The structure of an abequose - containing O-polysaccharide isolated from Pectobacterium aquaticum IFB5637.

Soft rot and blackleg diseases, caused by pectinolytic bacteria from the numerous species of Dickeya and Pectobacterium, pose a serious threat to the world potato production. Besides, infections triggered by these pectinolytic bacteria lead to huge economic losses in the cultivation of other crops, vegetables, and ornamentals. Strains belonging to the genus Pectobacterium tend to be isolated from various environments such as rotten or asymptomatic plants, weeds, soil or water. The main virulence factors of these phytopathogenic bacteria involve plant cell wall degrading enzymes (PCWDEs) i.e. pectinases, cellulases and proteases. Among accessory virulence factors, there is often lipopolysaccharide (LPS) listed. This constituent of the external part of bacterial cell wall contains lipid A, inner and outer core in addition to O-polysaccharide (OPS). LPS plays an important role in plant-microbe interactions, in particular during the first step of pathogen recognition. In this study we present the chemical structure of OPS of the first Pectobacterium aquaticum strain (IFB5637) isolated from water in Poland. The OPS consists of two common hexoses, such as mannose and glucose, as well as an abequose (3,6-dideoxy-d-xylo-hexose), the first 3,6-dideoxyhexose identified among the Pectobacteriaceae family: According to our best knowledge this is the first determined structure of the OPS of P. aquaticum.

Application of pulse-modulated radio-frequency atmospheric pressure glow discharge for degradation of doxycycline from a flowing liquid solution.

Doxycycline (DOX), an antibiotic commonly used in medicine and veterinary, is frequently detected in natural waterways. Exposition of bacteria to DOX residuals poses a selective pressure leading to a common occurrence of DOX-resistance genetic determinants among microorganisms, including virulent human pathogens. In view of diminishment of the available therapeutic options, we developed a continuous-flow reaction-discharge system generating pulse-modulated radio-frequency atmospheric pressure glow discharge (pm-rf-APGD) intended for DOX removal from liquid solutions. A Design of Experiment and a Response Surface Methodology were implemented in the optimisation procedure. The removal efficiency of DOX equalling 79 ± 4.5% and the resultant degradation products were identified by High-Performance Liquid Chromatography-Diode Array Detection, Liquid Chromatography Quadruple Time of Flight Mass Spectrometry, Ultraperformance Liquid Chromatography-Tandem Mass Spectrometry, total organic carbon, total nitrogen, Attenuated Total Reflectance Furrier Transform-Infrared, and UV/Vis-based methods. The pm-rf-APGD-treated DOX solution due to the generated Reactive Oxygen and Nitrogen Species either lost its antimicrobial properties towards Escherichia coli ATCC25922 or significantly decreased biocidal activities by 37% and 29% in relation to Staphylococcus haemolyticus ATCC29970 and Staphylococcus aureus ATCC25904, respectively. Future implementation of this efficient and eco-friendly antibiotic-degradation technology into wastewater purification systems is predicted.

Implementation of a Non-Thermal Atmospheric Pressure Plasma for Eradication of Plant Pathogens from a Surface of Economically Important Seeds.

Plant pathogenic bacteria cause significant economic losses in the global food production sector. To secure an adequate amount of high-quality nutrition for the growing human population, novel approaches need to be undertaken to combat plant disease-causing agents. As the currently available methods to eliminate bacterial phytopathogens are scarce, we evaluated the effectiveness and mechanism of action of a non-thermal atmospheric pressure plasma (NTAPP). It was ignited from a dielectric barrier discharge (DBD) operation in a plasma pencil, and applied for the first time for eradication of Dickeya and Pectobacterium spp., inoculated either on glass spheres or mung bean seeds. Furthermore, the impact of the DBD exposure on mung bean seeds germination and seedlings growth was estimated. The observed bacterial inactivation rates exceeded 3.07 logs. The two-minute DBD exposure stimulated by 3-4% the germination rate of mung bean seeds and by 13.4% subsequent early growth of the seedlings. On the contrary, a detrimental action of the four-minute DBD subjection on seed germination and early growth of the sprouts was noted shortly after the treatment. However, this effect was no longer observed or reduced to 9.7% after the 96 h incubation period. Due to the application of optical emission spectrometry (OES), transmission electron microscopy (TEM), and confocal laser scanning microscopy (CLSM), we found that the generated reactive oxygen and nitrogen species (RONS), i.e., N2, N2+, NO, OH, NH, and O, probably led to the denaturation and aggregation of DNA, proteins, and ribosomes. Furthermore, the cellular membrane disrupted, leading to an outflow of the cytoplasm from the DBD-exposed cells. This study suggests the potential applicability of NTAPPs as eco-friendly and innovative plant protection methods.

The First Polish Isolate of a Novel Species Pectobacterium aquaticum Originates from a Pomeranian Lake.

Pectinolytic bacteria from the genus Pectobacterium cause high economic losses in various crops, vegetables, and ornamentals including potato. Thus far, these strains have been isolated from distinct environments such as rotten or asymptomatic plants, soil, and waterways. The prevalence of soft rot Pectobacteriaceae in different depths of Pomeranian lakes was performed by a qualified scuba diver over 2 years of monitoring. It allowed for the isolation and broad characterization of a strain from the newly established species Pectobacterium aquaticum. Phylogenetic analysis on the sequences of dnaX and recA genes revealed the highest similarity of this strain to P. aquaticum CFBP 8637T. In addition to the determination of analytical profile index (API 20E), we discovered that this strain possesses a smooth form of a lipopolysaccharide with O-polysaccharide consisting of mannose, glucose, and abequose. Moreover, the characterized strain, described as P. aquaticum IFB5637, produced plant-cell-wall-degrading enzymes, such as pectinases, cellulases, proteases, and was capable of macerating potato and chicory tissues under laboratory conditions. In view of more frequent irrigation of seed potato fields resulting from the ongoing climate warming, it is important to monitor the occurrence of potential disease-causing agents in natural waterways.

