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Bacterial soft rot caused by Pectobacterium species is a serious disease in konjac (Amorphophallus konjac), a healthy source of starch particularly in East Asia. An effective diagnostic method is crucial to control the disease and reduce losses in konjac production. In this study, we evaluated a loop-mediated isothermal amplification (LAMP) assay with a specific primer set for the rapid and accurate detection of P. aroidearum. A comparative genomics approach was used to identify the specific genes suitable for the design of LAMP primers. The candidate target genes were determined through a first-round comparison with a 50-genome nucleotide database, and subjected to a second-round screening with the GenBank NR database. As a result, nine specific genes of P. aroidearum were selected for LAMP primer design. After screening of the primers, the primer set 1675-1 was chosen for LAMP detection owing to its high specificity and sensitivity. The LAMP assay could detect the presence of P. aroidearum genomic DNA at a concentration as low as 50 fg and 1.2 &times, 104 CFU/g artificially infected soil within 40 min at 65 &deg, C. Subsequently, this primer set was successfully used to specifically detect P. aroidearum in naturally infected and non-symptomatic plant samples or soil samples from the field. This study indicates that a comparative genomic approach may facilitate the development of highly specific primers for LAMP assays, and a LAMP diagnostic assay with the specific primer set 1675-1 should contribute to the rapid and accurate detection of soft-rot disease in konjac at an early stage.

Wash water from 66% of 401 samples of fresh fruits and vegetables collected in the marketplace and affected by bacterial soft rot were positive for suspected strains of Salmonella, i.e., black, hydrogen sulfide-positive colonies on Salmonella-Shigella agar incubated for 24 h at 37°C. By comparison, 30% of 402 healthy samples were positive. Incidence of suspected Salmonella in broth enrichment cultures was 59% in 533 soft rotted samples and 33% in 781 healthy samples. Thirty percent of 166 representative strains of suspected Salmonella, selected at random from 20 different commodities, were confirmed to be Salmonella by physiological and serological tests. Adjusting incidence values accordingly, Salmonella contamination was potentially present in at least 18 to 20% of soft rotted samples and in 9 to 10% of healthy samples. Wash water from 120 paired healthy and soft rotted fruits and vegetables contained an average of 1.0 × 105and 3.7 × 106CFU/ml, respectively, of suspected Salmonella-a ratio of 1:37. Average concentrations of suspected Salmonella in enrichment cultures of healthy and soft rotted samples were 7.5 × 107and 2.7 × 109CFU/ml, respectively, also in the ratio of 1:37. Fresh potato, carrot, and pepper disks coinoculated with the soft rot bacterium Erwinia carotovora and with Salmonella typhimurium, and incubated for up to 72 h at room temperature, contained approximately 10 times the concentration of S. typhimurium as did disks inoculated with Salmonella alone. Disks coinoculated with Pseudomonas viridiflava and S. typhimurium contained approximately three times the Salmonella populations as disks inoculated with Salmonella alone.

Bacterial root and stem rot of sweetpotato (Ipomoea batatas (L.) Lam.) was first fully characterized in the U.S. in 1977 (2). It was thought to be caused exclusively by Erwinia chrysanthemi. Although a previous report described sweetpotato as a host for E. carotovora subsp. carotovora, based on artificial inoculations, others have reported that neither E. carotovora subsp. carotovora nor E. carotovora subsp. atroseptica decay sweetpotato storage roots (1). In October 1995, storage roots of sweetpotato cv. Beauregard were received from St. Landry Parish, LA, that displayed typical bacterial root rot. Isolations from these roots yielded bacteria that showed a similarity of 0.945 to E. carotovora subsp. carotovora with the Biolog GN Bacterial Identification System (version 3.50). This isolate (Ecc-LH) also differed from isolates of E. chrysanthemi (Ech) from sweetpotato and other hosts in that it was insensitive to erythromycin, did not produce phosphatase or lecithinase, and did not produce gas from glucose. Ecc-LH differed from known strains of E. carotovora subsp. atroseptica in that it did not produce reducing substances from sucrose or acid from palatinose. When Beauregard storage roots were inoculated by inserting micropipette tips containing 50 μl of 1.0 × 108 CFU/ml, both Ecc-LH and Ech-48 produced typical bacterial root rot symptoms. However, when they were compared by infectivity titrations at 28 to 32°C, Ecc-LH was less virulent than Ech-48. Ecc-LH had an ED50 of approximately 1.0 × 106 CFU/ml and did not cause appreciable disease below inoculum concentrations of 1.0 × 105, whereas Ech-48 had an ED50 of approximately 1.0 × 108 and caused soft rot at the lowest concentration tested, 1.0 × 103. Similar disease incidence was observed in infectivity titrations at 22 to 24°C, but Ech-48 caused less severe soft rot. E. carotovora subsp. carotovora was reisolated from inoculated storage roots and its identity was reconfirmed by Biolog. When terminal vine cuttings of Beauregard were dipped in 1.0 × 108 CFU/ml and planted in a greenhouse, bacterial stem rot symptoms developed on plants inoculated with Ech-48 at about 4 weeks postinoculation, or when new growth began. However, no symptoms developed on plants inoculated with Ecc-LH. This is the first report of natural occurrence of E. carotovora subsp. carotovora causing bacterial root rot of sweetpotato in Louisiana. E. chrysanthemi remains the most important pathogen causing bacterial soft rot in sweetpotato since it is widely associated with sweetpotato, is more virulent on storage roots and also causes a stem rot. E. carotovora subsp. carotovora can cause root rot, but has been isolated in only one location to date, is less virulent on storage roots, and apparently does not cause stem rot on the predominant cultivar in U.S. sweetpotato production, Beauregard. References: (1) C. A. Clark and J. W. Moyer. 1988. Compendium of Sweet Potato Diseases. American Phytopathological Society, St. Paul, MN. (2) N. W. Schaad and D. Brenner. Phytopathology 67:302, 1977.

