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The research and development of bio-degumming technology is under a slow progress due to the shortage of proper efficient bacterial strains and processes. A degumming bacterial strain-Pectobacterium wasabiae (PW)-with broad-spectrum degumming abilities was screened out in this study. After the fermentation for 12 h, the residual gum contents of kenaf bast, ramie bast, hemp bast, flax bast, and Apocynum venetum bast were all lower than 15%. This bacterial strain could realize the simultaneous extracellular secretion of pectinase, mannase, and xylanase with the maximum enzyme activity levels of 130.25, 157.58, and 115.24 U/mL, respectively. The optimal degumming conditions of this bacterial strain were as follows: degumming time of 12 h, bath ratio of 1:10, temperature of 33 °C, and inoculum size of 2%. After the bio-degumming through this bacterial strain, the COD in wastewater was below 4000 mg/L, which was over 60% lower than that in boiling-off wastewater generated by chemical degumming. This technology achieves higher efficiency, higher quality, and lower pollution.

Pectobacterium parmentieri (former Pectobacterium wasabiae) is an emerging pectinolytic bacterial pathogen causing losses in potato (Solanum tuberosum L.) worldwide. Until now, there are no effective measures to protect crops from infections caused by P. parmentieri. Likewise, little is known about lytic bacteriophages that can infect and kill P. parmentieri, and that could be used in the (bio)control of this pathogen in agricultural applications. A novel lytic bacteriophage vB_Ppp_A38 (ϕA38), belonging to the order Caudovirales, was isolated and characterised in our previous studies. This report provides information about its complete genome sequence and accompanying phylogenomics. The genome of vB_Ppp_A38 consists of 75,764-bp-length, linear, double-stranded DNA with an average GC content of 48.7% and is predicted to have 97 open reading frames (ORFs), with an average length of 732 nucleotides. The open reading frames were classified into functional groups, including structural units, packing, DNA metabolism, regulation and additional (miscellaneous) functions. Comparative genomic analyses based on the whole-genome sequence indicated that the phage vB_Ppp_A38 is most similar to N4-like viruses of the family Schitoviridae, genus Cbunavirus, sharing more than 90% identity with the phages vB_PatP_CB1, vB_PatP_CB3 and vB_PatP_CB4 infecting P. atrosepticum. The bacteriophage vB_Ppp_A38 is the first Cbunavirus bacteriophage infecting other Pectobacterium spp. hosts than P. atrosepticum. As vB_Ppp_A38 is a lytic virus able to kill plant-pathogenic P. parmentieri, it can be used in the biological control of this pathogen in agricultural applications.

A BIO-PCR was developed for the detection of potato blackleg pathogens, including Pectobacterium wasabiae (Pw), P. atrosepticum (Pa), P. carotovorum subsp. brasiliense (Pcb), and Dickeya dianthicola (Ddi). The method was based on the semi-selective incubation of pathogens using pectate-based liquid enrichment medium (LEM) and PCR detection of blackleg pathogens (triplex PCR for Pw, Pa, and Ddi and simplex PCR for Pcb with previously reported primers). Incubation in LEM for 24–120 h at 25 °C was effective in detecting the pathogens from plants (stems and tubers) and the soil around the roots. The detection limit by BIO-PCR from the tuber sample (0.6 g) in 10 ml medium was 2–8 colony-forming units (cfu) for four pathogens, whereas that from soil (10 g) in 100 ml medium was 2 × 102, 3 × 103, 6 × 101, and 2 × 101 cfu for Pw, Pa, Pcb, and Ddi, respectively. All four blackleg pathogens could also be detected from symptomatic stems, progeny tubers, and the soil around the stem base of diseased plants by BIO-PCR. This method is a promising technique for the efficient detection of potato blackleg pathogens not only for seed inspection but also for epidemiological research.

We determined the species composition of 219 isolates of potato blackleg or soft rot pathogens in Japan collected after 2000 from potato plants by species-specific PCR. Of the total, Pectobacterium wasabiae (Pw) and P. carotovorum subsp. brasiliense (Pcb) comprised 88% (Pw: 94 isolates, Pcb: 98 isolates) of the isolates and Dickeya dianthicola (Ddi) 12% (27 isolates). P. atrosepticum (Pa) was not detected. Genetic diversity of Pw, Pcb and Ddi was investigated using repetitive sequence-based PCR (rep-PCR) fingerprinting with three primer pairs complementary to repetitive sequences in the bacterial genome (BOX, ERIC and REP). One hundred and twenty-one Pw isolates, one hundred Pcb isolates and thirty-three Ddi isolates were assigned to twenty-nine, twenty-one and eleven fingerprint genotypes (FGs) based on the combination of fingerprint patterns for each rep-PCR experiment (BOX-, ERIC- and REP-PCR). The genetic diversity of Pw and Ddi isolates in Japan was considered high, with Simpson’s diversity index and Shannon’s index of 0.87 and 2.44 for Pw, and 0.84 and 2.05 for Ddi, respectively. Pcb isolates were less diversified genetically (diversity index: 0.57 and 1.58). The FGs of the isolates obtained from blackleg-affected potato plants in the seed tuber field agreed with those of the isolates in the field where the seeds produced were grown. These results support the occurrence of seed-borne transmission of the blackleg pathogen between the fields.

Pectobacterium parmentieri (formerly Pectobacterium wasabiae), which causes soft rot disease in potatoes, is a newly established species of pectinolytic bacteria within the family Pectobacteriaceae. Despite serious damage caused to the potato industry worldwide, no field-deployable diagnostic tests are available to detect the pathogen in plant samples. In this study, we aimed to develop a reliable, rapid, field-deployable loop-mediated isothermal amplification (LAMP) assay for the specific detection of P. parmentieri. Specific LAMP primers targeting the petF1 gene region, found in P. parmentieri but no other Pectobacterium spp., were designed and validated in silico and in vitro using extensive inclusivity (15 strains of P. parmentieri) and exclusivity (94 strains including all other species in the genus Pectobacterium and host DNA) panels. No false positives or negatives were detected when the assay was tested directly with bacterial colonies, and with infected plant and soil samples. Sensitivity (analytical) assays using serially diluted bacterial cell lysate and purified genomic DNA established the detection limit at 10 CFU/mL and 100 fg (18–20 genome copies), respectively, even in the presence of host crude DNA. Consistent results obtained by multiple users/operators and field tests suggest the assay’s applicability to routine diagnostics, seed certification programs, biosecurity, and epidemiological studies.

