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Plant Disease Apr 2024Blackleg and soft rot are harmful diseases in potato (Solanum tuberosum) caused by Pectobacterium spp. and Dickeya spp. (Czajkowski et al. 2015). The occurrence of...
Blackleg and soft rot are harmful diseases in potato (Solanum tuberosum) caused by Pectobacterium spp. and Dickeya spp. (Czajkowski et al. 2015). The occurrence of potato blackleg was serious in potato-producing areas around Xiapu County in Fujian Province, China, in 2021 (6 ha) and 2022 (7 ha), with an incidence of approximately 5%, which reached nearly 23%. Three diseased plants were collected to isolate the pathogen. Single colonies from each sampled plant were isolated and streaked onto fresh plates. DNA from three colonies from different plants was PCR amplified with primer pair 27F/1492R (Lane 1991) for the 16S rRNA gene. Since the sequences were identical, we selected strain M2-3 for further analysis. The strain M2-3 was gram-negative, pectolytic on CVP, grew at 37°C and 5% NaCl. The bacterium was positive for phosphatase activity, erythromycin sensitivity, indole production, gelatin liquefaction, malonic utilization, and acid production from, melibiose, raffinose, and arabinose. The bacterium was negative for sucrose, α-methyl glucoside, sorbitol, trehalose, lactose, and sodium citrate (Fujimoto et al. 2018;),although sucrose and lactose did not provide the expected results, there are exception in all species. The genome of strain M2-3 was sequenced and deposited in the NCBI database under accession numbers: CP077422. An Average Nucleotide Identity (ANI) analysis showed that M2-3 clustered with other D. dadantii strains and has a 98.39% identity with D. dadantii strain DSM 18020 (CP023467). The housekeeping genes (recA, dnaX, acnA, gapA, icd, mdh, mtlD and pgi) were amplified with primer pairs designed previously(Fujimoto et al. 2018; Ma et al. 2007) and sequenced. A multilocus sequence analysis (MLSA) was performed by concatenating the 8 gene sequences and constructing a maximum likelihood phylogenetic tree using PhyloSuite version 1.2.1 (Zhang et al. 2020) and IQ-tree version 1.6.8 (Nguyen et al. 2015) software. Strain M2-3 was clustered together with Dickeya dadantii. For the pathogenicity test, three plants per treatment, totaling nine plants, were used. Bacterial suspensions (1×10^8 CFU/mL) were made in a 10mM PBS buffer. 10 μL of M2-3, D. dadantii type strain 18020 (positive control), and buffer (negative control) were injected into the plant stems near the base. Water stains appeared at the site of inoculation after 2 days and they gradually became black and rotten. The leaves became yellow and wilted, and the petiole base rotted within 5 days of inoculation completing the Koch postulate. According to average nucleotide identity and housekeeping gene sequence analysis, strain M2-3 was identified as Dickeya dadantii. Previous studies have reported several pathogens that cause potato blackleg in China, including P. atrosepticum, P. carotovorum, P. brasiliense, P. parmentieri, P. polaris, and P. punjabense (Li-ping et al. 2020; Wang et al. 2021). To the best of our knowledge, this study is the first to report potato blackleg disease caused by Dickeya dadantii in Fujian Province, China. This finding suggests that this pathogen may cause a threat to potato production in Fujian Province.
PubMed: 38687907
DOI: 10.1094/PDIS-04-23-0755-PDN -
Phytopathology May 2024Bacteria belonging to the genus cause blackleg and soft rot symptoms on many plant hosts, including potato. Although there is considerable knowledge about the genetic...
Bacteria belonging to the genus cause blackleg and soft rot symptoms on many plant hosts, including potato. Although there is considerable knowledge about the genetic determinants that allow to colonize host plants, as well as the genes that contribute to virulence, much is still unknown. To identify the genes important for fitness in potato stems, we constructed and evaluated randomly barcoded transposon mutant (RB-TnSeq) libraries of and . We identified 169 and 157 genes important for growth in and a in stems, respectively. This included genes related to metabolic pathways, chemotaxis and motility, transcriptional regulation, transport across membranes, membrane biogenesis, detoxification mechanisms, and virulence-related genes, including a potential virulence cluster , c-di-GMP modulating genes, and pectin degradation genes. When we compared the results of the stem assay with other datasets, we identified genes important for growth in stems versus tubers and in vitro conditions. Additionally, our data showed differences in fitness determinants for and . These data provide important insights into the mechanisms used by when interacting with and colonizing plants and thus might provide targets for management.
