First Report of Pectobacterium carotovorum subsp. carotovorum on Spathiphyllum wallisii in Argentina

Peace lily (Spathiphyllum wallisii Regel) is a popular ornamental potted plant in Argentina. During May of 2008 (austral autumn), necrotic lesions of unknown etiology were observed on S. wallisii in a nursery in Pontevedra (34°45′6″S, 58°42′42″W). Plants first showed water-soaked areas starting from the leaf tips. Infected tissue became irregular, brown, dark-to-black lesions on leaves ~12 to 14 mm in diameter surrounded by yellowish haloes. Disease incidence approached 30%. Abundant bacterial streaming was observed from lesions when examined at ×100. Bacteria isolated from lesions formed white-to-cream, glistening, convex colonies on yeast dextrose calcium carbonate agar. Three bacterial strains isolated from different symptomatic plants were selected for comparative analysis with Pectobacterium carotovorum subsp. carotovorum type strain ATCC 15713. All were facultatively, anaerobic, gram-negative rods, pectolytic on crystal violet pectate agar, nonfluorescent on King's medium B, and elicited a hypersensitive response in tobacco plants. All strains were oxidase and arginine dihydrolase negative, fermented glucose, did not hydrolyze starch, did not produce lecithinase, indole or the blue pigment indigoidine, reduced nitrates, hydrolyzed gelatin and esculin, able to rot onion slices, caused soft rot of potato tubers, resistant to erythromycin, and grew at 37°C. Acid was produced from cellobiose, d-glucose, d-melibiose, d-mannitol, d-mannose, l-rhamnose, d-sucrose, and l-arabinose but not from inositol and d-sorbitol. Bacteria utilized N-acetyl-glucosamine and citrate but not tartrate, benzoate, or propionate. Their identity was confirmed by 16S rRNA gene sequencing of strain F402Pcc (GenBank Accession No. FJ717337) showing a 99% homology with that of strain ATCC 3326 (FJ 5958691). Pathogenicity was verified on S. wallisii, Dieffenbachia picta, Aglaonema commutatum, and Anthurium andraeanum within the Araceae family by spraying two plants per strain tested with bacterial suspensions (108 CFU/ml) in sterile distilled water with and without wounding the leaves with sterile needles. Controls were sprayed with sterile distilled water. After 48 h in a humidity chamber, inoculated plants and controls were maintained at 25 ± 3°C in a greenhouse. Water-soaked areas developed from 24 to 48 h after inoculation and became necrotic within 4 to 5 days. Lesions expanded to resemble natural infection in S. wallisii within 20 days, while in the rest of the hosts tested, lesions were smaller and remained brown surrounded by yellowish haloes. All strains were reisolated from each host tested. The original and all reisolated strains were compared by enterobacterial repetitive intergeneric consensus-PCR (4) confirming that DNA fingerprints of the reisolated strains were identical to those of the original strains. No lesions were observed on controls. The pathogen was identified as P. carotovorum subsp. carotovorum based on biochemical, physiological, pathogenicity tests, and 16S rRNA sequencing (1–3).To our knowledge, this is the first report of this pathogen on S. wallisii in Argentina although it has been reported as causing tomato pith necrosis (1) and soft rot of vegetables after harvest (3). References: (1) A. M. Alippi et al. Plant Dis. 81:230, 1997. (2) L. Gardan et al. Int. J. Syst. Evol. Microbiol. 53:381, 2003. (3) L. Halperin and L. S. Spaini. Rev. Argent. Agron. 6:261, 1939. (4) F. J. Louws et al. Appl. Environ. Microbiol. 60:2286, 1994.

