCR) assay using the BOXA1R primer (5) resulted in highly similar DNA fragment banding patterns of the two NIB Z isolates and other reference strains of genomospecies 2 (minimum 95.1% identity with Pearson's correlation). Partial sequences of rpoD (3) of the two Slovenian isolates (600 bp; GenBank KJ744202, KJ744201) were identical to the P. savastanoi isolate from Mandevilla B200 (W. Wohanka, Germany; GenBank KJ744203) and P. s. pv. nerii strain NCPPB 3334 (GenBank AB039513). The sequences differed in two nucleotides relative to the sequence of the pathotype strain of pv. nerii NCPPB 3278 (positions 487 and 510 relative to GenBank FN433279) and had an insertion of six nucleotides compared to available P. savastanoi pv. savastanoi rpoD sequences (NZ_JOJV01000073, CM001834). Pathogenicity of isolated bacteria (two isolates) was determined on M. sanderi cv. Pretty Rose inoculated by two different methods, spraying foliage and pricking stems. The abaxial and adaxial surfaces of leaves were sprayed with a 30-ml bacterial suspension (5 × 106 CFU/ml). Three plants were inoculated with each isolate: NIB Z 1413 and 1415 and the reference strain NCPPB 3278. Necrotic spots developed on leaves after 14 days of incubation, under >80% high relative humidity, with 16 h of daylight at 25°C and 8 h of dark at 21°C. One month after inoculation, necrosis also developed on stems and new growth. Inoculation of bacteria by pricking nodes of healthy M. sanderi cv. Pretty Rose with a needle dipped in the isolates grown on KB for 24 h (each of NIB Z 1413, 1415, and NCPPB 3278 for positive control) led to development of galls in 14 days at the inoculation points. The re-isolation was performed separately from necrotic spots on leaves, stems, new growth above the inoculation points, and galls. The BOX-PCR profiles of the bacteria isolated from symptomatic tissues were identical to the original profiles, thus confirming the systemic spread of the bacteria. None of the three negative control plants sprayed with 0.01M MgSO4 or pricked with a sterile needle developed symptoms. This is the first report of P. savastanoi on Mandevilla sanderi plants in greenhouse production in Slovenia. The galls caused by P. savastanoi have previously been reported from the United States (4) and Germany (2). This report broadens the geographical area where P. savastanoi, causing both galls on stems and necrotic spots on leaves, can be found in commercial production of Mandevilla spp. References: (1) U. Edwards et al. Nucleic Acids Res. 17:7843, 1989. (2) N. Eltlbany et al. Appl. Environ. Microbiol. 78:8492, 2012. (3) N. Parkinson et al. Plant Pathol. 60:338, 2011. (4) M. L. Putnam et al. Phytopathology 100:S104, 2010. (5) J. Versalovic et al. Methods Mol. Cell Biol. 5:25, 1994.

Pseudomonas savastanoi pv. mandevillae pv. nov., a Clonal Pathogen Causing an Emerging, Devastating Disease of the Ornamental Plant Mandevilla spp.

“…However, cross-pathogenicity tests performed in olive and oleander could not confidently classify the isolates as members of any of these two pathovars (Eltlbany et al 2012). Although partial sequencing of the rpoD gene from two Slovenian isolates showed a closer proximity to Psn strains (Pirc et al 2015), results from pathogenicity tests in oleander plants were not reported.…”

“…This emergent disease is caused by the most prevalent bacterial pathogen of dipladenia, Pseudomonas savastanoi. The first report of this disease dates back to 2010 in the USA (Putnam et al 2010), with European outbreaks occurring over the following years in France, Germany (Eltlbany et al 2012), Slovenia (Pirc et al 2015) and Spain (Caballo-Ponce and Ramos 2016). With Spain and Italy as the main producers of dipladenia in Europe, this emergent disease has also become a serious concern for European growers.…”

“…With Spain and Italy as the main producers of dipladenia in Europe, this emergent disease has also become a serious concern for European growers. The rapid spread of the pathogen within greenhouses, difficulties for disease management and the visual symptoms of MaLSS are responsible for the loss of a large number of plants, which are classified as unmarketable, as reported for up to 70% of the dipladenia in Slovenian greenhouses (Pirc et al 2015).…”

“…Additionally, hybridization analyses of Southern-blotted plasmid restriction digests with probes generated from plasmid-borne P. syringae genes, showed that the profiles obtained from seven dipladenia isolates were highly similar but clearly distinct from those of Psv and Psn strains, which showed high strainspecific variability (Eltlbany et al 2012). Moreover, and unlike most P. savastanoi strains, dipladenia isolates do not trigger a hypersensitive response (HR) in tobacco leaves (Pirc et al 2015;Caballo-Ponce and Ramos 2016). All these initial observations suggest that P. savastanoi strains infecting dipladenia might constitute a separate, homogeneous lineage holding a distinct set of plasmids.…”

Knots Untie: Molecular Determinants Involved in Knot Formation Induced by Pseudomonas savastanoi in Woody Hosts

“…In contrast, the host range of Psr isolates is restricted to Spanish broom (Janse, 1982; Iacobellis et al, 1998; Ramos et al, 2012). Knot formation by P. savastanoi has also been described in several other hosts, including jasmine (Gardan et al, 1992b), privet (Gardan et al, 1992b), mock privet (Gardan et al, 1992b), forsythia (Bradbury, 1986), buckthorn (Temsah et al, 2007a), fontanesia (Mirik et al, 2011), pomegranate (Bozkurt et al, 2014), myrtle (Goumas et al, 2000; Temsah et al, 2007b; Cinelli et al, 2014) and dipladenia (Putnam et al, 2010; Eltlbany et al, 2012; Pirc et al, 2014; Caballo-Ponce and Ramos, 2016) ( Table 1 ). Although these P. savastanoi hosts are classified in several plant families, over 50% of them belong to the Oleaceae family.…”