Pectobacterium parmentieri is a Gram-negative plant-pathogenic bacterium able to infect potato (Solanum tuberosum L.). Little is known about lytic bacteriophages infecting P. parmentieri and how phage-resistance influences the environmental fitness and virulence of this species. A lytic phage vB_Ppp_A38 (ϕA38) has been previously isolated and characterized as a potential biological control agent for the management of P. parmentieri. In this study, seven P. parmentieri SCC 3193 Tn5 mutants were identified that exhibited resistance to infection caused by vB_Ppp_A38 (ϕA38). The genes disrupted in these seven mutants encoded proteins involved in the assembly of O-antigen, sugar metabolism, and the production of bacterial capsule exopolysaccharides. The potential of A38-resistant P. parmentieri mutants for plant colonization and pathogenicity as well as other phenotypes expected to contribute to the ecological fitness of P. parmentieri, including growth rate, use of carbon and nitrogen sources, production of pectinolytic enzymes, proteases, cellulases, and siderophores, swimming and swarming motility, presence of capsule and flagella as well as the ability to form biofilm were assessed. Compared to the wild-type P. parmentieri strain, all phage-resistant mutants exhibited a reduced ability to colonize and to cause symptoms in growing potato (S. tuberosum L.) plants. The implications of bacteriophage resistance on the ecological fitness of P. parmentieri are discussed.

Potato (Solanum tuberosum) is the second most important crop after maize in Kenya and plays a major role in national food production and security. Despite its importance, production is constrained by a number of factors, including diseases. Diseases caused by bacteria are a major biotic production constraint. Soft rot and blackleg diseases are considered as important constraints causing considerable losses in many production systems. However, there is scanty information on soft rot and blackleg in potato production systems in Kenya. A limited survey was conducted between September and October 2016 and in December 2016 to January 2017 in four major potato growing regions to determine the identity of pectinolytic bacteria causing blackleg and soft rot disease of potato in Kenya. Tubers and stems with symptoms of blackleg and soft rot disease were collected from 91 randomly selected farms. Identification of the pathogens was done using phenotypic methods which involved culturing on semi-selective medium, biochemical tests, pathogenicity tests and DNA based methods. The results identified occurrence of Pectobacterium carotovorum subsp. carotovorum (6 isolates) in all four regions and Pectobacterium wasabiae (5 isolates) for the first time in Elgeiyo Marakwet. The identity of twelve isolates could not be specified and therefore needs further investigation.

Pectobacterium wasabiae is a dreadful causal agent of potato soft rot. It relied mainly on the production a wide range of plant cell wall-degrading enzymes to cause disease. Of the different enzymes, the endo-b-1,4-glucanase may be important in pathogenesis, being responsible for hydrolyzing the cellulose. In the process of pathogen infection, cellulase secreted by the pathogen is major determinant of pathogenicity, which played an important role in softening and decomposition of cell wall. But so far, the related research very rarely. However, most characteristic of the cellulase in the soft rot bacteria was not clear so far. Here, a putative endo-b-1,4-glucanase gene from P. wasabiae SCC3193 was previously synthesized according the bias codon of yeast, heteroexpressed in Pichia pastoris, and further investigated for its characteristic. Pectobacterium wasabiae SCC3193 is a genetically well characterized model in soft rot research. SDS-PAGE analysis showed that the recombinant endo-b-1,4-glucanase (PcegAI) produced in P. pastoris has a molecular mass of approximately 30kDa. When CMC was used as substrate, it exhibited maximal activity at pH 4.0 with an optimum temperature of 40°C, and stable at pH 5-8. The Km and specific activity values for CMC are 64.5 mM and 7 ×103 U mg-1 respectively. Most metallic ions had no influence on the activity of endo-b-1,4-glucanase at a concentration of 1 mM except for Cu2+ and Mn2+. These characteristics suggested that PcegAI may play a major role in degrading of cellulose polymers. The high activity of this enzyme also has potential in biomass opening, such as the bioconversion of lignocellulosic materials and bioethanol production.

Pectobacterium parmentieri (formerly Pectobacterium wasabiae) is a newly established species of pectinolytic plant-pathogenic bacteria responsible for the symptoms of soft rot and blackleg on potato. In this work, we describe biodiversity and the population structure of P. parmentieri strains isolated during two consecutive growing seasons from the seed potato fields in Poland. About 450 samples of diseased potato tubers, potato plants, or accompanying weeds were collected throughout the country and tested for the presence of P. parmentieri by molecular identification methods. We found that P. parmentieri strains commonly occur in almost all regions of Poland. Furthermore, these isolates constituted significant fraction of pectinolytic bacteria from seed potato fields because 16% (2013) and 13% (2014) of the analyzed plant samples were infected with P. parmentieri. Subsequently, a detailed characterization of the obtained strains was conducted basing on repetitive sequences profiling, recA-gene-based phylogeny, and phenotypic features. By applying repetitive extragenic palindromic sequence-based polymerase chain reaction (REP-PCR), we revealed the presence of five distinct genomic profiles among P. parmentieri strains, with profile I being the most abundant (approximately 44%). The performed recA gene-based phylogenetic analysis divided P. parmentieri isolates into two distinct clades, although the strains originating from different years did not group separately. Evaluation of the phenotypic traits playing crucial roles for the virulence of pectinolytic bacteria (namely, pectinase, cellulase and protease activities, and siderophore production, in addition to potato tissue maceration, swimming, and swarming motility) indicated some differences among the characterized strains. To the best of our knowledge, this is the first study that describes biodiversity and the population structure of P. parmentieri isolated in two growing seasons under temperate climate conditions and, hence, illustrates high heterogeneity within this species.

Cabbage (Brassica oleracea var. capitata) is one of the important vegetables in Japan. In the summer of 2019, some cabbages with soft rot were found in commercial fields in Hokkaido, the northern island in Japan. All diseased plants showed grey to brown discoloration and expanding water-soaked lesions on leaves. We obtained two independent strains (NACAB191 and NACAB192) from diseased leaves. DNA from these strains yielded an expected single size amplicon with the primer set of PhF/PhR for P. wasabiae (De Boer et al. 2012) by PCR, but did not yield the expected amplicon with the primer set of BR1f/L1r for P. carotovorum subsp. brasiliense (Duarte et al. 2004) and Eca1f/Eca2r for P. atrosepticum (De Boer et al., 1995) by PCR. These two strains grew at 37°C, and their ability to utilize raffinose and lactose. These bacterial strains were gram-negative and rod-shaped. The bacterium was positive for O-nitrophenyl-beta-D-galactopyranoside, N-acetylglucosaminyl transferase, gelatin liquefaction, and acid production from D-galactose, lactose, melibiose, raffinose, citrate, and trehalose. The bacterium was negative for indole production and acid production from maltose, α-methyl-D-glucoside, sorbitol, D-arabitol, inositol, inulin, and melezitose. All strains exhibited pectolytic activity on potato slices. The sequence analysis of 16S rDNA (LC597897 and LC597898) showed more than 98% identities to P. wasabiae strain (e.g. HAFL01 in Switzerland) by BLAST analysis. In addition, Multi-locus sequence analysis (Ma et al. 2007) was performed by MEGA10 (Kumer et al. 2018) using concatenated DNA sequences of seven housekeeping genes (aconitate hydratase(acnA, LC597923 and LC597924), glyceraldehyde-3-phosphate dehydrogenase A(gapA, LC597970 and LC597971), isocitrate dehydrogenase (icdA, LC597996 and LC597997), malate dehydrogenase(mdh, LC598022 and LC598023), mannitol-1-phosphate dehydrogenase (mtlD, LC598048 and LC598049), glucose-6-phosphate isomerase (pgi, LC598074 and LC598075) and gamma-glutamyl phospate reductase (proA, LC598079 and LC598080)), and all clustered NACAB191 and NACAB192 into a clade containing other confirmed strains of P. wasabiae. As a result, these two strains shared high identity with each other (>98%, E-Values showed 0). The clade containing these two strains was consistently placed in a larger clade with the other P. wasabiae and 100% bootstrap support for its separation from other Pectobacterium species available in GenBank when the consensus tree constructed using Maximum Likelihood method. Pathogenicity of these strains against cabbage (cv. 'Rakuen') was confirmed by the field experiments with five weeks growth plants sprayed with bacterial suspension (1×107cfu/ml). Thirty cabbages per strain were used in this study, 12 plants treated the suspension of NACAB191 and 16 plants treated the suspension of NACAB192 which died with the same soft rot symptoms about four weeks after inoculation. Whereas water-inoculated plants remained symptomless. Strains re-isolated from the artificially diseased stems were confirmed as P. wasabiae using the methods as biochemical characterization and multiple genetic analyses. Based on the disease symptoms, the cultural, molecular, and pathological features of the strains, we conclude that the soft rot symptoms of cabbage in Hokkaido in 2019 were caused by P. wasabiae. To our knowledge, this is the first report of P. wasabiae as the soft rot disease agent of cabbage in Japan.