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First Report of Pectobacterium punjabense Causing Blackleg and Soft Rot on Potato in Hebei and Fujian Province, China

Plant Disease ◽

10.1094/pdis-12-21-2731-pdn ◽

2022 ◽

Author(s):

Utpal Handique ◽

Yaning Cao ◽

Dekang Wang ◽

Ruofang Zhang ◽

Wensi Li ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Housekeeping Genes ◽

Soft Rot ◽

Rrna Gene ◽

Pectobacterium Carotovorum ◽

Fujian Province ◽

First Report ◽

Potato Plants ◽

Carotovorum Subsp

Pectobacterium spp. and Dickeya spp. cause blackleg and soft rot on potato worldwide (Charkowski, 2018). Potato plants (cv. Favorita or Jizhang 8#) with blackleg symptoms (vascular browning of crown stems, Fig. S1) were observed in the field in Zhangjiakou, Hebei province in 2018, and in Ningde, Fujian Province in 2019, in China. The disease incidence was around 50% and 10% in Zhangjiakou (5 ha) and Ningde (4 ha), respectively. Diseased plants (3 from each site) were collected to isolate the pathogen. Blackleg symptomatic stems were soaked in 75% ethanol for 2 min, rinsed and ground in sterile distilled water. Serial tenfold dilutions of the above solution were plated onto the crystal violet pectate agar (CVP) plate (Ge et al., 2018). Two to 3 days after incubation at 28°C, 4 bacterial colonies in total which digested pectin from the media and developed pit on CVP plates were purified and sequenced for identification using the universal 16S rRNA gene primer set 27F/1492R (Monciardini et al., 2002). Two colony sequences that showed more than 99% sequence identity to Pectobacterium punjabense type strain SS95 (MH249622) were submitted to the GenBank ( accession numbers: OK510280, MT242589). Additionally, six housekeeping genes proA (OK546205, OK546199), gyrA (OK546206, OK546200), icdA (OK546207, OK546201), mdh (OK546208, OK546202), gapA (OK546209, OK546203), and rpoS (OK546210, OK546204) of these two isolates were amplified and sequenced (Ma et al., 2007, Waleron et al., 2008). All strains show 99% to 100% identity with MH249622T . Phylogenetic trees based on 16S rRNA gene sequences (Fig. S2) and concatenated sequences of the housekeeping genes (Fig. S3) of the 2 isolates were constructed using MEGA 6.0 software (Tamura et al., 2013). Koch’s postulate was performed on potato seedlings and potato tubers (cv. Favorita) by injecting 100 μl bacterial suspension (105 CFU/ml) or sterile phosphate-buffered solution into the crown area of the stems or the tubers and kept at 100% humidity and 21°C for 1 day. Four days after inoculation, the infected area of the inoculated seedlings rotten and turned black, while the controls were symptomless (Fig. S4). Two days after inoculation, the infected tubers rotten and turned black, while the controls were symptomless (Fig. S4). Bacterial colonies were reisolated from these symptomatic tissues and identified using the same methods described above. Blackleg on potato plants or soft rot on potato has been reported to be caused by Pectobacterium atrosepticum, Pectobacterium carotovorum subsp. carotovorum, Pectobacterium carotovorum subsp. brasiliense, Pectobacterium parmentieri, Pectobacterium polaris in China (Zhao et al., 2018; Cao et al., 2021; Wang et al., 2021). To our knowledge, this is the first report of blackleg/soft rot of potato caused by Pectobacterium punjabense in China. We believe that this report will draw attention to the management of this pathogen in China.

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First Report of Pectobacterium parmentieri Causing Blackleg on Potato in Inner Mongolia, China

Plant Disease ◽

10.1094/pdis-11-20-2502-pdn ◽

2021 ◽

Author(s):