EDTA and the antibiotic nisin, in combination with heat, were tested for inhibition of growth of six pectolytic, soft-rotting bacteria in 80% trypticase soy broth (TSB). Fifty percent reduction of growth by EDTA at 25°C in TSB occurred at 3.24 mM for Erwinia chrysanthemi, 2.57 mM for Pseudomonas fluorescens, 0.96 mM for E. carotovora (subsp. carotovora), 0.48 mM for P. viridiflava, 0.17 mM for Xanthomonas campestris (pv. campestris), and 0.16 mM for Cytophaga johnsonae. Nisin at 50 ug/ml was effective against X. campestris and C. johnsonae (over 90% inhibition of growth) but not against the other four bacteria (less than 20% inhibition), which are the more economically important soft-rotters. Combinations of EDTA and nisin were synergistic. A combination of 0.3 mM EDTA + nisin at 50 μg/ml inhibited growth of E. carotovora, E. chrysanthemi, and P. viridiflava by over 70%, and growth of P. fluorescens by 37%. Hot water treatments for 0.3 min at 37 or 49°C reduced survival of bacteria in the presence of EDTA + nisin, but not of EDTA, nisin, or water alone. EDTA + nisin at 37°C reduced CFU/ml of E. carotovora, E. chrysanthemi, P. fluorescens, and P. viridiflava by 2 log units, and at 49°C by 3 log units, compared with the 25°C treatment. Decay of carrot disks inoculated at two inoculum levels (103 and 104 CFU per disk) with E. carotovora, P. fluorescens, or P. viridiflava was reduced by a 1.5-min immersion in 45°C water, with or without EDTA and nisin additives. Immersion in 0.3 mM EDTA + nisin at 15 to 50 μg/ml at 45°C reduced decay due to E. carotovora and to P. fluorescens at the lower inoculum level by an average of about 50% compared with water alone at 45°C, but differences were statistically significant only at the 90% level of confidence and no different than a standard chlorine dip, current commercial practice for cut carrot slices.

Glucosinolate degradation products are known to suppress microbes. Brassica species produce glucosinolates. Previous investigations determined that susceptibility to bacterial soft rot of broccoli (Brassica oleracea (Italica group)) varied significantly by cultivar. To evaluate the impact of glucosinolates on Pseudomonas marginalis, a causal agent of bacterial soft rot, glucosinolates were measured in lyophilized florets from broccoli ‘Arcadia’, ‘Emperor’, ‘Green Comet’, ‘Green Valiant’, ‘Marathon’, ‘Packman’, ‘Premium Crop’, and ‘Shogun’. Total glucosinolate content was highest in ‘Shogun’ (29.8 μmol/g) and lowest in ‘Emperor’ (0.5 μmol/g). In an in vitro assay, simple linear regression analysis showed that 48% of differences in suppression of P. marginalis growth could be explained by differences in total glucosinolate content (P ≤ 0.01). Plant breeding efforts should include glucosinolate levels as a factor in selecting for disease resistance.

Chlorine dioxide (ClO2) gas was generated from a mixture of sodium chlorite and ferric chloride plus water (impregnated into zeolite) in a Tyvek sachet over a 2- or 24-h period. The gas was distributed by a fan over wound-inoculated tomato fruit (Lycopersicon esculentum) enclosed in a sealed aluminum pressure cooker. Within 24 h of inoculation with 6 log10 CFU of Erwinia carotovora subsp. carotovora per wound and storage at 22 to 24°C, bacterial soft rot was observed on >80% of the nontreated wounds (10 wounds/fruit and 4 or 6 fruit/treatment). By contrast, wounds that had been exposed to an atmosphere containing up to 99 mg ClO2 during a 2- or 24-h period remained firm and dry with no evidence of bacterial activity or soft rot. After 72 h of incubation, wounds exposed to 88 mg ClO2 produced over 24 h or 99 mg ClO2 produced over 2 h were free of decay, whereas bacterial soft rot was observed in ca. 12% and less than 5% of wounds treated with 0.75 mg or 7.5 mg, respectively, for either 2 or 24 h. Wounds that had not been inoculated remained free of bacterial soft rot throughout the entire storage period. Wounds exposed to the highest doses of ClO2, 88 mg/24 h or 99 mg/2 h, became bleached and sunken. Additionally, the stem scars on these fruit became cracked, sunken, and bleached. The intact cuticle was not visibly affected, and there was no observed change in overall fruit color. ClO2 gas may be effective for controlling postharvest decays of fruit that have been inoculated prior to or during harvest.

Vanda orchids are epiphytes grown for their attractive flowers by commercial producers and hobbyists throughout Florida. In August 2007, five Vanda hybrids, with an economic value of $150 each, were found at a nursery in central Florida with leaves that were macerated, brown, and water soaked. According to the growers, the plants were normal the previous day but symptoms developed rapidly. The plants were immediately removed from the greenhouse to prevent potential disease spread. Bacteria were isolated according to the method of Schaad et al. (1). Isolated bacteria grew at 37°C, were gram negative, degraded pectate, and produced phosphatase. MIDI (Sherlock version TSBA 4.10; Microbial Identification 16 System, Newark, DE) (SIM 0.906) identified the bacteria as Erwinia chrysanthemi (Dickeya chrysanthemi Burkholder et al. 1953) Samson et al. 2005. PCR was performed on the 16S rRNA gene (GenBank Accession No. EU526397) with primers 27f (5′-GAGAGTTTGATCCTG GCTCAG-3′) and 1495r (5′-TACGGCTACCTTGTTACGA-3′) (2). Subsequent DNA sequencing and GenBank search showed the isolated strain is 99% identical to that of Dickeya chrysanthemi. Four leaves each of six Vanda hybrids were inoculated by injecting approximately 150 μl of a bacteria suspension at 1 × 108 CFU/ml into each leaf. One plant was inoculated with water in each of four leaves. Plants were enclosed in plastic bags and returned to the greenhouse under 50% shade at 29°C day and 17°C night temperatures. Within 24 h, soft rot symptoms appeared on inoculated leaves. The water control appeared normal. D. chrysanthemi was reisolated and identified with the above method, thus Koch's postulates were fulfilled. To our knowledge, this is the first report of a soft rot caused by D. chrysanthemi on Vanda hybrids. Because of the popularity and high value of Vanda orchids, proper identification of this rapidly progressing bacterial disease is of great importance for the commercial producer and homeowner alike. References: (1) 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. (2) W. G. Weisburg. J. Bacteriol. 173:697, 1991.

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.

