Characterization of Pectobacterium carotovorum subsp. carotovorum isolates from a recent outbreak on cabbage in Bosnia and Herzegovina

The causal agent of soft rot disease associated with a cabbage outbreak in Semberija region, Bosnia and Herzegovina, in 2018 was identified and characterized. Symptoms appeared in the form of water-soaked lesions on leaves and specific odour. Disease incidence ranged from 20% to 30%. The causal pathogen was isolated on nutrient agar (NA), King?s B and crystal violet pectate (CVP) media. Eight creamy-white, round and convex bacterial isolates, which produced characteristic pits on CVP medium were taken as representative. They were gram negative, facultative anaerobe, oxidase negative, catalase positive, nonfluorescent on King?s B medium, levan and arginine dehydrolase negative. The isolates were able to cause soft rot on cabbage and potato tuber slices 24 h after inoculation under conditions of high relative humidity. Polymerase chain reaction (PCR) was performed for preliminary identification by using three specific primer sets: F0145/E2477 (specific for Pectobacterium carotovorum subsp. carotovorum), Br1f/L1r (specific for P. carotovorum subsp. brasiliensis) and ECA1f/ECA2r (specific for P. atrosepticum). All isolates produced the band size of 666 bp with F0145/E2477 primer pair, indicating that they belong to the species P. carotovorum subsp. carotovorum. Further genetic characterization was based on sequence analysis of the gapA and mdh housekeeping genes. BLAST analysis confirmed 99.39% (Q. cover 100%, E. value 0.0) and 100% (Q. cover 100%, E. value 0.0) identity of the isolates with P. carotovorum subsp. carotovorum strains deposited in the NCBI database as M34 (KY047594) for gapA and Pcc t0437 (KC337296) for mdh genes, respectively. Phylogenetic analysis showed genetic homogeneity among the cabbage isolates.

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Efficacy of potent antagonistic yeast Wickerhamiella versatilis against soft rot disease of potato caused by Pectobacterium carotovorum subsp. carotovorum

Archives of Phytopathology and Plant Protection ◽

10.1080/03235408.2019.1693236 ◽

2019 ◽

Vol 52 (15-16) ◽

pp. 1125-1148 ◽

Cited By ~ 1

Author(s):

Elhagag Ahmed Hassan ◽

Hadeel M. M. Khalil Bagy ◽

Shymaa R. Bashandy

Keyword(s):

Soft Rot ◽

Pectobacterium Carotovorum ◽

Soft Rot Disease ◽

Antagonistic Yeast ◽

Carotovorum Subsp ◽

Rot Disease

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The Ribosomal Protein RplY Is Required for Pectobacterium carotovorum Virulence and Is Induced by Zantedeschia elliotiana Extract

Phytopathology ◽

10.1094/phyto-04-17-0161-r ◽

2017 ◽

Vol 107 (11) ◽

pp. 1322-1330 ◽

Cited By ~ 6

Author(s):

Huan Jiang ◽

Mengyi Jiang ◽

Liuke Yang ◽

Peiyan Yao ◽

Lin Ma ◽

...

Keyword(s):

Plant Extract ◽

Plant Cell Wall ◽

Insertion Site ◽

Bacterial Pathogen ◽

Soft Rot ◽

Kanamycin Resistance ◽

Pectobacterium Carotovorum ◽

Promoter Trap ◽

Carotovorum Subsp ◽

Rot Disease

Pectobacterium carotovorum subsp. carotovorum strain PccS1, a bacterial pathogen causing soft rot disease of Zantedeschia elliotiana (colored calla), was investigated for virulence genes induced by the host plant. Using a promoter-trap transposon (mariner), we obtained 500 transposon mutants showing kanamycin resistance dependent on extract of Z. elliotiana. One of these mutants, PM86, exhibited attenuated virulence on both Z. elliotiana and Brassica rapa subsp. pekinensis. The growth of PM86 was also reduced in minimal medium (MM), and the reduction was restored by adding plant extract to the MM. The gene containing the insertion site was identified as rplY. The deletion mutant ΔrplY, exhibited reduced virulence, motility and plant cell wall-degrading enzyme production but not biofilm formation. Analysis of gene expression and reporter fusions revealed that the rplY gene in PccS1 is up-regulated at both the transcriptional and the translational levels in the presence of plant extract. Our results suggest that rplY is induced by Z. elliotiana extract and is crucial for virulence in P. carotovorum subsp. carotovorum.