Topics: Solanum tuberosum; Plant Diseases; Dickeya; Plant Stems; Virulence; Genes, Bacterial; Genetic Fitness
PubMed: 38170668
DOI: 10.1094/PHYTO-09-23-0351-KC -
Scientific Reports Oct 2021RecA plays a central role in DNA repair and is a main actor involved in recombination and activation of the SOS response. It is also used in the context of... (Comparative Study)
Comparative Study
RecA plays a central role in DNA repair and is a main actor involved in recombination and activation of the SOS response. It is also used in the context of biotechnological applications in recombinase polymerase isothermal amplification (RPA). In this work, we studied the biological properties of seven RecA variants, in particular their recombinogenic activity and their ability to induce the SOS response, to better understand the structure-function relationship of RecA and the effect of combined mutations. We also investigated the biochemical properties of RecA variants that may be useful for the development of biotechnological applications. We showed that Dickeya dadantii RecA (DdRecA) had an optimum strand exchange activity at 30 °C and in the presence of a dNTP mixture that inhibited Escherichia coli RecA (EcRecA). The differences between the CTD and C-tail of the EcRecA and DdRecA domains could explain the altered behaviour of DdRecA. D. radiodurans RecA (DrRecA) was unable to perform recombination and activation of the SOS response in an E. coli context, probably due to its inability to interact with E. coli recombination accessory proteins and SOS LexA repressor. DrRecA strand exchange activity was totally inhibited in the presence of chloride ions but worked well in acetate buffer. The overproduction of Pseudomonas aeruginosa RecA (PaRecA) in an E. coli context was responsible for a higher SOS response and defects in cellular growth. PaRecA was less inhibited by the dNTP mixture than EcRecA. Finally, the study of three variants, namely, EcPa, EcRecAV1 and EcRecAV2, that contained a combination of mutations that, taken independently, are described as improving recombination, led us to raise new hypotheses on the structure-function relationship and on the monomer-monomer interactions that perturb the activity of the protein as a whole.
Topics: DNA-Binding Proteins; Deinococcus; Dickeya; Escherichia coli; Escherichia coli Proteins; Pseudomonas aeruginosa; Rec A Recombinases; Species Specificity
PubMed: 34702889
DOI: 10.1038/s41598-021-00589-9 -
Plant Disease Jul 2023Cariru (Talinum fruticosum) is a non-conventional food plant and a source of vitamins A, B2, B5, and C, calcium, phosphorus, and iron. It is an important crop for...
Cariru (Talinum fruticosum) is a non-conventional food plant and a source of vitamins A, B2, B5, and C, calcium, phosphorus, and iron. It is an important crop for producers of the Amazon region and it is used as a leafy green vegetable, similar to spinach, as well as for medicinal purposes. It is cultivated by producers in the states of Pará and Amazonas (Amorim et al. 2014). In February 2020, in gardens of approximately 1.0 and 0.18 ha in the municipality of Ananindeua (01° 34' 12" S - 48° 38' 11" W), Pará, Brazil, we observed approximately 10% of the plants exhibited soft rot symptoms, bacterial oozing, and stem pith disintegration. From cariru infected leaves, the bacteria were selectively isolated using healthy pepper fruit, as described by Moraes et al. (2017). Two strain (UFRADD17 and UFRADD18) were obtained and both showed white, small, "broken glass" colonies on CPG (peptone-casamino acid-glucose) medium when observed in a stereoscope under oblique lighting. Pathogenicity tests were performed on 20-day-old cariru seedlings by depositing 10 ul of the bacterial suspension (106 CFU/ml) onto a wound made on the leaf blade with an entomological pin. After inoculation, seedlings were incubated in a greenhouse for 48 h at 30 ± 2°C and 90 ± 2% RH. Plants treated similarly with sterile distilled water were used as a negative control. Symptoms of soft rot were observed between 12 and 24 h after inoculation. Seedlings used as negative control remained symptomless. Forty-eight hours after inoculation, we reisolated the pathogen and performed rep-PCR (REP, ERIC, and BOX-PCR) analyses (Gama et al. 2018) with the strains used for inoculation and those reisolated