Konnyaku (Amorphophallus rivieri Durieu) is grown in some rural areas of China as an important cash crop. In 2011, there was a serious outbreak of Konnyaku soft rot in Xuanwei District of Yunnan Province of China. The disease was characterized by partial or complete tuber rot. At its anaphase, the soft rot may move up the stem, causing the caudex to decay and the whole plant to collapse. If the stem is strong or big enough, the soft rot may develop on one side of the stem, leaving the other side healthy for several days. In this case, the stem does not collapse, and etiolation may be observed on the rotten tissue. In serious cases, up to 80% of the plants were infected. The disease is even more serious if Konnyaku is grown continuously in the same field for more than one year. At its worst, the disease can wipe out the whole crop. In 2012 and 2013, we isolated 46 strains of bacteria from 60 Konnyaku tuber samples with soft rot symptoms from Xuanwei District. All strains grew on CVP medium, and produced iridescent, cross-hatched translucent colonies in deep, cuplike depressions or pits. All strains were facultatively anaerobic, gram-negative, straight rod-shaped cells with peritrichous flagella. All strains were catalase-positive, but oxidase-negative. They were able to ferment glucose, reduce nitrate, produce β-galactosidase and H2S, and they utilized L-arabinose, D-galactose, D-glucose, glycerol, D-mannose, D-ribose, and sucrose, but did not produce urease, or acid from adonitol. Pectobacterium carotovorum subsp. carotovorum (syn. Erwina carotovora subsp. carotovora) has been commonly accepted as the causal agent of Konnyaku soft rot in Japan and China (1,3). Our studies also confirmed that P. carotovorum subsp. carotovorum caused Konnyaku soft rot, but the colony morphology and physiological and biochemical characteristics of these bacteria differed greatly from those of P. carotovorum subsp. carotovorum and other pectolytic Pectobacterium species. They grew at 37°C, caused potato soft rot, produced acid from melibiose, citrate, raffinose, and lactose, but did not produce acid from sorbitol and arabitol. The strain also utilized malonate but not keto-methyl glucoside as the sole carbon source. All strains were positive for phosphatase. Forty-one of 46 strains were sensitive to erythromycin. Thirty-seven of 46 strains produced indole. All tests were conducted with P. carotovorum subsp. carotovorum standard strain C2 isolated from Chinese cabbage as a positive control. Healthy Konnyaku tubers were inoculated with suspensions of the strains with a concentration of 108 CFU/ml in sterile water to confirm pathogenicity. After ~48 h, tuber rot symptoms were observed on all inoculated Konnyaku tubers. In comparison, there were no symptoms on tubers inoculated with sterile water. The bacterium was re-isolated from the infected Konnyaku tubers and identified as Dickeya dadantii, in accordance with Koch's postulates. All strains were confirmed by using the species-specific primers ERWFOR/CHRREV (2), which amplified a 450-bp DNA fragment by PCR assay. To our knowledge, this is the first report of Konnyaku soft rot caused by D. dadantii in China. References: (1) N. Hayashi. Gunma J. Agric. Res. A (Genera1) 5:25, 1988. (2) E. J. Smid et al. Plant Pathol. 44:1058, 1995. (3) J. Y. Tang et al. J. Yunnan Agric. Univ. 16:185, 2001.

Horn lian (Typhonium giganteum) is a perennial herb of the family Aracea and is commonly used for expelling phlegm and as an antispasmodic treatment. In August 2012, horn lian grown in Changchun, Jilin Province of China, exhibited soft rot disease with ~60% incidence and experienced great losses. Water-soaked and dark green lesions on leaves expanded along main veins. Semitransparent, water-soaked, and sunken lesions on stems expanded rapidly and caused the whole plant to collapse with a foul smell. Nine representative strains were isolated from infected leaves and stems on nutrient agar (NA) medium after 36 h incubation at 28°C (1). Colonies were round, shiny, grayish white, and convex on NA medium. All strains were gram-negative, non-fluorescent on King's B medium (KB), facultatively anaerobic, motile with three to six peritrichous flagella (observed by electron transmission microscope), positive for catalase and pectolytic activity test on potato slices, but negative for oxidase, urease, and lecithinase. Strains grew at 37°C and in yeast salts broth medium containing 5% NaCl. They also liquefied gelatin and reduced nitrate, but did not reduce sucrose. Strains were also negative for starch hydrolysis, malonate utilization, gas production from glucose and indole. Results were variable for the Voges-Proskauer test. The strains utilized sucrose, arabinose, fructose, D-galactose, D-glucose, inositol, lactose, D-mannose, D-mannitol, melibiose, rhamnose, salicin, trehalose, maltose, raffinose, glycerol, D-xylose, and cellobiose as carbon sources, but not melezitose, α-CH3-D-gluconate, sorbitol, or dulcitol. Species identity was confirmed by molecular characterization of one of the nine strains, DJL1-2. DNA GC content indicated by high performance liquid chromatography (HPLC) was 51.7%. The 16S rDNA sequence (KC07897) of DJL1-2 showed 99% identity to that of a Pectobacterium carotovorum subsp. carotovorum (Pcc) strain (CP001657) and the sequence of the 16S-23S rDNA spacer region (KJ623257) was 93% similar to that of another known strain of Pcc (CP003776). As a result, the strains were identified as Pcc (2). Pathogenicity of the nine strains was evaluated by spraying 1 ml of bacterial cell suspension (108 CFU/ml) onto healthy leaves and injecting 0.1 ml of cell suspension into stems of 3-year-old horn lian plants with a sterile pipette tip. Three seedlings were used for each strain and sterilized water served as negative controls. Pcc SMG-2 reference strain (from milk thistle) was also inoculated into horn lian leaves and stems. Inoculated plants were covered with plastic bags for 24 h in a greenhouse at 28 to 30°C. After 72 h, water-soaked lesions similar to the naturally infected plants were observed on leaves and stems inoculated by the nine isolated strains and Pcc SMG-2, while negative control plants remained symptomless. Biochemical tests and 16S rDNA sequence analysis confirmed that the re-isolated bacteria were Pcc. To our knowledge, this is the first report of Pcc causing bacterial soft rot of horn lian in Changchun, Jilin Province, China. References: (1) Z. D. Fang. Research Method of Phytopathology. China Agricultural Press, 1998. (2) N. W. Schaad, et al. Laboratory Guide for Identification of Plant Pathogenic Bacteria, 3rd ed. American Phytopathological Society, St. Paul, MN, 2001.

During 2014 to 2015, a devastating bacterial soft rot on cucumber stems and leaves occurred in Shandong, Shanxi, Hebei, Henan, and Liaoning provinces of China, resulting in serious economic losses for cucumber production. The gummosis emerged on the surface of leaves, stems, petioles, and fruit of cucumber. The basal stem color was dark brown and the stem base turned to wet rot. Yellow spots and wet rot emerged at the edge of the infected cucumber leaves and gradually infected the leaf centers. In total, 45 bacterial strains were isolated from the infected tissues. On the basis of phenotypic properties of morphology, physiology, biochemistry, and 16S ribosomal RNA gene sequence analysis, the pathogen was identified as Pectobacterium carotovorum. Multilocus sequence analysis confirmed that the isolates were P. carotovorum subsp. brasiliense, and the pathogens fell in clade II. The pathogenicity of isolated bacteria strains was confirmed. The strains reisolated were the same as the original. The host range test confirmed that strains had a wide range of hosts. As far as we know, this is the first report of cucumber stem soft rot caused by P. carotovorum subsp. brasiliense in China as well as in the world, which has a significant economic impact on cucumber production.