Two bacteriophages, ϕA38 and ϕA41, infecting Pectobacterium parmentieri strain SCC 3193 (former Pectobacterium wasabiae strain SCC 3193) were isolated from arable soil samples collected in different regions of Poland. ϕA38 and ϕA41 have a typical morphology of the members of the family Podoviride and order Caudovirales, with a head diameter of ca. 60 nm and tail length of ca. 20 nm. Phages ϕA38 and ϕA41 exhibited a similar RFLP pattern with Csp6I restriction endonuclease. They were stable in a range of pHs, temperatures and osmolarities but were rapidly inactivated by UV light. During the first 20 min., 74 and 69% of ϕA38 and ϕA41 phages, respectively, were adsorbed to SCC 3193 cells. In one-step growth experiments, ϕA38 and ϕA41 showed latent period of ca. 20–30 min and burst size of 102 and 141 phages, respectively. The optimal multiplicity of infection (MOI) was calculated to be 0.01 for both bacteriophages. In the host range experiments, both phages were able to infect six from 21 of the tested P. parmentieri isolates but the phages were unable to infect other members of the Pectobacterium spp. or Dickeya spp. In the proof-of-concept experiments, ϕA38 and ϕA41 were able to inhibit the growth of P. parmentieri strain SCC 3193 and to protect potato tuber tissue maceration caused by the bacterium. The potential use of ϕA38 and ϕA41 bacteriophages for the biocontrol of P. parmentieri in potato is discussed.

Pectobacterium atroseptica is known as a rod-shaped gram-negative bacterial pathogen associated with the blackleg of potato. P. atroseptica has been widely identified as the predominant agent causing tuber rot in temperate regions, a disease that leads to severe economic losses to potato industry. In this study, we provide the complete genome sequence of P. atroseptica JG10-08, which revealed that P. atroseptica strain JG10-08 carries a single 5,004,926 bp chromosome with 51.15% G+C content and harbors 4252 predicted coding genes. Phylogenetic analysis based on the genome sequences showed a close evolutionary relationship between P. atroseptica and Pectobacterium wasabiae. We discovered total 168 genes were potentially related to pathogenesis including 9 strain-specific genes encoding toxins on the genome of JG10-08. Further comparison with other species in Pectobacterium revealed a better understanding of pathogenic factors, especially secretion systems in P. atroseptica JG10-08. Collectively, the results of this research provide a solid foundation for discovering the underlying pathogenic mechanisms of P. atroseptica and offer the information to develop more effective strategies against blackleg of potatoes.

Gram-stain-negative, rod-shaped pectinolytic bacteria strains designated as DPMP315T, DPMP316, DPMP317 and DPMP318 isolated from groundwater sampled from a vegetable field in the North of Poland, were subjected to the polyphasic analyses. Multilocus sequence analyses based on five housekeeping genes (gyrA, recA, recN, rpoA and rpoS) revealed their distinctiveness from the other species of the genus, simultaneously indicating that the newly described species, Pectobacterium punjabense , as well as Pectobacterium parmentieri and P. wasabiae , to be the closest relatives. In silico DNA–DNA hybridization (

Aerial stem rot of potato (Solanum tuberosum), also known as bacterial stem rot, is often caused by the pectolytic bacteria Pectobacterium (Erwinia) carotovorum subsp. carotovorum, P. atrosepticum, or Dickeya spp. (3). A survey was carried out in August 2008 in ‘Russet Burbank’ potato fields exhibiting aerial stem rot symptoms in the Columbia Basin of Washington State. One bacterial strain isolated during the survey, PwO405, exhibited pectolytic ability on crystal violet pectate (CVP) agar and potato slices and failed to grow at 37°C, but physiological tests did not conclusively distinguish the bacterium as P. atrosepticum (1). The bacterium was positive for ONPG, N-acetylglucosaminyl transferase, gelatin liquefaction, and acid production from D-galactose, lactose, melibiose, raffinose, citrate, and trehalose. The bacterium was negative for indole production and acid production from maltose, α-methyl-D-glucoside, sorbitol, D-arabitol, inositol, inulin, and melezitose. Molecular identification of the bacterium was performed with 16S rRNA, aconitase (acnA), and malate dehydrogenase (mdh) coding sequences as previously described (2,4). Partial sequences of 16S rRNA (1,408 bp) and acnA (412 bp) genes (GenBank Accession Nos. JQ723958 and JQ723959, respectively) exhibited 99% shared identities with P. wasabiae strain WPP163, while the mdh sequence (435 bp) (GenBank Accession No. JQ723960) exhibited 100% shared identity with mdh sequences from three P. wasabiae strains (NZEC9, NZEC10, and NZEC8974). Maximum parsimony analysis using concatenated acnA and mdh sequences from this study and Pectobacterium sequences previously deposited in GenBank (2,4) clustered strain PwO405 with other P. wasabiae strains. Three 7-week-old ‘Russet Norkotah’ potato plants were wound-inoculated by inserting a sterile 23 gauge needle just above a central leaf axil at a depth of 1 mm. A 10-μl drop of inoculum (104 CFU) was placed on the wound. Plants were exposed to a 24-h leaf wetness period (90 to 100% RH in a mist chamber) and lesions were measured. All three inoculated plants exhibited aerial stem rot symptoms similar to those observed in the field, including brown water-soaked lesions that spread acropetally and basipetally. Upon drying, the lesions became shriveled and turned dark brown to black. Some plants exhibited hollowing of the stems and unilateral wilt on the side of the lesion. Symptoms were not observed on water-inoculated controls. The bacteria that were reisolated into pure culture from all three inoculated stems caused pitting on CVP and exhibited the same morphology as the original culture and were confirmed as P. wasabiae using 16S rRNA, acnA, and mdh coding sequences, fulfilling Koch's postulates. Stem rot ability of the bacterium was also confirmed on four potato cultivars: ‘Ranger Russet,’ ‘Russet Burbank,’ ‘Russet Norkotah,’ and ‘Umatilla Russet’ by wound-inoculating six single-stem plants of each cultivar as described above. To our knowledge, this is the first report of aerial stem rot of potato caused by P. wasabiae in Washington State. References: (1) S. De Boer and A. Kelman. Page 56 in: Laboratory Guide for Identification of Plant Pathogenic Bacteria, 3rd ed. N. Schaad et al., ed. APS Press, St. Paul, 2001. (2) A. Pitman et al. Eur. J. Plant Pathol. 32:211, 2010. (3) M. Powelson and G. Franc. Page 10 in: Compendium of Potato Diseases. W. Stevenson et al., ed. APS Press, St. Paul, 2002. (4) M. Yap et al. Appl. Environ. Microbiol. 70:3013, 2004.