from the pathogenicity test to confirm the identity of the strains and to fulfill Koch's postulates. The two re-isolated strains showed the same REP, ERIC, and BOX-PCR profiles as the strains used for inoculation. In addition, there were no differences between the rep-PCR profiles of the isolates UFRADD17 and UFRADD18. These isolates were Gram-negative, grew at 37°C, and were positive for maceration in pepper fruit and potato tubers. Molecular identification of the isolate UFRADD18 was performed from sequencing fragments from the 16S rDNA region and dnaX, fusA, gyrA, and mdH housekeeping genes as previously described (Van der Wolf et al. 2014; Ma et al. 2007). Sequences were deposited in Genbank under accessions OP142347, OP191704, OP191705, OP191706, and OP191707. Blastn analysis showed 96.15% identity with the 16S rDNA region of accession KY231142.1 (400/416 bp), 99.5% identity with dnaX of accession KC844490.1 (396/398 bp), 99.86% identity with fusA of accession CP023467.1 (714/715 bp), 99.74% identity with gyrA of accession KC844598 (387/388 bp), and 99.10% identity with mdH of accession GQ891979.1 (563/560 bp) from the type strain of Dickeya dadantii. A phylogenetic analysis performed by Bayesian inference with the dnaX, fusA, gyrA, and mdH genes grouped the isolate UFRADD18 along with NCPPB898T, with a 1.00 posterior probability. To our knowledge, this is the first report of D. dadantii causing soft rot in cariru in the Brazilian territory. In addition, this report increases understanding of the host range of this bacterium, which is important for adopting management strategies based on the control of alternative hosts.
PubMed: 37408126
DOI: 10.1094/PDIS-02-23-0397-PDN -
Frontiers in Microbiology 2023Tailocins are nanomolecular machines with bactericidal activity. They are produced by bacteria to contribute to fitness in mixed communities, and hence, they play a...
Tailocins are nanomolecular machines with bactericidal activity. They are produced by bacteria to contribute to fitness in mixed communities, and hence, they play a critical role in their ecology in a variety of habitats. Here, we characterized the new tailocin produced by strain 3937, a well-characterized member of plant pathogenic Soft Rot (SRP). Tailocins induced in were 166 nm long tubes surrounded by contractive sheaths with baseplates having tail fibers at one end. A 22-kb genomic cluster involved in their synthesis and having high homology to the cluster coding for the tail of the Peduovirus P2 was identified. The tailocins, termed dickeyocins P2D1 (phage P2-like dickeyocin 1), were resistant to inactivation by pH (3.5-12), temperature (4-50°C), and elevated osmolarity (NaCl concentration: 0.01-1 M). P2D1 could kill a variety of different spp. but not any strain of spp. tested and were not toxic to .
PubMed: 38098664
DOI: 10.3389/fmicb.2023.1307349 -
Biomolecular NMR Assignments Oct 2022The ability to interact and adapt to the surrounding environment is vital for bacteria that colonise various niches and organisms. One strategy developed by...
The ability to interact and adapt to the surrounding environment is vital for bacteria that colonise various niches and organisms. One strategy developed by Gram-negative bacteria is to secrete exoprotein substrates via the type II secretion system (T2SS). The T2SS is a proteinaceous complex spanning the bacterial envelope that translocates folded proteins such as toxins and enzymes from the periplasm to the extracellular milieu. In the T2SS, a cytoplasmic ATPase elongates in the periplasm the pseudopilus, a non-covalent polymer composed of protein subunits named pseudopilins, and anchored in the inner membrane by a transmembrane helix. The pseudopilus polymerisation is coupled to the secretion of substrates. The T2SS of Dickeya dadantii secretes more than 15 substrates, essentially plant cell wall degrading enzymes. In D. dadantii, the major pseudopilin or the major subunit of the pseudopilus is called OutG. To better understand the mechanism of secretion of these numerous substrates via the pseudopilus, we have been studying the structure of OutG by NMR. Here, as the first part of this study, we report the H, N and C backbone and sidechain chemical shift assignment of the periplasmic domain of OutG and its NMR derived secondary structure.