International audience; Pectobacterium carotovorum M022(T) has been isolated from a waterfall source in Selangor district (Malaysia). Using genomic and phenotypic tests, we re-examined the taxonomical position of this strain. Based on 14 concatenated housekeeping genes (fusA, rpoD, rpoS, acnA, purA, gyrB, recA, mdh, mtID, groEL, secY, glyA, gapA and rpIB), multi-locus sequence analysis revealed that strain M022(T) falls into a novel Glade separated from the other Pectobacterium species. The in silico DNA-DNA hybridization and average nucleotide identity values were lower than the 70 and 95% threshold values, respectively. In addition, by combining genomic and phenotypic tests, strain M022(T) may be distinguished from the other Pectobacterium isolates by its incapacity to grow on D(+)-xylose, L-rhamnose, cellobiose and lactose. Strain M022(T) (=CFBP 8629(T)=LMG 30744(T)) is proposed as the type strain of the Pectobacterium fontis sp. nov.

In late September 2019, seven stalks of about 1400 stalks of sugarcane cultivar Zhongzhe 1 exhibited soft rot symptoms in a trial plot in Beihai city, Guangxi province of China. Symptoms included scorched and collapsed leaves, maceration of stalks, and sour smelling exudates from the stalks (Supplementary Fig. S1). Severely diseased stalks had collapsed and were dead. Internal stalk fragments of 5 × 5 mm were collected at the junction of healthy and diseased tissue after surface-sterilization of stalks with 70% ethanol for one minute, and three times rinsing with sterile distilled water. Stalk fragments were placed on Luria-Bertani agar medium (1 % w/v tryptone, 0.5 % w/v yeast extract, 1 % w/v NaCl, 1 % w/v agar, pH7.0) and plates were put in an incubator at 30°C for 48h. Four types of bacterial colonies were obtained, and small and white colonies with irregular margins were the most dominant. A single colony of each type was diluted in sterile distilled water and aliquots of each suspension were streaked on fresh medium plates to obtain pure cultures. Ten eight-week-old stalks (11 th leaf stage) of sugarcane plants, which derived from cuttings of symptomless cultivar Zhongzhe 1, were inoculated by injection of 300 μl of bacterial suspension (3.5x108 CFU/ml) into the stalks. Another 10 stalks were injected with pure water and served as control. The inoculated plants were kept in a greenhouse at 25-37℃.Among the four types of bacteria, only strain BH9 induced symptoms that were identical to those of diseased canes observed in the field (Supplementary Fig. S1). Elongated water-soaked lesions were observed around the inoculation sites three days post inoculation. Five of the 10 BH9-inoculated plants had collapsed two days later. Water-soaked stalks had a sour smell similar to the filed diseased plants. Eight days post inoculation, all BH9-inoculated plants exhibited symptoms but control plants remained symptomless up to 30 days after inoculation. Uniform white colonies with irregular margins were isolated from the inoculated stalks that developed soft rot symptom, and these bacteria caused again stalk soft rot symptoms when inoculated to a new batch of 10 healthy plants. The 16S rRNA gene of strain BH9 was amplified by PCR with primer pair fD2/rP1 and the PCR amplicons from three independent colonies were sequenced. The sequences of the three amplicons were identical (Accession No. MT723897). BLAST alignments of the 16S rDNA sequence from BH9 strain with the GenBank database revealed that BH9 belonged to the genus Dickeya (98.5% identity between D. zeae BH9 and D. zeae EC1). Further PCR assays and sequencing of three genes, DNA polymerase III gamma subunit gene dnaX with primers dnaXf/dnaXr, DNA gyrase gene gyrB with primers gyrBf1/gyrBr1, and recombinase A gene recA with primers recAf/recAr, were performed to identify the species within the genus Dickeya (Zhang et al., 2014). BH9 sequences of these genes (Accession No. MT723898 to MT723900) had highest identity (97.5%, 97.6%, and 97.7%, respectively) with those from D. zeae EC1 (GenBank accession No. CP006929.1). To determine the evolutionary relationship of BH9 to other Dickeya species and strains, a phylogenetic analysis was performed using dnaX, gyrB, and recA sequences. As shown in Supplementary Fig. S2, BH9 clustered with D. zeae strains and formed a lineage distinguishable from other Dickeya species. Among the closest strains, D. zeae NCPPB3531 (Accession No. CM001980.1) was isolated from potato and D. zeae CSL RW192 (Accession No. CM001972.1) from river water (Pritchard et al., 2013). Consequently, strain BH9 was identified as D. zeae. This bacterial species has been reported to cause soft rot in rice (Pu et al., 2012), banana (Zhang et al., 2014), maize (Martinez-Cisneros et al., 2014), and clivia (Hu et al., 2018). To the best of our knowledge, this is the first report of a bacterial stalk rot caused by D. Zeae in sugarcane. In fact, low incidence of D. zeae-caused stalk soft rot was recently found in sugarcane fields in Fusui County, about 150 km north to Beihai. Given the potential threat of this disease to the local sugarcane industry, the mode of transmission, cultivar resistance, and measures to control the disease should be investigated.