In August 2011, sweet potato (Ipomoea batatas), tomato (Solanum lycopersicum), and eggplant (S. melongena) crops from major growing areas of the Cameron highlands and Johor state in Malaysia were affected by a soft rot disease. Disease incidence exceeded 80, 75, and 65% in severely infected fields and greenhouses of sweet potato, tomato, and eggplant, respectively. The disease was characterized by dark and small water-soaked lesions or soft rot symptoms on sweet potato tubers, tomato stems, and eggplant fruits. In addition, extensive discoloration of vascular tissues, stem hollowness, and water-soaked, soft, dark green lesions that turned brown with age were observed on the stem of tomato and eggplant. A survey was performed in these growing areas and 22 isolates of the pathogen were obtained from sweet potato (12 isolates), tomato (6 isolates), and eggplant (4 isolates) on nutrient agar (NA) and eosin methylene blue (EMB) (4). The cultures were incubated at 27°C for 2 days and colonies that were emerald green on EMB or white to gray on NA were selected for further studies. All bacterial cultures isolated from the survey exhibited pectolytic ability on potato slices. These bacterial isolates were gram negative; rod shaped; N-acetylglucosaminyl transferase, gelatin liquefaction, and OPNG positive; and were also positive for acid production from D-galactose, lactosemelibiose, raffinose, citrate, and trehalose. They were negative for indol production, phosphatase activity, reducing substances from sucrose, and negative for acid production from maltose, sorbitol, inositol, inolin, melezitose, α-mathyl-D-glocoside, and D-arabitol. The bacteria did not grow on NA at 37°C. Based on these biochemical and morphological assays, the pathogen was identified as Pectobacterium wasabiae (2). In addition, DNA was extracted and PCR assay with two primers (16SF1 and 16SR1) was performed (4). Partial sequences of 16S rRNA (GenBank Accession Nos. JQ665714, JX494234, and JX513960) of sweet potato, tomato, and eggplant, respectively, exhibited a 99% identity with P. wasabiae strain SR91 (NR_026047 and NR_026047.1). A pathogenicity assay was carried out on sweet potato tubers (cv. Oren), tomato stems (cv. 152177-A), and eggplant fruits (cv. 125066x) with 4 randomly representative isolates obtained from each crop. Sweet potato tubers, tomato stems, and eggplant fruits (4 replications) were sanitized in 70% ethyl alcohol for 30 s, washed and rinsed in sterile distilled water, and needle punctured with a bacterial suspension at a concentration of 108 CFU/ml. Inoculated tubers, stems, and fruits were incubated in a moist chamber at 90 to 100% RH for 72 h at 25°C when lesions were measured. All inoculated tubers, stems, and fruits exhibited soft rot symptoms after 72 h similar to those observed in the fields and greenhouses and the same bacteria were consistently reisolated. Symptoms were not observed on controls. The pathogenicty test was repeated with similar results. P. wasabiae have been previously reported to cause soft rot on Japanese horseradish (3), and aerial stem rot on potato in New Zealand (4), the U.S. (2), and Iran (1). To our knowledge, this is the first report of sweet potato, tomato, and eggplant soft rot caused by P. wasabiae in Malaysia. References: (1) S. Baghaee-Ravari et al. Eur. J. Plant Pathol. 129:413, 2011. (2) S. De Boer and A. Kelman. Page 56 in: Laboratory Guide for Identification of Plant Pathogenic Bacteria, 3rd ed. N. Schaad et al., eds. APS Press, St. Paul, 2001. (3) M. Goto et al. Int. J. Syst. Bacteriol. 37:130, 1987. (4) A. R. Pitman et al. Eur. J. Plant Pathol. 126:423, 2010.

In western Europe, Pectobacterium carotovorum subsp. brasiliense is emerging as a causal agent of blackleg disease. In field experiments in the Netherlands, the virulence of this pathogen was compared with strains of other Dickeya and Pectobacterium species. In 2013 and 2014, seed potato tubers were vacuum infiltrated with high densities of bacteria (106 CFU mL−1) and planted in clay soil. Inoculation with P. carotovorum subsp. brasiliense and P. atrosepticum resulted in high disease incidences (75–95%), inoculation with D. solani and P. wasabiae led to incidences between 5% and 25%, but no significant disease development was observed in treatments with P. carotovorum subsp. carotovorum, D. dianthicola or the water control. Co-inoculations of seed potatoes with P. carotovorum subsp. brasiliense and D. solani gave a similar disease incidence to inoculation with only P. carotovorum subsp. brasiliense. However, co-inoculation of P. carotovorum subsp. brasiliense with P. wasabiae resulted in a decrease in disease incidence compared to inoculation with only P. carotovorum subsp. brasiliense. In 2015, seed potatoes were inoculated with increasing densities of P. carotovorum subsp. brasiliense, D. solani or P. atrosepticum (103–106 CFU mL−1). After vacuum infiltration, even a low inoculum density resulted in high disease incidence. However, immersion without vacuum caused disease only at high bacterial densities. Specific TaqMan assays were evaluated and developed for detection of P. carotovorum subsp. brasiliense, P. wasabiae and P. atrosepticum and confirmed the presence of these pathogens in progeny tubers of plants derived from vacuum-infiltrated seed tubers.