Yaning Cao ◽

Qinghua Sun ◽

Zhiwen Feng ◽

Utpal Handique ◽

Jian Wu ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Inner Mongolia ◽

Rrna Gene ◽

Pectobacterium Carotovorum ◽

Distilled Water ◽

First Report ◽

Potato Plants ◽

Sequence Identity ◽

Carotovorum Subsp

Blackleg on potato plants (Solanum tuberosum) is caused by Pectobacterium spp. and Dickeya spp. (Charkowski, 2018) worldwide. From June to August in both 2018 and 2019, cases of blackleg were investigated in potato-producing areas in Hulunbuir, Ulanqab, and Hohhot in Inner Mongolia, China. The total surveyed field area was about 200 hectares. The plants showed typical blackleg symptoms, such as black and stunted stems or curled leaves (Fig. S1), and the number of infected plants were counted. The disease showed an incidence of around 8%. Five diseased plants were collected from a 10 ha potato field with approximately 75,000 potato plants (cv. mainly Favorita and Xisen) per hectare. Two-centimeter-long samples of symptomatic stems were removed from the selected plants using a sterile scalpel. The surfaces of the samples were disinfected with 75% ethanol for 2 min. They were then rinsed with sterile distilled water and soaked in 5 ml sterile distilled water for 30 min. Aliquots of three tenfold dilutions of this solution were plated onto the crystal violet pectate agar (CVP) plate and incubated for 3 days at 28°C (Ge et al., 2018). A single bacterial colony that showed pitting on CVP plates (Fig. S2) was picked with a toothpick, streaked onto nutritional agar (She et al., 2013) to obtain pure colonies. Amplification of a 1.4-kb segment containing 16S rRNA gene was performed on the pure colonies using the universal primer set 27F/1492R (Monciardini et al., 2002). The amplicons were sequenced and submitted to the GenBank Nucleotide Basic Local Alignment Search Tool analysis. The 16S rRNA gene sequences of four isolates (GenBank accession numbers: MN626412, MN626449, MN625916, and MT235556) showed more than 99% sequence identity to Pectobacterium parmentieri type strain RNS 08-42-1A (NR_153752.1) (Fig. S3). Six housekeeping genes proA (MT427753-MT427756), gyrA (MT427757–MT427760), icdA (MT427761-MT427764), mdh (MT427765–MT427768), gapA (MT427769-MT427772), and rpoS (MT427773–MT427776) of these four isolates were amplified and sequenced (Ma et al., 2007, Waleron et al., 2008). All sequences showed 99% to 100% sequence identity with Pectobacterium parmentieri strains. Phylogenetic trees (Fig. S4) were constructed by multi-locus sequence analysis (MLSA) using MEGA 6.0 software (Tamura et al., 2013). The samples were tested against Koch’s postulates on potato seedlings (cv. Favorita) by injecting 100 μl bacterial suspension (107 CFU/ml) or sterile phosphate buffered solution into the stems 2 cm above the base (Ge et al., 2018). The seedlings were incubated at 21°C and 80% humidity (She et al., 2013). Three to 5 days after inoculation, only infected seedlings showed similar symptoms as those observed in the field: the infected area turned black and rotten (Fig. S5). Bacterial colonies isolated from these symptomatic seedlings were identified using the same methods described above and were identified as inoculated Pectobacterium parmentieri strains. Blackleg on potato plants has been reported to be caused by Pectobacterium atrosepticum, Pectobacterium carotovorum subsp. carotovorum, and Pectobacterium carotovorum subsp. brasiliense in China (Zhao et al., 2018). To our knowledge, this is the first report of blackleg of potato caused by Pectobacterium parmentieri in Inner Mongolia, China. We believe that this report will draw attention to the identification of this pathogen, which is essential to disease management.

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First Report of Pectobacterium carotovorum subsp. carotovorum Causing Soft Rot of Orostachys japonica in Korea

Plant Disease ◽

10.1094/pdis-10-13-1039-pdn ◽

2014 ◽

Vol 98 (7) ◽

pp. 989-989 ◽

Cited By ~ 2

Author(s):

W. Cheon ◽

Y. H. Jeon

Keyword(s):