Carrot (Daucus carota) is an important root vegetable planted and consumed worldwide (Stein and Nothnagel 1995). In June 2020, carrots (cv. New Kuroda) showing soft rot symptoms were observed in a 600 sqft plot located in Pitou, Changhua, Taiwan (23°54'00.9"N, 120°28'37.3"E; with around 400 plants). About 10% of the plants on site had similar symptoms; infected taproot tissues were macerated (Figure S1) and emitted a foul odor. In most cases, the peels above the rotten tissues remain intact. Two infected plants were brought to the lab. Macerated tissues were suspended in water and examined under a microscope at 600X (without staining). Rod, motile bacteria were observed in all of the samples and the bacteria were isolated onto nutrient agar. Three bacterial strains were obtained from two taproots; strain Car1 was isolated from one plant, and strains Car2 and Car3 were isolated from the other. Their colonies were translucent, round and convex. All isolates could ferment glucose and induce soft rot symptoms on potato tuber slices (Schaad et al. 2001). They were not able to produce indigoidine on yeast dextrose calcium carbonate agar and were tested negative for phosphatase activity (Schaad et al. 2001). The 16S rDNA of Car1 to Car3 were amplified using primers 27F/1492R (Lane 1991). Cloning and sequencing of their 16S rDNA (GenBank accession no. MT889640) revealed that their sequences shared 99.9% identity (1,463/1,464 bp) with that of Pectobacterium aroidearum CFBP 8168T (SCRI 109T; GenBank accession no. NR_159926.1). Multilocus sequence analyses targeting the three isolates' dnaX, leuS and recA genes were conducted. The concatenated sequences (1,596 bp) of Car1 to Car3 and those included in a previous work (Portier et al. 2019) were subjected to phylogenetic analysis. The sequences of Car1 to Car3 were identical (GenBank accession nos. MT892671-MT892673). A maximum-likelihood tree showed that the three isolates belonged to the same clade as P. aroidearum CFBP 8168T (GenBank accession nos. MK516971, MK517115 and MK517259; Figure S2). For the concatenated sequences analyzed, the identity between P. aroidearum CFBP 8168T and our three isolates was 99.4% (1,587/1,596 bp). The pathogenicity of these isolates was determined by inoculating the bacteria into carrot (cv. Xiangyang No.2) taproots. Strains Car1 to Car3 were grown on NA for 48 h (28 °C) and cell suspensions with OD600 values of 0.3 (2.4 x 108 CFU/ml; in water) were prepared. The suspensions of each strain (100 μl) were loaded into 200 μl pipette tips. The tips were then pierced into intact carrot taproots (2.4 cm deep), ejected and left on the plants (one tip per plant). Three taproots were tested for each strain. Tips loaded with 100 μl of water were used for the controls (three replicates). The plants were incubated in a sealed plastic container kept in a growth chamber set at 28°C. After 48 h, all of the inoculated taproots produced soft rot symptoms resembling those observed in the field and plants in the control group did not. Bacteria were re-isolated from macerated tissues of the artificially infected plants and found to share the same leuS sequence with Car1 to Car3. Occurrences of carrot soft rot in Taiwan have only been attributed to Dickeya spp. (Erwinia chrysanthemi) in previous studies (Hsu and Tzeng 1981). The present study is the first report of P. aroidearum infecting carrots in Taiwan. The findings may add to our understanding of the diversity of soft rot pathogens affecting carrot production in Taiwan.

Bacterial soft rot is a devastating disease that harms vegetables. Lactic acid bacteria (LAB) can be effective antagonist against soft rot pathogens. In this study, Lactobacillus paracasei WX322 and its produced bacteriocin had antibacterial activities against a variety of foodborne pathogens and spoilage bacteria, including both Gram-positive and Gram-negative bacteria, especially the soft rot pathogen Pectobacterium carotovorum subsp. brasiliense BZA12 (Pcb BZA12). The in vivo antibacterial test proved that the bacteriocin produced by the L. paracasei WX322 could control soft rot disease of peppers caused by Pcb BZA12. The L. paracasei WX322 produced bacteriocin after fermentation for 36 h, which had broad antimicrobial activity. In addition, the bacteriocin produced by L. paracasei WX322 had good thermal stability and broad pH adaptation. Then, the complete genome of L. paracasei WX322 was sequenced with one circular chromosome and eight plasmids. Three bacteriocin gene clusters were identified in the genome by antiSMASH. Furthermore, seven genes in the three gene clusters were identified to be bacteriocins. Moreover, three of the seven bacteriocins were novel. The complete genome sequences of the L. paracasei WX322 can provide a better molecular basis of its antibacterial activity and biocontrol effect on bacterial soft rot disease.

The postharvest development of bacterial soft rot in tomato fruit caused by Pectobacterium carotovorum is herein linked with inoculation of lenticel-like apertures located around the stem attachment. Stem scars misted with aqueous cell suspensions of P. carotovorum (1 × 106 CFU/ml) or briefly (5 s) immersed in the suspension were likely to become infected during subsequent storage, with a disease incidence exceeding 70% within 7 days at 24°C. Water soaking was initially observed beneath the fruit surface at the juncture of radial walls of endocarp tissues and a corky ring around the stem attachment. If fruit were swirled for 1 min in chlorine at 150 ppm, pH 6.5, within 5 s after inoculation, lesions did not occur. However, if the chlorine treatment was delayed by 120 s, lesions occasionally developed. A 60-min delay usually assured that decay would occur before fruit ripened (within 5 days). Delaying the chlorine wash by 18 h, as would occur if commercial harvests were held overnight prior to packing operations, led to the same incidence of bacterial soft rot as observed among control fruit. Inoculation of fruit with attached stems did not lead to as much decay during storage compared with stemless fruit, suggesting that an intact calyx physically reduced lenticel exposure to external aqueous cell suspensions. Tomato fruit exposed to uncontrolled free moisture during or shortly after harvest have a high potential for postharvest decays and the likely site for infection development is the area surrounding the stem attachment.

Bacterial soft rot caused by Pectobacterium carotovorum subsp. carotovorum is one of the most important diseases of Chinese cabbage. To develop a microbial pesticide that is not only safe but also easily acceptable by consumers, we screened strains of lactic acid bacteria for biocontrol potential. One of 1166 isolates, Lactobacillus plantarum strain BY significantly reduced soft rot disease severity in Chinese cabbage in six different fields in Japan. This isolate also suppressed disease in onion, potato, and tomato. BY populations persisted on wounded Chinese cabbage leaves after spraying. Moreover, BY inhibits pathogen growth in an antibiosis assay. These results strongly suggest that BY inhibits the invasion of the pathogen at wounded sites on leaves and its proliferation in host tissues. This is the first report indicating the application of lactic acid bacteria for biological control of a plant disease under field conditions.