Topics: Adenosine Triphosphatases; Bacterial Proteins; Dickeya; Nuclear Magnetic Resonance, Biomolecular; Periplasm; Polymers; Protein Binding; Protein Subunits; Type II Secretion Systems
PubMed: 35482172
DOI: 10.1007/s12104-022-10085-4 -
Microbiology Spectrum Dec 2023Bacteria respond to environmental changes and adapt to host systems. The response regulator VfmH of the Vfm quorum sensing system regulates a crucial virulence factor,...
Bacteria respond to environmental changes and adapt to host systems. The response regulator VfmH of the Vfm quorum sensing system regulates a crucial virulence factor, pectate lyase (Pel), in . At high c-di-GMP concentrations, VfmH binds c-di-GMP, resulting in the loss of its activation property in the Pel and virulence regulation in . VfmH binds to c-di-GMP three conserved arginine residues, and mutations of these residues eliminate the c-di-GMP-related phenotypes of VfmH in Pel synthesis. Our data also show that VfmH interacts with CRP to regulate transcription, thus integrating cyclic AMP and c-di-GMP signaling pathways to control virulence in . We propose that VfmH is an important intermediate factor incorporating quorum sensing and nucleotide signaling pathways for the collective regulation of pathogenesis.
Topics: Bacterial Proteins; Enterobacteriaceae; Cyclic GMP; Gene Expression Regulation, Bacterial
PubMed: 37811940
DOI: 10.1128/spectrum.01537-23 -
Environmental Microbiology Reports Oct 2014Pectate lyases are enzymes involved in plant cell wall degradation. They cleave pectin using a β-elimination mechanism, specific for acidic polysaccharides. They are... (Review)
Review
Pectate lyases are enzymes involved in plant cell wall degradation. They cleave pectin using a β-elimination mechanism, specific for acidic polysaccharides. They are mainly produced by plant pathogens and plant-associated organisms, and only rarely by animals. Pectate lyases are also commonly produced in the bacterial world, either by bacteria living in close proximity with plants or by gut bacteria that find plant material in the digestive tract of their hosts. The role of pectate lyases is essential for plant pathogens, such as Dickeya dadantii, that use a set of pectate lyases as their main virulence factor. Symbiotic bacteria produce their own pectate lyases, but they also induce plant pectate lyases to initiate the symbiosis. Pectin degradation products may act as signals affecting the plant–bacteria interactions. Bacterial pectate lyases are also essential for using the pectin of dead or living plants as a carbon source for growth. In the animal gut, Bacteroides pectate lyases degrade the pectin of ingested food, and this is particularly important for herbivores that depend on their microflora for the digestion of pectin. Some human pathogens, such as Yersinia enterocolitica, produce a few intracellular pectate lyases that can facilitate their growth in the presence of highly pectinolytic bacteria, at the plant surface, in the soil or in the animal gut.
Topics: Bacteria; Bacterial Infections; Bacterial Proteins; Humans; Plant Diseases; Polysaccharide-Lyases
PubMed: 25646533
DOI: 10.1111/1758-2229.12166 -
Plant Disease Mar 2023(satin pothos or silver vine) is an evergreen climbing plant belonging to the Araceae family, subfamily Monstereae (Bown, 2000), which is also cultivated as a foliage...