16S Rrna ◽

Pathogenic Bacteria ◽

Sequence Similarity ◽

Soft Rot ◽

Bacterial Suspension ◽

Pectobacterium Carotovorum ◽

First Report ◽

Biolog System ◽

Carotovorum Subsp ◽

Physiological Tests

Orostachys japonica (Maxim) A. Berger is an important traditional medicine in Korea. The extract of this plant has antioxidant activity and suppresses cancer cell proliferation (1). From summer through fall of 2012 and 2013, a high incidence (~10% to 30%) of disease outbreaks of all plants characterized by water-soaked lesions and soft rot with a stinky odor was observed in cultivated O. japonica around Uljin (36°59′35.04″N, 126°24′1.51″E), Korea. Water-soaked lesions were first observed on the stem base of plants. Subsequently, the plants collapsed, although the upper portion remained asymptomatic. Thereafter, the lesions expanded rapidly over the entire plant. To isolate potential pathogens from infected leaves, small sections (5 to 10 mm2) were excised from the margins of lesions. Ten bacteria were isolated from ten symptomatic plants. Three representative isolates from different symptomatic plants were used for identification and pathogenicity tests. Isolated bacteria were gram negative, pectolytic on crystal violet pectate agar, nonfluorescent on King's medium B, and elicited a hypersensitive response in tobacco plants. All isolates caused soft rot of potato tubers. These isolates also differed from isolates of Erwinia chrysanthemi (Ech) that they were insensitive to erythromycin and did not produce phosphatase. These isolates differed from known strains of E. carotovora subsp. atroseptica in that they did not produce reducing substances from sucrose (2). Use of the Biolog GN microplate and the Release 4.0 system identified the isolate as Pectobacterium carotovorum subsp. carotovorum with 81.2% similarity. The 16S rRNA of the isolated bacteria was amplified by PCR and sequenced as described by Weisburg et al. (3). A BLAST analysis for sequence similarity of the 16S rRNA region revealed 99% similarity with nucleotide sequences for P. carotovorum subsp. carotovorum isolates (KC790305, KC790280, JF926758, JX196705, and AB680074). The pathogenicity of three bacterial isolates was examined on three 2-year-old O. japonica plants by adding 50 μl of a bacterial suspension containing 108 CFU/ml when wounding the leaves with sterile needles. Ten control plants were inoculated with sterilized water. After inoculation, plants were maintained in a growth chamber at 25°C with relative humidity ranging from 80 to 90%. After 2 to 3 days, tissue discoloration, water-soaked lesions, and soft rot developed around the inoculation point. Severe symptoms of soft rot and darkening developed on leaves of inoculated plants within 3 to 5 days after inoculation. All controls remained healthy during these experiments. The bacterial strains re-isolated from the parts of the leaf showing the symptoms and identified as P. carotovorum subsp. carotovorum on the basis of the biochemical and physiological tests, as well as Biolog system. The results obtained for pathogenicity, Biolog analysis, and molecular data corresponded with those for P. carotovorum subsp. carotovorum. To our knowledge, this is the first report of the presence of P. carotovorum on O. japonica in Korea. References: (1) C.-H. Kim et al. Kor. J. Med. Crop Sci. 11:31, 2003. (2) N. W. Schaad et al. Erwinia Soft Rot Group. Page 56 in: Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3rd ed. N. W. Schaad et al. eds. American Phytopathological Society, St. Paul. MN, 2001. (3) W. G. Weisburg et al. J. Bacteriol. 173:697, 1991.

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First Report of Pectobacterium polaris Causing Blackleg on Potato in Inner Mongolia and Sichuan Province, China

Plant Disease ◽

10.1094/pdis-08-21-1788-pdn ◽

2021 ◽

Author(s):

UTPAL HANDIQUE ◽

Yaning Cao ◽

Zhiwen Feng ◽

Qinghua Sun ◽

Ruofang Zhang ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Inner Mongolia ◽