Twenty seven newly introduced genotypes of lettuce collected from different indigenous and exotic sources were characterized with respect to economically important traits by using principle component and regression analyses. Significant differences were observed among the genotypes for all the characters under study. CGN-10944 performed best among all the genotypes for most of the traits under study. Principal component analysis indicated that the first five factors had variances of 3.88, 2.78, 1.47, 1.42 and 1.24 with 25.87%, 18.57%, 9.81%, 9.49% and 8.26% of total variation, respectively and aggregating to 72.01% of total variation. The highest positive values of different traits in different components indicated that main emphasis should be given on the gross head weight (X 2 ), polar (X 6 ) & equatorial diameter (X 7 ) and seed vigour index-I (X 12 ) & II (X 13 ) to have higher yields in lettuce. To quantify the importance of each variable in predicting yield, multiple linear regression model was developed. The coefficient of determination (R 2 ) in linear regression model revealed that 94% of total variation in yield per plot (X 15 ) was influenced by days to marketable maturity (X 1 ), gross head weight (X 2 ), heading percentage (X 4 ), seed germination (X 11 ), seed vigour index-II (X 13 ) and severity of bacterial soft rot (X 14 ). Therefore, main emphasis should be given on days to marketable maturity, gross head weight, polar & equatorial diameter, heading percentage, seed germination, seed vigour index-I & II and severity of bacterial soft rot during selection and hybridization programmes for yield improvement in lettuce.

Soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) is a serious disease in Chinese cabbage (Brassica rapa L. subsp. pekinensis). To reduce the severity of soft rot symptoms in Chinese cabbage, Arabidopsis AtWRKY75 was introduced into Chinese cabbage by Agrobacterium-mediated transformation, which was previously reported to reduce susceptibility to Pcc infection in Arabidopsis. Three independent Chinese cabbage transgenic lines carrying AtWRKY75 were obtained. The growth phenotypes of AtWRKY75 overexpression (OE) lines were normal. Bacterial soft rot symptoms and Pcc growth were reduced in AtWRKY75-OE Chinese cabbage lines compared with WT plants. In contrast, overexpression of AtWRKY75 had no effect on infection with a hemibiotrophic pathogen, Xanthomonas campestris pv. campestris (Xcc) causing black rot disease. These results are consistent with those observed in the transgenic Arabidopsis. We found that AtWRKY75 activated a subset of Chinese cabbage genes related to defense against Pcc infection, such as Meri15B, BrPR4, and BrPDF1.2 (but not BrPGIP2). Moreover, overexpression of AtWRKY75 caused H2O2 production and activation of H2O2 scavenge enzyme genes, suggesting that H2O2 played a role in AtWRKY75-mediated resistance to Pcc. Together, these results demonstrated that AtWRKY75 decreased the severity of Pcc-caused bacterial soft rot and activated a subset of Pcc infection defense-related genes in Chinese cabbage similar to in Arabidopsis. It is suggested that AtWRKY75 is a candidate gene for use in crop improvement, because it results in reduced severity of disease symptoms without concurrent growth abnormalities.

In vitro screening techniques were used to evaluate 46 genotypes of Iranian potato collection for resistance to bacterial soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc). One month old in vitro rooted potato plantlets were inoculated by two inoculation techniques under in vitro conditions: 1) sterile toothpicks dipped into bacterial suspension and pressed into the crown of plantlets and 2) the freshly cut crown of plantlets were dipped into bacterial suspension of 108 cfu ∙ ml-1 for 10 min. Typical soft rot disease symptoms, including the percentage of wilted leaves were recorded on inoculated plantlets 3, 6, 9, 12 and 15 days post-inoculation. The potato genotypes which were examined responded differently to Pcc and varying levels of resistance were observed. Potato genotype AG showed the highest level of resistance. Results obtained from in vitro screening were then verified by classical tuber slice assay. The verifications were conducted using five representative cultivars: Milva, Ramus, Picaso, Marfona and Agria which responded similarly to both in vitro and classical evaluation systems. Similar results obtained from these tests indicated that the in vitro screening technique developed in this study could provide a simple and rapid whole plant assay in selecting resistant potato genotypes against bacterial soft rot.

Potato (Solanum tuberosum L.) production in Zimbabwe is an important element of the agricultural production sector. The local industry is however affected by low yields due to soft rot disease (Pectobacterium carotororum subsp. carotovorum) and poor quality produce. The main thrust of this study was to come up with recommendations that would increase potato production through improved knowledge of cultivar choices according to ranked performance in terms of soft rot infestations. Four experiments were conducted, in the field; in greenhouse, in storage at 10 o C and at room temperature on soft rot inoculated and un-inoculated potato tubers. The experiments were conducted on five locally available varieties in Zimbabwe. Tubers were screened for soft rot infection using counts and weights of infected tubers and BP1 showed significant difference (p

Gamma irradiation was evaluated for its in vitro and in vivo antibacterial activity against a postharvest bacterial pathogen, Erwinia carotovora subsp. carotovora (Ecc). Gamma irradiation in a bacteria cell suspension resulted in a dramatic reduction of the viable counts as well as an increase in the amounts of DNA and protein released from the cells. Gamma irradiation showed complete inactivation of Ecc, especially at a dose of 0.6 kGy. In addition, scanning electron microscopy of irradiated cells revealed severe damage on the surface of most bacterial cells. Along with the morphological changes of cells by gamma irradiation, it also affected the membrane integrity in a dose-dependent manner. The mechanisms by which the gamma irradiation decreased the bacterial soft rot can be directly associated with the disruption of the cell membrane of the bacterial pathogen, along with DNA fragmentation, results in dose-dependent cell inactivation. These findings suggest that gamma irradiation has potential as an antibacterial approach to reduce the severity of the soft rot of paprika.

Bacillus amyloliquefaciens BGP20 is a promising antagonist in controlling post-harvest bacterial soft rot of vegetables caused by Erwinia carotovora subsp. carotovora (Ecc). The objective of this study was to screen a kind of natural and safe additive which could enhance the bio-control activity of BGP20 against post-harvest bacterial soft rot of green pepper. The results of this study indicated that the additive natamycin had stronger inhibition against the pathogen Ecc compared with bamboo vinegar and chitosan in the 2× Yeast extract and Tryptone (2YT) medium. However, natamycin had a slight negative effect on the growth of BGP20 in the 2YT medium. In preventative treatments, natamycin significantly improved the bio-efficacy of BGP20, and enhanced its competitive position against Ecc in the wounds of green pepper. Compared with the treatment with BGP20 alone, the viable count of BGP20 after 72 h of incubation increased by 115.8% in the wounds of green pepper treated with BGP20 and 0.1% natamycin...