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Effect of some chemicals on incidence of poato soft rot disease in Bangladesh

Bangladesh Journal of Scientific and Industrial Research ◽

10.3329/bjsir.v52i2.32936 ◽

2017 ◽

Vol 52 (2) ◽

pp. 135-140 ◽

Cited By ~ 1

Author(s):

MM Rahman ◽

AA Khan ◽

IH Mian ◽

AM Akanda ◽

MZ Alam

Keyword(s):

Acetic Acid ◽

Boric Acid ◽

Soft Rot ◽

Pectobacterium Carotovorum ◽

Potato Tubers ◽

Soft Rot Disease ◽

Bleaching Powder ◽

Carotovorum Subsp ◽

Rot Disease

Bactericidal effect was investigated by chemicals against potato soft rot bacteria in vitro and in storage. The chemicals were acetic acid, boric acid, bleaching powder, lactic acid, calcium hydroxide, calcium chloride, potassium chloride and sodium hypo-chloride. Among eight chemicals only three chemicals viz. acetic acid, boric acid and bleaching powder showed bactericidal activity against potato soft rot bacteria Pectobacterium carotovorum subsp. carotovorum (E. carotovora subsp. carotovora) P-138 in vitro. Based on the results of in vitro experiment three chemicals, acetic acid, boric acid and bleaching powder were used to control soft rot disease of potato in storage. Fresh potato tubers were dipped in 0.2% solution/suspensions of acetic acid, boric acid and bleaching powder for 30 min. Then soft rot bacteria Pectobacterium carotovorum subsp. carotovorum P-138 was inoculated on potato. Finally potatoes were stored for 22 weeks in net bags in sterilized condition. All the three chemicals significantly decreased the infection rate, loss in weight and increased percentage of disease reduction (PDR) of potato. Boric acid was the most effective in controlling the soft rot disease of potato in storage followed by acetic acid and bleaching powder. So these chemicals may be used for seed purpose storage of potato tubers for year round storage at farmers level.Bangladesh J. Sci. Ind. Res. 52(2), 135-140, 2017

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A novel subspecies-specific primer targeting the gyrase B gene for the detection of Pectobacterium carotovorum subsp. brasiliense

Biodiversitas Journal of Biological Diversity ◽

10.13057/biodiv/d201037 ◽

2019 ◽

Vol 20 (10) ◽

Author(s):

Tri Joko ◽

ALAN SOFFAN ◽

MUHAMMAD SAIFUR ROHMAN

Keyword(s):

Sequence Data ◽

Soft Rot ◽

Pectobacterium Carotovorum ◽

Specific Primer ◽

Base Pairs ◽

Detection And Identification ◽

Wide Range ◽

Carotovorum Subsp ◽

Rot Disease ◽

The Tropics

Abstract. Joko T, Soffan A, Muhammad Saifur Rohman MS. 2019. A novel subspecies-specific primer targeting the gyrase B gene for the detection of Pectobacterium carotovorum subsp. brasiliense. Biodiversitas 20: 3042-3048. Pectobacterium carotovorum subsp. brasiliense is one of the major causative bacterial pathogens of the soft rot disease in various crops. It has a high virulence and a wide range of hosts in the tropics and the subtropics. Most often, conventional methods are not able to accurately distinguish P. carotovorum subsp. brasiliense from other subspecies. Thus, this study aimed to design a specific gyrase B gene (gyrB) -based primers for the detection and identification of soft rot pathogen. The specific primers design was based on the alignment using gyrB gene sequence data from P. carotovorum subsp. brasiliense and other data from the GenBank. The primers comprised of F-gyr-Pcb (5’-CAC AGG CAC CGC TGG CTG TT-3’) and R-gyr-Pcb (5’-CGT CGT TCC ACT GCA ATG CCA-3’) with an amplicon of 336 base pairs. The specificity of the primers pair was verified both in silico and in polymerase chain reaction (PCR) assays, where the primers could only detect P. carotovorum subsp. brasiliense. Primers’ sensitivity was determined by qualitative PCR with a detection limit of less than 0.5 ng/µL of genomic DNA. Hence, the proposed detection tool can be beneficial to advance further studies on the ecology and epidemiology of soft rot diseases.

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First Report of Bacterial Soft Rot of Potato Caused by Pectobacterium carotovorum subsp. carotovorum in Guangdong Province of China

Plant Disease ◽

10.1094/pdis-07-12-0620-pdn ◽

2012 ◽

Vol 96 (12) ◽

pp. 1819-1819 ◽

Cited By ~ 1

Author(s):

J. X. Zhang ◽

B. R. Lin ◽

H. F. Shen ◽

X. M. Pu ◽

Z. N. Chen ◽

...