Housekeeping Genes ◽

Sichuan Province ◽

Rrna Gene ◽

Pectobacterium Carotovorum ◽

First Report ◽

Potato Plants ◽

Carotovorum Subsp

Pectobacterium spp. and Dickeya spp. cause Blackleg on potato worldwide (Charkowski, 2018). Potato plants (cv. Innovator V4 or Favorita) with blackleg symptoms (vascular browning of crown stems or curled leaves, Fig. S1) were observed in the field in Xilingol League, Inner Mongolia in 2018, and in Chengdu, Sichuan Province in 2020, in China. The disease incidence were around 10% and 20% in Xilingol League (20 ha) and Chengdu (40 ha), respectively. Diseased plants (5 from Xilingol League, and 2 from Chengdu) were collected to isolate the pathogen. Blackleg symptomatic stems were soaked in 75% ethanol for 2 min, rinsed and ground in sterile distilled water. Serial tenfold dilutions of the above solution were plated onto the crystal violet pectate agar (CVP) plate (Ge et al., 2018). Two to 3 days after incubation at 28°C, the bacterial colonies which digested pectin from the media and developed pit on CVP plates were purified and sequenced for identification using the universal 16S rRNA gene primer set 27F/1492R (Monciardini et al., 2002). Three colony sequences that showed more than 99% sequence identity to Pectobacterium polaris type strain NIBIO1392 (NR_159086.1) were submitted to the GenBank ( accession numbers: MT242579, MT242580, and MZ489432). Additionally, six housekeeping genes proA (MZ39581–MZ395583), gyrA (MZ395569–MZ395571), icdA (MZ395572–MZ39574), mdh (MZ395575–MZ395577), gapA (MZ395578– MZ395580), and rpoS (MZ39584–MZ395586) of these three isolates were amplified and sequenced (Ma et al., 2007, Waleron et al., 2008). All strains show 99% to 100% identity with Pectobacterium polaris strain NIBIO1392. Phylogenetic trees based on 16S rRNA gene sequences (Fig. S2) and concatenated sequences of the housekeeping genes (Fig. S3) of the 3 isolates were constructed using MEGA 6.0 software (Tamura et al., 2013). Koch’s postulate was performed on potato seedlings (cv. Favorita) by injecting 100 μl bacterial suspension (107 CFU/ml) or sterile phosphate-buffered solution into the crown area of the stems and kept at 80% humidity and 21°C for 2 days. Seven days after inoculation, the infected area of the inoculated seedlings rotten and turned black or even lodged, while the controls were symptomless (Fig. S4). It was observed that isolate MZ489432 from Chengdu, Sichuan Province was more virulent than the isolates from Xilingol League (Fig. S4). Bacterial colonies were reisolated from these symptomatic seedlings and identified using the same methods described above. Blackleg on potato plants has been reported to be caused by Pectobacterium atrosepticum, Pectobacterium carotovorum subsp. carotovorum, Pectobacterium carotovorum subsp. brasiliense, and Pectobacterium parmentieri in China (Zhao et al., 2018; Cao et al., 2021). To our knowledge, this is the first report of blackleg of potato caused by Pectobacterium polaris in China. We believe that this report will draw attention to the management of this pathogen in China.

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First Report of Pectobacterium carotovorum subsp. brasiliense Causing Soft Rot on Squash and Watermelon in Serbia

Plant Disease ◽

10.1094/pdis-12-18-2213-pdn ◽

2019 ◽

Vol 103 (10) ◽

pp. 2667-2667 ◽

Cited By ~ 3

Author(s):

N. Zlatković ◽

A. Prokić ◽

K. Gašić ◽

N. Kuzmanović ◽

M. Ivanović ◽

...

Keyword(s):

Soft Rot ◽

Pectobacterium Carotovorum ◽

First Report ◽

Carotovorum Subsp

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First Report of Pectobacterium carotovorum subsp. brasiliense Causing Soft Rot on Potato and Other Vegetables in Poland

Plant Disease ◽

10.1094/pdis-02-15-0180-pdn ◽

2015 ◽

Vol 99 (9) ◽

pp. 1271-1271 ◽

Cited By ~ 16

Author(s):

M. Waleron ◽

K. Waleron ◽

E. Lojkowska

Keyword(s):

Soft Rot ◽

Pectobacterium Carotovorum ◽

First Report ◽

Carotovorum Subsp

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First Report of Pectobacterium aroidearum and Pectobacterium carotovorum subsp. brasiliensis Causing Soft Rot of Cucurbita pepo in Brazil

Plant Disease ◽

10.1094/pdis-08-16-1168-pdn ◽

2017 ◽

Vol 101 (2) ◽

pp. 379 ◽

Cited By ~ 12

Author(s):

A. J. G. Moraes ◽

E. B. Souza ◽

R. L. R. Mariano ◽

A. M. F. Silva ◽

N. B. Lima ◽

...