Pectobacterium genus comprises numerous soft-causing bacteria affecting vegetable, ornamental, and fruit crops (Charkowski 2018; Ma et al. 2007). Potato is the major host for several Pectobacterium species including P. parmentieri, a newly described species formerly known as P. wasabiae (Khayi et al. 2016). During surveys conducted in January 2018 and January 2019 at Oahu, 67 infected potato samples exhibiting wilting, water-soaked regions with watery ooze, and darkened and necrotic basal stem symptoms extending upward were collected. Symptomatic stems and tubers were cut into 0.5- to 1-cm pieces and surface sterilized with 10% sodium hypochlorite solution for 30 s, followed by three rinses with sterile distilled water. After sterilization, each stem piece was macerated, streaked on crystal violet pectate (CVP) medium, and incubated at 26 ± 2°C for 48 h. Colonies, forming pits on the CVP medium, were restreaked on 2,3,5-triphenyl-tetrazolium chloride medium. DNA was isolated from infected plant tissues using the DNeasy Plant Mini Kit (Qiagen, Germantown, MA). Bacterial genomic DNA from purified cultures was isolated using a DNeasy Blood and Tissue Kit (Qiagen). End-point PCR was performed using the primers Pec.dnaA-F1 and Pec.dnaA-R1 (Dobhal et al. 2019) to amplify the dnaA gene; PCR products were enzymatically purified by adding 2 µl of ExoSAP-IT. Sequencing was performed at the GENEWIZ facility; both forward and reverse strands were aligned and manually edited using Geneious. The BLASTn results of the dnaA region displayed 99 to 100% identity and 100% query cover with all P. parmentieri strains sequences in the NCBI GenBank database. Eleven strains originating from the same field (growing multiple potato cultivars) but from different plants, namely PL30 (MN428432), PL32 (MN428433), PL67 (MN428434), PL70 (MN428435), PL71 (MN428436), PL72 (MN428437), PL74 (MN428438), PL75 (MN428439), PL123 (MN428440), PL124 (MN428441), and PL128 (MN428442), were identified as P. parmentieri. Three-week-old potato plants, grown from healthy tubers under temperature-controlled conditions, were artificially inoculated at three consecutive nodes starting from the base of the stem by injecting 100 µl of bacterial inoculum of PL70 (3.56 × 10⁸ CFU/ml) and PL72 (2.78 × 10⁸ CFU/ml); control plants were inoculated with 100 µl of sterile water, and the experiment was performed in triplicates with consistent results. After ∼72 h, blackleg and stem rot symptoms resembling those observed in the field were observed; no symptoms on control plants were observed. Bacteria from inoculated symptomatic stems were reisolated on CVP; pits were produced by both strains. To fulfill Koch’s postulates, DNA was isolated, amplified, and sequenced for the dnaA region; BLASTn results showed 100% identity with the original P. parmentieri strains. Additionally, a phylogenetic analysis based on the dnaA gene was conducted with the 11 P. parmentieri Hawaiian strains; all these strains were clustered together with 18 other P. parmentieri strains in the GenBank database, further confirming the identity and genetic relationship of these new P. parmentieri strains. This is the first report of P. parmentieri causing soft rot and blackleg disease on potato at Oahu, Hawaii. The presence of this pathogen along with the previous report of Dickeya dianthicola (Boluk and Arif 2019) outline the importance of exerting proper crop management and of accurate and efficient diagnostics to prevent its spread; it also warrants a statewide survey for pectolytic bacteria on vegetable crops.

Pectobacterium species cause serious bacterial soft rot diseases worldwide on economically important fruit and vegetable crops including tomato and potato. Accurate and simple methods are essential for rapid pathogen identification and timely management of the diseases. Recombinase polymerase amplification (RPA) combined with a lateral flow device (LFD) was developed for specific detection of Pectobacterium sp. directly from infected plant materials with no need for DNA isolation. The specificity of RPA-LFD was tested with 26 Pectobacterium sp. strains and 12 non-Pectobacterium species and no false positive or false negative outcomes were observed. RPA primers and probe for host control were also developed to detect the host genome for enhanced reliability and accuracy of the developed assay. The detection limit of 10 fg was obtained with both sensitivity and spiked sensitivity assays. No inhibitory effects were observed on the RPA assay when targets (pathogen and host) were directly detected from infected potato and tomato sap. The developed RPA assay has numerous applications from routine diagnostics at point-of-care, biosecurity, surveillance and disease management to epidemiological studies. In addition, this tool can also be used to discover reservoir hosts for Pectobacterium species.

Lettuce (Lactuca sativa L.) and Chinese cabbage (Brassica pekinensis Rupr.) are leafy vegetables of great socioeconomic value in Brazil. Bacterial diseases are among the factors that might limit their yield, particularly soft rot caused by bacteria of the genus Pectobacterium. In July 2014, vegetable gardens of approximately 1.0 and 0.18 ha in the cities of Juazeiro (09° 24′ 42″ S, 40° 29′ 55″ W), state of Bahia, and Petrolina (09° 23′ 55″ S, 40° 30′ 03″ W), state of Pernambuco, Brazil, showed 5% of lettuce plants variety Elba and 44.4% of Chinese cabbage cultivar Michihilli plants with wilt of leaves, foul-smelling soft rot, and inner medulla disintegration. Bacterial isolates were selectively isolated from stem and midribs of lettuce and Chinese cabbage by transferring rot tissue to a healthy pepper fruit using a sterilized toothpick (Takatsu et al. 1981). Pepper fruits were incubated at 26°C for 36 h in a moist chamber, and after incubation bacterial isolates were isolated in Petri dishes containing casamino acid–peptone–glucose (CPG) medium. Small white colonies with “broken glass” aspect (Kelman and Dickey 1995) were observed under a stereoscope, and after that, they were preserved in sterile distilled water (SDW). Twelve isolates were obtained from lettuce and Chinese cabbage plants with soft rot symptoms, which showed the same cultural characteristics in CPG medium. Pathogenicity testing was performed using seedlings of lettuce variety Elba (30 ± 2°C; 62 ± 2% relative humidity [RH]) and detached leaves of Chinese cabbage cultivar Michihilli (28 ± 2°C; 65 ± 2% RH), which are susceptible to bacterial soft rot. Ten microliters of bacterial suspension (10⁶ CFU/ml) were injected into the intermediary portion of the leaf midrib, followed by 48 h of incubation in a moist chamber. For the negative control, seedlings and detached leaves were treated similarly with SDW. The isolate CRMP1 of Pectobacterium aroidearum (Moraes et al. 2017) was used as a reference. Typical symptoms of soft rot were observed 12 to 24 h after inoculation, followed by pathogen reisolation onto CPG medium, fulfilling Koch’s postulates. Seedlings and detached leaves used as negative control remained symptomless, whereas seedlings and detached leaves inoculated with CRMP1 showed soft rot symptoms. Both isolates (UNEB 3 and CCRMPA35) presented gram-negative cells, grew at 37°C, were erythromycin resistant, and were positive for maceration in potato tubers. The Biolog Gen III System identified the isolates as Pectobacterium sp. with a level of 70% similarity. Two isolates were selected for molecular identification, one from lettuce (UNEB3) and one from Chinese cabbage (CCRMPA35). Fragments of the 16S rDNA region (Hauben et al. 1998) and icdA, proA, and mdH genes (Ma et al. 2007) of these isolates were sequenced, and BLASTn analysis of the 16S rRNA region (GenBank MH507334 and MH507330), icdA (GenBank MH500095 and MH500096), proA (GenBank MH507331 and MH507332), and mdH (GenBank MH507329 and MH507330) genes of the isolates UNEB 3 and CCRM35 showed similarity levels of 99, 99, 98, and 99% with sequences of the 16S rRNA region and icdA, proA, and mdH genes of the type isolate of P. aroidearum (isolate SCRI109ᵀ; GenBank JN600323, HM156915, HM157162, and HM156976). Phylogenetic analysis of concatenated sequences of the icdA, proA, and mdH genes using Bayesian inference between isolates UNEB 3 and CCRM35 and species and subspecies of Pectobacterium from GenBank grouped these two isolates with SCRI109ᵀ, with a posterior probability of 100%. To our knowledge, this is the first report of P. aroidearum causing soft rot in lettuce and Chinese cabbage in Brazilian territory. In addition, the present research helps to better know the host range of P. aroidearum in Brazil, which until now was only reported in zucchini (Moraes et al. 2017).