Keyword(s):

Guangdong Province ◽

Housekeeping Genes ◽

Soft Rot ◽

Bacterial Suspension ◽

Pectobacterium Carotovorum ◽

Bacterial Soft Rot ◽

First Report ◽

Wide Range ◽

Carotovorum Subsp ◽

Agar Media

Potato (Solanum tuberosum L.) is a major crop in China, with 80.0 million tons being produced in 2010 on 3.3 million ha. Pectobacterium carotovorum subsp. carotovorum Jones 1901; Hauben et al. 1999 causes soft rot worldwide on a wide range of hosts including potato, carrot, and cabbage. During spring 2010, a soft rot with a foul smell was noted in stored potato tubers of different cultivars in the Guangdong Province. Symptoms on tubers appeared as tan, water-soaked areas with watery ooze. The rotted tissues were white to cream colored. Stems of infected plants with typical inky black symptoms could also be found in the fields prior to harvest. Three different potato fields were surveyed, and 13% of the plants had the symptoms. Twenty-seven samples (three symptomatic tubers per sample) were collected. Bacteria were successfully isolated from all diseased tissues on nutrient agar media supplemented with 5% sucrose and incubated at 26 ± 1°C for 36 h. After purification on tripticase soy agar media, four typical strains (7-3-1, 7-3-2, 8-3-1, and 8-3-2) were identified using the following deterministic tests: gram-negative rods, oxidase negative, facultatively anaerobic, able to degrade pectate, sensitive to erythromycin, negative for phosphatase, unable to produce acid from α-methyl-glucoside, and produced acid from trehalose. Biolog analysis (Ver 4.20.05, Hayward, CA) identified the strains as P. carotovorum subsp. carotovorum (SIM 0.808, 0.774, 0.782, and 0.786, respectively). The identity of strains 7-3-1 (GenBank Accession No. JX258132), 7-3-2 (JX258133), and 8-3-1 (JX196705) was confirmed by 16S rRNA gene sequencing (4), since they had 99% sequence identity with other P. carotovorum subsp. carotovorum strains (GenBank Accession Nos. JF926744 and JF926758) using BLASTn. Further genetic analysis of strain 8-3-1 was performed targeting informative housekeeping genes, i.e., acnA (GenBank Accession No. JX196704), gabA (JX196706), icdA (JX196707), mdh (JX196708), mtlD (JX196709), pgi (JX196710), and proA (JX196711) (2). These sequences from strain 8-3-1 were 99 to 100%, homologous to sequences of multiple strains of P. carotovorum subsp. carotovorum. Therefore, strain 8-3-1 grouped with P. carotovorum subsp. carotovorum on the phylogenetic trees (neighbor-joining method, 1,000 bootstrap values) of seven concatenated housekeeping genes when compared with 60 other strains, including Pectobacterium spp. and Dickeya spp. (3). Pathogenicity of four strains (7-3-1, 7-3-2, 8-3-1, and 8-3-2) was evaluated by depositing a bacterial suspension (106 CFU/ml) on the potato slices of cultivar ‘Favorita’ and incubating at 30 ± 1°C. Slices inoculated with just water served as non-inoculated checks. The strains caused soft rot within 72 h and the checks had no rot. Bacteria were reisolated from the slices and were shown to be identical to the original strains based on morphological, cultural, and biochemical tests. Although this pathogen has already been reported in northern China (1), to our knowledge, this is the first report of P. carotovorum subsp. carotovorum causing bacterial soft rot of potato in Guangdong Province of China. References: (1) Y. X. Fei et al. J. Hexi Univ. 26:51, 2010.(2) B. Ma et al. Phytobacteriology 97:1150, 2007. (3) S. Nabhan et al. Plant Pathol. 61:498, 2012. (4) W. G. Weisbury et al. J. Bacteriol. 173:697, 1991.

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Characterization of Pectobacterium carotovorum subsp. carotovorum causing soft-rot disease on Pinellia ternata in China

European Journal of Plant Pathology ◽

10.1007/s10658-007-9219-4 ◽

2007 ◽

Vol 120 (3) ◽

pp. 305-310 ◽

Cited By ~ 14

Author(s):

Xiu-Fang Hu ◽

Fei-Xiang Ying ◽

Yu-Bo He ◽

Yuan-Yuan Gao ◽

Hai-Min Chen ◽

...

Keyword(s):

Soft Rot ◽

Pectobacterium Carotovorum ◽

Pinellia Ternata ◽

Soft Rot Disease ◽

Carotovorum Subsp ◽

Rot Disease

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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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