In plant soft-rotting bacteria Pectobacterium, quorum sensing (QS) regulates the secretion of an arsenal of plant cell wall degrading extracellular enzymes (PCWDEs) and flagella-mediated motility via two different signaling molecules such as 3-oxohexanoyl-L-homoserine lactone (3-oxo-C6-AHL) and 3-oxooctanoyl-L-homoserine lactone (3-oxo- C8-AHL). In the present investigation, the phytochemical compound curcumin was assessed for its QS inhibitory potential against AHL-dependent PCWDEs production and motility in P. wasabiae SCC3193, P. carotovorum subsp. Carotovorum Pcc21 and P. carotovorum subsp. Carotovorum Pcc. Interestingly, curcumin at sub-MIC effectively inhibited the production of PCWDEs as well as the swimming and swarming motility of all tested pathogens in a non-bactericidal fashion. Subsequently, the in vitro root pathogenicity assay in Arabidopsis thaliana unveiled the rescuing efficacy of curcumin against the pathogenicity of tested plant pathogens by attenuating its QS mediated virulence factors production. Besides, studies with QS mutant strain Pcc21-M10 also evidenced the QSI property of curcumin.

In August 2012, vegetable crops including potato, cabbage, bell pepper and carrot showing symptoms of maceration and water soaked lesions on their tuber, leaf and fruit were collected from major vegetable growing areas in north-west of Iran. Physiological and biochemical assays divided the isolates into two main groups according to their ability to grow at 37°C. In addition, these two groups were differentiated by some biochemical characteristics such as utilization of α –methyl glucoside and maltose, reducing substrates from sucrose and gelatin liquefaction. Partial sequence of polymerase chain reaction (PCR) product from reaction of putative Pectobacterium spp. with 16S rRNA confirmed the results obtained from physiological and biochemical assays used for identification of the bacterium. All of the causal organisms isolated from infected tissues were identified as Pectobacterium spp. based on biochemical characteristics and PCR amplification of the 16S rRNA gene with specific primers PCCSSF and PCCSSR. Twenty of the isolates were identified as Pectobacterium carotovorum subsp. carotovorum using a polyphasic approach including physiological, biochemical and molecular characteristics. Only six isolates from infected bell peppers and five isolates from infected potatoes identified as Pectobacterium wasabiae and Pectobacterium atrosepticum respectively. Our findings based on physiological, biochemical and molecular assays indicated the occurrence of P. wasabiae as a novel species in bell pepper in Iran. Moreover, genetic diversity of Pcc strains in this study was also performed based on Repetitive extragenic palindromic-PCR (rep-PCR). High genetic variability was revealed among these Pcc strains by rep-PCR typing using two primers sets for ERIC-PCR and BOX-PCR. Key words: 16S rRNA, repetitive extragenic palindromic- polymerase chain reaction (rep-PCR), Pectobacterium, soft rot, vegetable.

International audience; Pectobacterium isolates SS95(T), SS54 and SS56 were collected from a potato field in the Chiniot district in the plains of the Punjab province, Pakistan. Sequencing of the gapA barcode revealed that these strains belong to a novel phylogenetic group separated from P. ectobacterium wasabiae and Pectobacterium parmentieri species. Furthermore, multilocus sequence analyses of 13 housekeeping genes (fusA, rpoD, acnA, purA, gyrB, recA, mdh, mtlD, groEL, secY, glyA, gapA and rplB) clearly distinguished the type strain, SS95(T), from its closest relatives, i.e.P. parmentieri RNS 08-42-1A(T) and P. wasabiae CFBP3304(T), as well as from all the other known Pectobacterium species. In silico DNA-DNA hybridization (\textless44.1 %) and average nucleotide identity (\textless90.75 %) values of strain SS95(T) compared with other Pectobacterium type strains supported the delineation of a new species. Genomic and phenotypic comparisons permitted the identification of additional traits that distinguished the Pakistani isolates from all other known Pectobacterium type strains. The name Pectobacterium punjabense sp. nov. is proposed for this taxon with the type strain SS95(T) (=CFBP 8604(T) =LMG 30622(T)).

Four Gram-negative, rod-shaped pectinolytic bacterial strains designated as 2M, 9M, DPMP599 and DPMP600 were subjected to polyphasic analyses that revealed their distinctiveness from the other Pectobacterium species. Strains 2M and 9M were isolated from Calla lily bulbs cultivated in Central Poland. DPMP599 and DPMP600 strains were isolated from Calla lily leaves from plants grown in Serbia. Phylogenetic analyses based on nine housekeeping genes (gapA, gyrA, icdA, pgi, proA, recA, recN, rpoA, and rpoS), as well as phylogeny based on the 381 most conserved universal proteins confirmed that Pectobacterium zantedeschiae strains were distantly related to the other Pectobacterium, and indicated Pectobacterium atrosepticum, Pectobacterium betavasculorum, Pectobacterium parmentieri and Pectobacterium wasabiae as the closest relatives. Moreover, the analysis revealed that Pectobacterium zantedeschiae strains are not akin to Pectobacterium aroidearum strains, which were likewise isolated from Calla lily. The genome sequencing of the strains 2M, 9M and DPMP600 and their comparison with whole genome sequences of other Pectobacterium type strains confirmed their distinctiveness and separate species status within the genus based on parameters of in silico DNA–DNA hybridization and average nucleotide identity (ANI) values. The MALDI-TOF MS proteomic profile supported the proposition of delineation of the P. zantedeschiae and additionally confirmed the individuality of the studied strains. Based on of all of these data, it is proposed that the strains 2M, 9M, DPMP599, and DPMP600 isolated from Calla lily, previously assigned as P. atrosepticum should be reclassified as Pectobacterium zantedeschiae sp. nov. with the strain 9M T (PCM2893 = DSM105717 = IFB9009) as the type strain.

Potato blackleg and soft rot cause major losses and are caused by two bacterial genera, Pectobacterium and Dickeya. Species affecting potatoes are Pectobacterium atrosepticum (Pba), Pectobacterium carotovorum subsp. carotovorum (Pcc), Pectobacterium carotovorum subsp. brasiliense (Pcb), Pectobacterium wasabiae (Pwa), Dickeya dadantii (Dda) and Dickeya solani (Dso). Pathogenicity of these species is dependent on temperature, with each species having its own optimal temperature and temperature range for growth, leading to varying degrees of losses. Pectobacterium atrosepticum, a temperature sensitive species, mainly occurs in temperate climates, Pcc in temperate to tropical, and Dickeya spp. in subtropical environments. The aim of this study was to determine the cardinal growth temperatures for the species responsible for blackleg and soft rot in vitro. Bacterial isolates were incubated in a temperature gradient shaking incubator at 30 different temperatures ranging from ±5 °C to ±56 °C, and growth measured at two set time intervals. Results were statistically analysed using the Gaussian function. The optimal temperature of 31 °C and temperature range of 20 °C to 38 °C for Pectobacterium carotovorum subsp. brasiliense, was similar to those recorded for Pcc. Pectobacterium wasabiae grew at an optimal temperature of 29 °C and range of 20 °C to 34 °C. Higher optimal temperatures of 32 °C and 34 °C, with ranges of 21 °C to 38 °C and 23 °C to 41 °C were recorded for Dda and Dso, respectively. The minimal variation in optimal temperatures between different species might be an indication that temperature ranges, rather than optimal temperature, play an important role in disease development. Results for Dso, which has not yet been reported in South Africa, are especially important in light of prevailing temperatures in South African potato production regions.