(satin pothos or silver vine) is an evergreen climbing plant belonging to the Araceae family, subfamily Monstereae (Bown, 2000), which is also cultivated as a foliage ornamental (Masnira et al. 2019). In September of 2022, soft rot symptoms were observed on potted . plants grown in a greenhouse in Nantun District, Taichung, Taiwan, in which soft rot of another aroid (philodendron) has also been reported (Wu et al. 2023). The symptoms appeared on the petioles and most of them tended to extend to the leaf blades; the colors of leaf lesions ranged from dark brown to gray (Fig. S1). Some 70% of the plants in the greenhouse showed similar symptoms and losses were estimated to be 15-30%. Four symptomatic plants were sampled. Macerated tissues from rotting petioles were soaked in 10 mM MgCl and observed under a light microscope (Nikon, Japan) at 400 x magnification. Motile, rod-shaped bacteria were observed, and 1-2 loopfuls of undiluted sample suspension were streaked onto nutrient agar (NA; Gibco, USA). After culturing at 28°C for 1 day, all samples yielded round, creamy-white colonies (0.9 mm in diameter) and from each of the four samples a pure culture was obtained (Spi1-Spi4). All isolates exhibited oxidative and fermentative metabolism of glucose (Schaad et al. 2001). They caused pitting on crystal violet pectate agar, induced maceration on potato tuber and were tested positive for phosphatase activity and indigoidine production (Lee and Yu 2006; Schaad et al. 2001). Polymerase chain reaction tests using -specific primers 5A and 5B (Chao et al. 2006) amplified the expected amplicon (0.5 kb) in extracted DNA samples of all isolates. Identification of the strains was achieved by amplifying and sequencing fragments of the housekeeping genes , , , , and (Marrero et al. 2013); the lengths of the five gene fragments analyzed were 822, 762, 720, 672, and 450 bp, respectively (accession nos. OP985528-OP985532). The five sequences were concatenated for every isolate; the resulting 3,426 bp sequences were aligned with ClustalW and found to be identical. A maximum-likelihood analysis was conducted using the 3,426-bp sequences and those of known species' type strains. Spi1 to Spi4 clustered with . subsp. NCPPB 2976 and . subsp. CFBP 1269 (Fig. S2) with sequence identities of 98.4 and 98%, respectively. To fulfil Koch's Postulates, stab inoculations of the four isolates into the petioles of cutting propagated, 38-day-old . plants (3 plants per isolate) were conducted using sterile toothpicks. The amounts of bacteria used was approximately 10 cfu per toothpick; the bacterial loads were estimated by suspending the cells in 10 mM MgCl and spread-plating diluted suspensions on NA. Sterile toothpicks were used as control. All tested plants were sealed in plastic bags (containing wet paper towel) and kept in a growth chamber (28°C; 12-h photoperiod). After 1 day, all isolates induced soft rot symptoms resembling those observed under natural conditions in the greenhouse. Bacteria were re-isolated, and they all shared the same sequence with strains Spi1 to Spi4. This is the first report of . affected by . in Taiwan. Further investigation is needed to determine whether Spi1-Spi4 belong to . subsp. . has been found infecting different aroids (Lee and Chen 2021; Lin et al. 2012). The species has also been reported in Taiwan on poinsettia (Wei et al., 2019) and philodendron (Wu et al. 2023). Because these plants are often grown closely in the same facilities, growers should be wary of . 's spread among these plants. Reduction of environmental humidity and avoiding overhead irrigation may be effective in preventing the pathogen's transmission.
PubMed: 36947837
DOI: 10.1094/PDIS-12-22-2885-PDN -
The Journal of Biological Chemistry Jan 2022The catabolism of pectin from plant cell walls plays a crucial role in the virulence of the phytopathogen Dickeya dadantii. In particular, the timely expression of pel...
The catabolism of pectin from plant cell walls plays a crucial role in the virulence of the phytopathogen Dickeya dadantii. In particular, the timely expression of pel genes encoding major pectate lyases is essential to circumvent the plant defense systems and induce massive pectinolytic activity during the maceration phase. Previous studies identified the role of a positive feedback loop specific to the pectin-degradation pathway, whereas the precise signals controlling the dynamics of pectate lyase expression were unclear. Here, we show that the latter is controlled by a metabolic switch involving both glucose and pectin. We measured the HPLC concentration profiles of the key metabolites related to these two sources of carbon, cAMP and 2-keto-3-deoxygluconate, and developed a dynamic and quantitative model of the process integrating the associated regulators, cAMP receptor protein and KdgR. The model describes the regulatory events occurring at the promoters of two major pel genes, pelE and pelD. It highlights that their activity is controlled by a mechanism of carbon catabolite repression, which directly controls the virulence of D. dadantii. The model also shows that quantitative differences in the binding properties of common regulators at these two promoters resulted in a qualitatively different role of pelD and pelE in the metabolic switch, and also likely in conditions of infection, justifying their evolutionary conservation as separate genes in this species.
Topics: Bacterial Proteins; Catabolite Repression; Dickeya; Digestion; Enterobacteriaceae; Gene Expression Regulation, Bacterial; Pectins; Polysaccharide-Lyases
PubMed: 34826421
DOI: 10.1016/j.jbc.2021.101446