Keyword(s):

Cucurbita Pepo ◽

Soft Rot ◽

Pectobacterium Carotovorum ◽

First Report ◽

Carotovorum Subsp

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The First Report of Stolbur Phytoplasma Associated with Phyllody of Calendula officinalis in Serbia

Plant Disease ◽

10.1094/pdis-01-14-0085-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1152-1152 ◽

Cited By ~ 3

Author(s):

S. Pavlovic ◽

M. Starovic ◽

S. Stojanovic ◽

G. Aleksic ◽

S. Kojic ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Natural Infection ◽

Cytotoxic Activities ◽

Rrna Gene ◽

Calendula Officinalis ◽

Aster Yellows ◽

First Report ◽

Pot Marigold ◽

Stolbur Phytoplasma

Pot marigold (Calendula officinalis L.) is native to southern Europe. Compounds of marigold flowers exhibit anti-inflammatory, anti-tumor-promoting, and cytotoxic activities (4). In Serbia, pot marigold is cultivated as an important medicinal and ornamental plant. Typical phyllody, virescence, proliferation of axillary buds, and witches' broom symptoms were sporadically observed in 2011 in Pancevo plantation, Serbia (44°51′49″ N, 20°39′33″ E, 80 m above sea level). Until 2013, the number of uniformly distributed affected pot marigold plants reached 20% in the field. Due to the lack of seed production, profitability of the cultivation was seriously affected. Leaf samples from 10 symptomatic and 4 symptomless marigold plants were collected and total nucleic acid was extracted from midrib tissue (3). Direct PCR and nested PCR were carried out with primer pairs P1/16S-SR and R16F2n/R16R2n, respectively (3). Amplicons 1.5 and 1.2 kb in length, specific for the 16S rRNA gene, were amplified in all symptomatic plants. No PCR products were obtained when DNA isolated from symptomless plants was used. Restriction fragment length polymorphism (RFLP) patterns of the 1.2-kb fragments of 16S rDNA were determined by digestion with four endonucleases separately (TruI1, AluI, HpaII, and HhaI) and compared with those of Stolbur (Stol), Aster Yellows (AY), Flavescence dorée-C (FD-C), Poinsettia Branch-Inducing (PoiBI), and Clover Yellow Edge (CYE) phytoplasmas (2). RFLP patterns from all symptomatic pot marigold plants were identical to the Stol pattern, indicating Stolbur phytoplasma presence in affected plants. The 1.2-kb amplicon of representative Nv8 strain was sequenced and the data were submitted to GenBank (accession no. KJ174507). BLASTn analysis of the sequence was compared with sequences available in GenBank, showing 100% identity with 16S rRNA gene of strains from Paeonia tenuifolia (KF614623) and corn (JQ730750) from Serbia, and peach (KF263684) from Iran. All of these are members of the 16SrXII ‘Candidatus Phytoplasma solani’ group, subgroup A (Stolbur). Phytoplasmas belonging to aster yellows (16SrI) (Italy and Canada) and peanut witches' broom related phytoplasma (16SrII) group (Iran) have been identified in diseased pot marigold plants (1). To our knowledge, this is the first report of natural infection of pot marigold by Stolbur phytoplasma in Serbia. References: (1) S. A. Esmailzadeh-Hosseini et al. Bull. Insectol. 64:S109, 2011. (2) I. M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (3) J. P. Prince. Phytopathology 83:1130, 1993. (4) M. Ukiya et al. J. Nat. Prod. 69:1692, 2006.

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