Bacteriophage vB_PpaP_PP74 (PP74) is a novel virulent phage that infects members of the species Pectobacterium parmentieri, a newly established species of soft-rot-causing bacteria in the family Pectobacteriaceae, derived from potato-specific Pectobacterium wasabiae. vB_PpaP_PP74 was identified as a member of the family Podoviridae by transmission electron microscopy. The phage has a 39,790-bp dsDNA genome containing 50 open reading frames (ORFs). Because of the absence of genes encoding toxins or lysogeny factors, PP74 may be considered a candidate phage for pathogen biocontrol applications. The genome layout is similar to genomes of T7-like phages within the subfamily Autographivirinae, and therefore, functions can be attributed to most of ORFs. However, the closest nucleotide sequence homologs of phage PP74 are unclassified Escherichia phages. Based on phylogenetic analysis, vB_PpaP_PP74 is a sensu lato T7-like phage, but it forms a distant subgenus group together with homologous enterobacterial phages.

Seven Gram-negative, rod-shaped pectinolytic bacteria strains designated as IFB5227, IFB5228, IFB5229, IFB5230, IFB5231, IFB5232, IFB5636, isolated from potato tubers cultivated in Peru at high altitude (2400-3800m) were subjected to polyphasic analyses that revealed their distinctiveness from the other Pectobacterium species. Phylogenetic analyses based on five housekeeping genes (gyrA, recA, recN, rpoA and rpoS) clearly showed strains separateness, simultaneously indicating Pectobacterium atrosepticum, Pectobacterium wasabiae, Pectobacterium parmentieri and Pectobacterium betavasculorum as the closest relatives. In silico DNA-DNA hybridization of strain IFB5232T with other Pectobacterium type strains revealed significant drop in DDH value below 70%, which is a prerequisite to distinguish Pectobacterium peruviense. The ANI values supported the proposition of delineation of the P. peruviense. Genetic REP-PCR fingerprint and detailed MALDI-TOF MS proteomic profile sealed the individuality of the studied strains. However, phenotypic assays do not indicate immense differences. Provided results of analyses performed for seven Peruvian strains are the basis for novel species distinction and reclassification of the strains IFB5227-5232 and IFB5636, previously classified as Pectobacterium carotovorum subsp. carotovorum. Here, we propose to establish the IFB5232 isolate as a type strain (=PCM2893T=LMG30269T=SCRI179T) with the name Pectobacterium peruviense sp. nov.

Bacterial communities can sense their neighbors, regulating group behaviors in response to cell density and environmental changes. The diversity of signaling networks in a single species has been postulated to allow custom responses to different stimuli; however, little is known about how multiple signals are integrated and the implications of this integration in different ecological contexts. In the plant pathogen Pectobacterium wasabiae (formerly Erwinia carotovora), two signaling networks—the N-acyl homoserine lactone (AHL) quorum-sensing system and the Gac/Rsm signal transduction pathway—control the expression of secreted plant cell wall-degrading enzymes, its major virulence determinants. We show that the AHL system controls the Gac/Rsm system by affecting the expression of the regulatory RNA RsmB. This regulation is mediated by ExpR2, the quorum-sensing receptor that responds to the P. wasabiae cognate AHL but also to AHLs produced by other bacterial species. As a consequence, this level of regulation allows P. wasabiae to bypass the Gac-dependent regulation of RsmB in the presence of exogenous AHLs or AHL-producing bacteria. We provide in vivo evidence that this pivotal role of RsmB in signal transduction is important for the ability of P. wasabiae to induce virulence in response to other AHL-producing bacteria in multispecies plant lesions. Our results suggest that the signaling architecture in P. wasabiae was coopted to prime the bacteria to eavesdrop on other bacteria and quickly join the efforts of other species, which are already exploiting host resources., IMPORTANCE Quorum-sensing mechanisms enable bacteria to communicate through small signal molecules and coordinate group behaviors. Often, bacteria have various quorum-sensing receptors and integrate information with other signal transduction pathways, presumably allowing them to respond to different ecological contexts. The plant pathogen Pectobacterium wasabiae has two N-acyl homoserine lactone receptors with apparently the same regulatory functions. Our work revealed that the receptor with the broadest signal specificity is also responsible for establishing the link between the main signaling pathways regulating virulence in P. wasabiae. This link is essential to provide P. wasabiae with the ability to induce virulence earlier in response to higher densities of other bacterial species. We further present in vivo evidence that this novel regulatory link enables P. wasabiae to join related bacteria in the effort to degrade host tissue in multispecies plant lesions. Our work provides support for the hypothesis that interspecies interactions are among the major factors influencing the network architectures observed in bacterial quorum-sensing pathways.

International audience; Pectobacterium wasabiae was originally isolated from Japanese horseradish (Eutrema wasabi), but recently some Pectobacterium isolates collected from potato plants and tubers displaying blackleg and soft rot symptoms were also assigned to P. wasabiae. Here, combining genomic and phenotypical data, we re-evaluated their taxonomic position. PacBio and Illumina technologies were used to complete the genome sequences of P. wasabiae CFBP 3304T and RNS 08-42-1A. Multi-locus sequence analysis showed that the P. wasabiae strains RNS 08-42-1A, SCC3193, CFIA1002 and WPP163, which were collected from potato plant environment, constituted a separate clade from the original Japanese horseradish P. wasabiae. The taxonomic position of these strains was also supported by calculation of the in-silico DNA-DNA hybridization, genome average nucleotide indentity, alignment fraction and average nucleotide indentity values. In addition, they were phenotypically distinguished from P. wasabiae strains by producing acids from (+)-raffinose, α-d(+)-α-lactose, d(+)-galactose and (+)-melibiose but not from methyl α-d-glycopyranoside, (+)-maltose or malonic acid. The name Pectobacterium parmentieri sp. nov. is proposed for this taxon; the type strain is RNS 08-42-1AT (=CFBP 8475T=LMG 29774T).