Multivariate Optimization of the FLC-dc-APGD-Based Reaction-Discharge System for Continuous Production of a Plasma-Activated Liquid of Defined Physicochemical and Anti-Phytopathogenic Properties.

To the present day, no efficient plant protection method against economically important bacterial phytopathogens from the Pectobacteriaceae family has been implemented into agricultural practice. In this view, we have performed a multivariate optimization of the operating parameters of the reaction-discharge system, employing direct current atmospheric pressure glow discharge, generated in contact with a flowing liquid cathode (FLC-dc-APGD), for the production of a plasma-activated liquid (PAL) of defined physicochemical and anti-phytopathogenic properties. As a result, the effect of the operating parameters on the conductivity of PAL acquired under these conditions was assessed. The revealed optimal operating conditions, under which the PAL of the highest conductivity was obtained, were as follows: flow rate of the solution equaled 2.0 mL min-1, the discharge current was 30 mA, and the inorganic salt concentration (ammonium nitrate, NH4NO3) in the solution turned out to be 0.50% (m/w). The developed PAL exhibited bacteriostatic and bactericidal properties toward Dickeya solani IFB0099 and Pectobacterium atrosepticum IFB5103 strains, with minimal inhibitory and minimal bactericidal concentrations equaling 25%. After 24 h exposure to 25% PAL, 100% (1-2 × 106) of D. solani and P. atrosepticum cells lost viability. We attributed the antibacterial properties of PAL to the presence of deeply penetrating, reactive oxygen and nitrogen species (RONS), which were, in this case, OH, O, O3, H2O2, HO2, NH, N2, N2+, NO2-, NO3-, and NH4+. Putatively, the generated low-cost, eco-friendly, easy-to-store, and transport PAL, exhibiting the required antibacterial and physicochemical properties, may find numerous applications in the plant protection sector.

Antibacterial Activity of Fructose-Stabilized Silver Nanoparticles Produced by Direct Current Atmospheric Pressure Glow Discharge towards Quarantine Pests.

Development of efficient plant protection methods against bacterial phytopathogens subjected to compulsory control procedures under international legislation is of the highest concern having in mind expensiveness of enforced quarantine measures and threat of the infection spread in disease-free regions. In this study, fructose-stabilized silver nanoparticles (FRU-AgNPs) were produced using direct current atmospheric pressure glow discharge (dc-APGD) generated between the surface of a flowing liquid anode (FLA) solution and a pin-type tungsten cathode in a continuous flow reaction-discharge system. Resultant spherical and stable in time FRU-AgNPs exhibited average sizes of 14.9 ± 7.9 nm and 15.7 ± 2.0 nm, as assessed by transmission electron microscopy (TEM) and dynamic light scattering (DLS), respectively. Energy dispersive X-ray spectroscopy (EDX) analysis revealed that the obtained nanomaterial was composed of Ag while selected area electron diffraction (SAED) indicated that FRU-AgNPs had the face-centered cubic crystalline structure. The fabricated FRU-AgNPs show antibacterial properties against Erwinia amylovora, Clavibacter michiganensis, Ralstonia solanacearum, Xanthomonas campestris pv. campestris and Dickeya solani strains with minimal inhibitory concentrations (MICs) of 1.64 to 13.1 mg L-1 and minimal bactericidal concentrations (MBCs) from 3.29 to 26.3 mg L-1. Application of FRU-AgNPs might increase the repertoire of available control procedures against most devastating phytopathogens and as a result successfully limit their agricultural impact.

Application of Silver Nanostructures Synthesized by Cold Atmospheric Pressure Plasma for Inactivation of Bacterial Phytopathogens from the Genera Dickeya and Pectobacterium.

Pectinolytic bacteria are responsible for significant economic losses by causing diseases on numerous plants. New methods are required to control and limit their spread. One possibility is the application of silver nanoparticles (AgNPs) that exhibit well-established antibacterial properties. Here, we synthesized AgNPs, stabilized by pectins (PEC) or sodium dodecyl sulphate (SDS), using a direct current atmospheric pressure glow discharge (dc-APGD) generated in an open-to-air and continuous-flow reaction-discharge system. Characterization of the PEC-AgNPs and SDS-AgNPs with UV/Vis absorption spectroscopy, transmission electron microscopy, energy dispersive X-ray spectroscopy, and selected area electron diffraction revealed the production of spherical, well dispersed, and face cubic centered crystalline AgNPs, with average sizes of 9.33 ± 3.37 nm and 28.3 ± 11.7 nm, respectively. Attenuated total reflection-Fourier transformation infrared spectroscopy supported the functionalization of the nanostructures by PEC and SDS. Antibacterial activity of the AgNPs was tested against Dickeya spp. and Pectobacterium spp. strains. Both PEC-AgNPs and SDS-AgNPs displayed bactericidal activity against all of the tested isolates, with minimum inhibitory concentrations of 5.5 mg∙L-1 and 0.75-3 mg∙L-1, respectively. The collected results suggest that the dc-APGD reaction-discharge system can be applied for the production of defined AgNPs with strong antibacterial properties, which may be further applied in plant disease management.