Pectobacterium carotovorum is the most common causative organism of bacterial soft rot which gives rise to great economic losses. It is important to implement disease control measures and management strategies on this bacterium in order to prevent further crop loss. Among the various management strategies, using the antibacterial nature of spices offer promising solutions to pesticidal hazards and also to increasing cost of plant protection operations. Although there are many reports on the antibacterial nature of spices on various pathogens, its effect on P. carotovorum is limited. Objectives of this study were to determine the antimicrobial activity of selected spices and then to ascertain the minimum inhibitory concentration (MIC) of aqueous and ethanolic extracts of those spices against P. carotovorum. Among the nine tested spices, four were active against P. caratovorum. As depicted by agar well diffusion assay, the highest inhibitory effect on P. carotovorum was shown by the aqueous and ethanol extracts of Allium sativum while the aqueous and ethanolic extracts of Cinnamomum verum gave the smallest zones of inhibition. Garcinia cowa and Tamarindus indica displayed moderate antibacterial activity. There was no significant difference (p>0.05) in inhibition zones between aqueous and ethanol extract of a particular spice. MIC of the spices, which inhibited the growth of P. carotovorum was 0.5 g/ml within the test range. Positive extracts can be used as a cost effective and eco-friendly preventive strategy to minimize the effects of P. carotovorum. Evaluation of the antibacterial activity of these extracts under field conditions is necessary before a cost-effective formulation is developed.

Bacterial soft rot is a disease complex caused by multiple genera of gram-negative and gram-positive bacteria, with Dickeya and Pectobacterium being the most widely studied soft-rot bacterial pathogens. In addition to soft rot, these bacteria also cause blackleg of potato, foot rot of rice, and bleeding canker of pear. Multiple Dickeya and Pectobacterium species cause the same symptoms on potato, complicating epidemiology and disease resistance studies. The primary pathogen species present in potato-growing regions differs over time and space, further complicating disease management. Genomics technologies are providing new management possibilities, including improved detection and biocontrol methods that may finally allow effective disease management. The recent development of inbred diploid potato lines is also having a major impact on studying soft-rot pathogens because it is now possible to study soft-rot disease in model plant species that produce starchy vegetative storage organs. Together, these new discoveries have changed how we face diseases caused by these pathogens.

Four hundred and fifty bacteria were evaluated for antagonistic activity against bacterial soft rot of potato caused by Pectobacterium carotovorum sp strain II16. A strain Ar10 exhibiting potent antagonist activity has been identified as Bacillus amyloliquefaciens on the basis of biochemical and molecular characterization. Cell free supernatant showed a broad spectrum of antibacterial activity against human and phytopathogenic bacteria in the range of 10-60 AU/mL. Incubation of P. carotovorum cells with increasing concentrations of the antibacterial compound showed a killing rate of 94.8 and 96% at MIC and 2xMIC respectively. In addition, the antibacterial agent did not exert haemolytic activity at the active concentration and has been preliminary characterized by TLC and GC–MS as a glycolipid compound. Treatment of potato tubers with strain Ar10 for 72 h significantly reduced the severity of disease symptoms (100 and 85.05% reduction of necrosis deep / area and weight loss respectively). The same levels in disease symptoms severity was also recorded following treatment of potato tubers with cell free supernatant for 1 h. Data suggest that protection against potato soft rot disease may be related to glycolipid production by strain Ar10. The present study affords new alternatives for anti-Pectobacterium carotovorum bioactive compounds against the soft rot disease of potato.

Amorphophallus konjac soft rot can cause severe yield losses, and the effects agent to prevent the disease had not been found currently in production. In this dissertation, inhibition on konjac soft rot bacteria of seven agricultural fungicides, as benzene ring yl ether, methyl thiophanate, streptomycin sulfate, methane frost hymexazol, bismerthiazol WP, gray mold and Dyson Mn-Zn, was determined by antagonistic petri dish method. The results indicated that: the tested fungicide s had certain inhibition on konjac soft rot bacteria, and the inhibitory effect of different fungicides was significant difference. 80% thiophanate-methyl and 72% streptomycin sulfate had a good inhibitory effect on konjac soft rot disease bacteria.

Potato crop is one of the most economically important crops, and is considered as the fourth main food crop in the world. One of the most important serious diseases of potato is bacterial soft rot disease. In our study, soil samples were collected from potato rhizosphere from a potato cultivated field in Giza, Egypt. Two bacteriophages were isolated on Kosakonia sacchari as a host, causing potato soft rot disease. Based on the genome characterization and electron microscopy, the two phages were named as vB_KsaM-C1 belonging to Myoviridae and vB_KsaO-C2 belonging to Microviridae. Both phages sustained their activity more than 20 months with a remarkable decrease in phage titre. In addition both recorded the same TIP at 65°C and showed the optimum activity at pH 7. The DEP of vB_KsaM-C1 was 10-7, while that of phage vB_KsaO-C2 was 10-9. A small scale application of the two isolated bacteriophages on infected potato tubers discs, in the lab, successfully inhibited bacterial soft rot caused by Kosakonia sacchari.