The objective of this study was to develop a multiplex detection and identification protocol for bacterial soft rot coliforms, namely Pectobacterium wasabiae (Pw), Pectobacterium atrosepticum (Pba) and Dickeya spp., responsible for potato blackleg and tuber soft rot. The procedures were derived from the phylogenetic relationships of these and other Enterobacteriaceae based on recA sequences. The group of Pw strains was highly homogeneous and could be distinguished from the other species. A ligation-based method for detection of Pw was developed. Five padlock probes (PLPs) were designed, targeting recA sequences to identify the Pw, Pba or Dickeya spp., whereas a sixth probe recognised recA sequences of all soft rot coliforms including Pectobacterium carotovorum subsp. carotovorum (Pcc). Two PLP-based applications were developed: one using real-time PCR and one using universal microarrays. Assay sensitivity and specificity were demonstrated using 71 strains of Pw, Pcc, Pba and Dickeya spp. Both multiplex methods can be potentially used for seed testing and in ecological studies, but further validation is required

To identify bacteria causing soft rot and blackleg in potato in Finland, pectinolytic enterobacteria were isolated from diseased potato stems and tubers. In addition to isolates identified as Pectobacterium atrosepticum and Dickeya sp., many of the isolated strains were identified as Pectobacterium carotovorum subsp. carotovorum. Phylogenetic analysis and biochemical tests indicated that one of the isolates from potato stems resembled Pectobacterium wasabiae. Furthermore, two blackleg-causing P. carotovorum strains recently isolated in Europe clustered with P. wasabiae, suggesting that at least some of these isolates were originally misidentified. All the other Finnish P. carotovorum isolates resembled the subsp. carotovorum type strain in biochemical tests but could be clustered into two distinct groups in the phylogenetic analysis. One of the groups mainly contained strains isolated from diseased tubers, whereas the other mainly included isolates from potato stems. In contrast to the tuber isolates, the stem isolates lacked genes in Type III secretion genes, were not able to elicit a hypersensitive response in tobacco leaves and produced only small amounts of autoinducers in the stationary phase in vitro. P. wasabiae isolate was able to cause similar amount of blackleg-like symptoms as P. atrosepticum in a field experiment with vacuum-infiltrated tubers, whereas both P. atrosepticum and P. carotovorum isolates reduced emergence and delayed growth more than P. wasabiae. Our findings confirm the presence of P. wasabiae in Finland and show that the Finnish P. carotovorum subsp. carotovorum isolates can be divided into two groups with specific characteristics and possibly also different ecologies

Potato early dying (PED) is a disease complex primarily caused by the fungus Verticillium dahliae. Pectolytic bacteria in the genus Pectobacterium can also cause PED symptoms as well as aerial stem rot (ASR) of potato. Both pathogens can be present in potato production settings, but it is not entirely clear if additive or synergistic interactions occur during co-infection of potato. The objective of this study was to determine if co-infection by V. dahliae and Pectobacterium results in greater PED or ASR severity using a greenhouse assay and quantitative real-time PCR to quantify pathogen levels in planta. PED symptoms caused by Pectobacterium carotovorum subsp. carotovorum isolate Ec101 or V. dahliae isolate 653 alone included wilt, chlorosis and senescence and were nearly indistinguishable. Pectobacterium wasabiae isolate PwO405 caused ASR symptoms including water-soaked lesions and necrosis. Greater Pectobacterium levels were detected in plants inoculated with PwO405 compared to Ec101, suggesting that ASR can result in high Pectobacterium populations in potato stems. Significant additive or synergistic effects were not observed following co-inoculation with these strains of V. dahliae and Pectobacterium. However, infection coefficients of V. dahliae and Ec101 were higher and premature senescence was greater in plants co-inoculated with both pathogens compared to either pathogen alone in both trials, and V. dahliae levels were greater in basal stems of plants co-inoculated with either Pectobacterium isolate. Overall, these results indicate that although co-infection by Pectobacterium and V. dahliae does not always result in significant additive or synergistic interactions in potato, co-infection can increase PED severity.

In this study, symptoms closely resembling those caused by Pseudomonas syringae pv. actinidiae were found in kiwifruit plants (Actinidia chinensis cv. Hort16A) growing in Jeju province, Korea. Bacterial strains including strain KKH3T were isolated from representative lesions on kiwifruit plants. Comparative 16S rRNA gene sequence-based phylogenetic analysis placed these bacteria in a separate cluster within the genus Pectobacterium. Strain KKH3T showed the highest similarity (98.7%) to the recognised bacteria Pectobacterium carotovorum subsp. carotovorum and Pectobacterium wasabiae. To justify subspecies level differentiation, the partial nucleotide sequences of atpD, carA and recA from strain KKH3T were compared with those from other members of the family Enterobacteriaceae. Distant relationships between strain KKH3T and six reference strains of Pectobacterium spp. were evident in the phylogenetic tree. The results of the DNA–DNA hybridisation and polyphasic analysis revealed that strain KKH3T b...

O-Polysaccharides were isolated from the lipopolysaccharides of two strains of plant pathogenic bacteria Pectobacterium wasabiae isolated in Poland in 2013 (IFB5408 and IFB5427). The purified polysaccharides were analyzed using 1D and 2D NMR spectroscopy ((1)H, DQF-COSY, TOCSY, ROESY, HSQC, HSQC-TOCSY, and HMBC) and the chemical methods. Sugar and methylation analyses of native polysaccharides, absolute configuration assignment of constituent monosaccharides together with NMR spectroscopy data revealed that the chemical structures of both O-polysaccharides are the same.

Taimepatogeenide levimise piiramiseks on oluline mõista nende liikumist taimes. Selleks kasutatakse meie uurimisgrupis mudelorganismina märgmädanikku põhjustavat Pectobacterium wasabiae SCC3193 tüve. Selles bakalaurusetöös analüüsitakse nii metanooliga kartulimugulatest ekstraheeritud fenoolsete ühendite kui ka üksikute fenoolsete ühendite, nagu salitsüülhape ja klorogeenhape, mõju mudelorganismi voogamisele in vitro.

Pectobacterium wasabiae (previously known as Erwinia carotovora ) is an important plant pathogen that regulates the production of plant cell wall-degrading enzymes through an N -acyl homoserine lactone-based quorum sensing system and through the GacS/GacA two-component system (also known as ExpS/ExpA). At high cell density, activation of GacS/GacA induces the expression of RsmB, a noncoding RNA that is essential for the activation of virulence in this bacterium. A genetic screen to identify regulators of RsmB revealed that mutants defective in components of a putative Trk potassium transporter ( trkH and trkA ) had decreased rsmB expression. Further analysis of these mutants showed that changes in potassium concentration influenced rsmB expression and consequent tissue damage in potato tubers and that this regulation required an intact Trk system. Regulation of rsmB expression by potassium via the Trk system occurred even in the absence of the GacS/GacA system, demonstrating that these systems act independently and are both required for full activation of RsmB and for the downstream induction of virulence in potato infection assays. Overall, our results identified potassium as an essential environmental factor regulating the Rsm system, and the consequent induction of virulence, in the plant pathogen P. wasabiae . IMPORTANCE Crop losses from bacterial diseases caused by pectolytic bacteria are a major problem in agriculture. By studying the regulatory pathways involved in controlling the expression of plant cell wall-degrading enzymes in Pectobacterium wasabiae , we showed that the Trk potassium transport system plays an important role in the regulation of these pathways. The data presented further identify potassium as an important environmental factor in the regulation of virulence in this plant pathogen. We showed that a reduction in virulence can be achieved by increasing the extracellular concentration of potassium. Therefore, this work highlights how elucidation of the mechanisms involved in regulating virulence can lead to the identification of environmental factors that can influence the outcome of infection.

Pectolytic bacteria were isolated from potato tubers and stems showing tuber soft rot and blackleg symptoms. Approximately half (52 %) of the isolates could grow at both 27 and 37 °C while another half (48 %) failed to grow at 37 °C. All isolates could be amplified with primers specific to the pectate lyase (pel) gene. Carbon utilisation profiles could not conclusively identify these isolates. PCR amplification using primers specific for Pectobacterium carotovorum subsp. brasiliensis was positive for all isolates that grew at 37 °C. However, the group that did not grow at 37 °C failed to amplify with P. atrosepticum specific primers. To characterise this group of isolates, the intergenic transcribed spacer region (ITS) was amplified and PCR products digested with two restriction enzymes (RsaI and CfoI) to generate ITS-PCR-RFLP profiles. The profiles of these new isolates were compared to those of the type strains of other pectolytic bacteria. Profiles of five of the selected atypical strains generated with the enzyme CfoI appeared to be most similar to those of P. wasabiae type strain. Phylogenetic analysis using concatenated partial gene sequences of housekeeping genes mdh and gapA clustered these isolates together with those of P. wasabiae reference strains thus confirming their identity. These strains were virulent on potato tubers and stems but did not elicit hypersensitive response on tobacco plants. This is the first report of P. wasabiae causing soft rot and blackleg of potatoes in South Africa.

Ramie (Boehmeria nivea and Boehmeria tenacissima) is a widely used fiber crop. Traditional water retting or chemical boiling method performed in order to extract ramie fiber seriously pollute the environment and severely damage the fiber, so biological method is the general trend of the fiber-extracting industry. Some strains (687), involving 26 genera and 43 species, were collected from the three samples, which produce hydrolyzed circles in the selective culture medium in order to detect the degumming effect and to compare the enzyme activity. Among these strains, 13 of them did not produce cellulase and had a ramie decreasing weight rate above 25 %, which were regarded as efficient ramie-degumming strains named from R1 to R13. R1 to R13 belonged to Amycolata autotrobutylicun, Bacillus subtilis, Clostridium acetobutylicum, Bacillus subtilis, Rhizobium leguminosarum, Bacteroides finegoldii, Streptomyces lividans, Bacillus amyloliquefaciens, Clostridium acetobutylicum, Pseudomonas brassicacearum, Bacillus pumilus, Bacillus licheniformis, Pectobacterium wasabiae respectively. Bacteroides sp., Rhizobium sp. and Pseudomonas sp. were firstly reported to be used in ramie-degumming. At the same time, the pectinase was the key enzyme in the ramie-degumming process.

Amplified fragment length polymorphism (AFLP) markers and multilocus sequence analysis (MLSA) were used to analyse 63 bacterial strains, including 30 soft-rot-causing bacterial strains collected from Syrian potato fields and 33 reference strains. For the MLSA, additional sequences of 41 strains of Pectobacterium and Dickeya, available from the NCBI GenBank, were included to produce a single alignment of the 104 taxa for the seven concatenated genes (acnA, gapA, proA, icd, mtlD, mdh and pgi). The results indicate the need for a revision of the previously classified strains, as some potato-derived Pectobacterium carotovorum strains were re-identified as P. wasabiae. The strains that were classified as P. carotovorum during the analyses demonstrated high heterogeneity and grouped into five P. carotovorum highly supported clusters (PcI to PcV). The strains represented a wide range of host plants including potatoes, cabbage, avocados, arum lilies, sugar cane and more. Host specificity was detected in PcV, in which four of the six strains were isolated from monocotyledonous plants. The PcV strains formed a clearly distinct group in all the constructed phylogenetic trees. The number of strains phylogenetically classified as subspecies ‘P. c. subsp. brasiliensis’ in PcIV dramatically increased in size as a result of the characterization of new isolates or re-identification of previous P. carotovorum and P. atrosepticum strains. The P. carotovorum strains from Syria were grouped into PcI, PcII and PcIV. This grouping indicates a lack of correlation between the geographical origin and classification of these pathogens.

This paper briefly reviews research on the causative agents of blackleg and soft rot diseases of potato, namely Pectobacterium and Dickeya species, and the disease syndrome, including epidemiological and aetiological aspects. It critically evaluates control methods used in practice based on the avoidance of the contamination of plants, in particular the use of seed testing programmes and the application of hygienic procedures during crop production. It considers the perspective of breeding and genetic modification to introduce resistance. It also evaluates the application of physical and chemical tuber treatments to reduce inoculum load and examines the possibility of biocontrol using antagonistic bacteria and bacteriophages.

Dickeya species (formerly Erwinia chrysanthemi) cause diseases on numerous crop and ornamental plants world-wide. Dickeya spp. (probably D. dianthicola) were first reported on potato in the Netherlands in the 1970s and have since been detected in many other European countries. However, since 2004–5 a new pathogen, with the proposed name ‘D. solani’, has been spreading across Europe via trade in seed tubers and is causing increasing economic losses. Although disease symptoms are often indistinguishable from those of the more established blackleg pathogen Pectobacterium spp., Dickeya spp. can initiate disease from lower inoculum levels, have a greater ability to spread through the plant’s vascular tissue, are considerably more aggressive, and have higher optimal temperatures for disease development (the latter potentially leading to increased disease problems as Europe’s climate warms). However, they also appear to be less hardy than Pectobacterium spp. in soil and other environments outside the plant. Scotland is currently the only country in Europe to enforce zero tolerance for Dickeya spp. in its potato crop in an attempt to keep its seed tuber industry free from disease. However, there are a number of other ways to control the disease, including seed tuber certification, on-farm methods and the use of diagnostics. For diagnostics, new genomics-based approaches are now being employed to develop D. dianthicola- and ‘D. solani’-specific PCR-based tests for rapid detection and identification. It is hoped that these diagnostics, together with other aspects of ongoing research, will provide invaluable tools and information for controlling this serious threat to potato production

Characteristics of forty strains from macerated potato tubers and water-soaked lesions of some ornamental plants were studied in north parts of Iran. The causal organisms isolated from infected tissues were identified as Pectobacterium spp. based on their physiological and biochemical assays and confirmed by species and subspecies specific PCR and RFLP analysis of 16S–23S intergenic transcribed spacer region. Artificial inoculation of isolates to their related hosts generated the same symptoms on potato and ornamental plants, from which the same bacteria were isolated and identified. We detected two groups of atypical isolates in this study. The first group from potato classified as Pectobacterium carotovorum subsp. carotovorum by phenotypic tests but was unable to elicit HR on tobacco leaves, to grow at 37°C and to amplify the pel gene relevant to this subspecies. The second one from ornamental plants which was again characterized as Pectobacterium carotovorum subsp. carotovorum in biochemical assays, produced a unique ITS-RFLP profile different from all of known Pectobacterium species and subspecies. Our findings based on phylogenetic analysis using concatenated partial sequences of housekeeping genes mdh and gapA, indicated the occurrence of P. wasabiae as a novel species in potato storage in Iran. Furthermore we detected a distinct clade of Pectobacterium spp. from some ornamental plants including Schlumbergera bridgesii, Syngonium podophyllum and Iris spp.