Biodegradation of low-density polyethylene by plasma-activated Bacillus strain.

Plastic biodegradation by microorganisms is an eco-friendly and sustainable method without any ramifications. Herein, we used a cultivation method and 16S rRNA sequencing to screen bacteria that can efficiently colonize and degrade low-density polyethylene (LDPE) from various plastic wastes. We identified Bacillus safensis BS-10L through whole-genome sequencing analysis and verified its LDPE-degradation ability. However, the decomposition mechanism of the isolated bacteria was unclear and the decomposition efficiency was insufficient, so low-temperature plasma was used to increase the decomposition efficiency of the bacteria. The population and viability of bacteria treated with cold plasma increased. Plasma-activated bacteria could induce cracks, holes, and roughness on the surface of LDPE films over 90 days, and over 30 days; the LDPE film lost 13.40 ± 0.013% and 27.78 ± 0.014% of its mass by BS-10L and plasma-treated BS-10L, respectively. Fourier-transform infrared spectroscopic analysis identified new peaks of the C=O and C-O groups in the plasma-treated LDPE film, exhibiting high transmittance in the LDPE film that was inoculated with bacteria. X-ray photoelectron spectroscopic analysis showed that C-O bonds were generated by BS-10L strain, and relatively strong C=O bonds were generated in the film inoculated with plasma-treated BS-10L strain. Plasma treatment increased the colonization of the BS-10L strain and changed the chemical bonding of the LDPE film, suggesting that plasma-activated BS-10L could accelerate decomposition by oxidation by increasing the carbonyl group of the PE film. Therefore, plasma technology may be effective for enhancing the plastic-degrading ability of microorganisms.

Enhancement of vitality and activity of a plant growth-promoting bacteria (PGPB) by atmospheric pressure non-thermal plasma.

The inconsistent vitality and efficiency of plant growth promoting bacteria (PGPB) are technical limitations in the application of PGPB as biofertilizer. To improve these disadvantages, we examined the potential of micro Dielectric Barrier Discharge (DBD) plasma to enhance the vitality and functional activity of a PGPB, Bacillus subtilis CB-R05. Bacterial multiplication and motility were increased after plasma treatment, and the level of a protein involved in cell division was elevated in plasma treated bacteria. Rice seeds inoculated with plasma treated bacteria showed no significant change in germination, but growth and grain yield of rice plants were significantly enhanced. Rice seedlings infected with plasma treated bacteria showed elevated tolerance to fungal infection. SEM analysis demonstrated that plasma treated bacteria colonized more densely in the broader area of rice plant roots than untreated bacteria. The level of IAA (Indole-3-Acetic Acid) and SA (Salicylic Acid) hormone was higher in rice plants infected with plasma treated than with untreated bacteria. Our results suggest that plasma can accelerate bacterial growth and motility, possibly by increasing the related gene expression, and the increased bacterial vitality improves colonization within plant roots and elevates the level of phytohormones, leading to the enhancement of plant growth, yield, and tolerance to disease.

Dynamics of nitric oxide level in liquids treated with microwave plasma-generated gas and their effects on spinach development.

In this study, we generated water and phosphate buffer treated with microwave plasma-generated gas in which the major component was nitric oxide (PGNO), and investigated the efficiency of the treated water and buffer in fertilization and sanitation. Real time NO level monitored by an electrode sensor was linearly increased over PGNO injection time, and removal of O2 from liquid before PGNO injection accelerated NO assimilation into liquids. Residual NO was still present 16 h after PGNO injection was stopped. H2O2, NO2-, and NO3- were also detected in PGNO-treated liquids. Spinach plants applied with 10 and 30 times diluted PGNO-treated water and 0.5 mM phosphate buffer showed slightly higher height and dry weight than control after 5 weeks. Plants grown with 10 and 30 times diluted PGNO-treated water exhibited the increased tolerance to water deficiency. Significant anti-microbial activity within 1 h was observed in un-diluted and in half-diluted PGNO-treated water and 0.5 mM phosphate buffer. Our results suggest that water or phosphate buffer containing NO, H2O2, NO2-, and NO3- can be produced by PGNO treatment, and that PGNO-treated water or buffer can be used as a potential fertilizer enhancing plant vitality with sanitation effect.

The Major Postharvest Disease of Onion and Its Control with Thymol Fumigation During Low-Temperature Storage.

Onion (Allium cepa L.) is one of the major vegetable crops in Korea that are damaged and lost by pathogenic fungal infection during storage due to a lack of proper storage conditions. The aim of this study was to determine an appropriate control measure using thymol to increase the shelf life of onions. To control fungal infections that occur during low-temperature storage, it is necessary to identify the predominant fungal pathogens that appear in low-temperature storage houses. Botrytis aclada was found to be the most predominant fungal pathogen during low-temperature storage. The antifungal activity of the plant essential oil thymol was tested and compared to that of the existing sulfur treatments. B. aclada growth was significantly inhibited up to 16 weeks with spray treatments using a thymol solution. To identify an appropriate method for treating onions in a low-temperature storage house, thymol was delivered by two fumigation treatment methods, either by heating it in the granule form or as a solution at low-temperature storage conditions (in vivo). We confirmed that the disease severity was reduced up to 96% by fumigating thymol solution compared to the untreated control. The efficacy of the fumigation of thymol solution was validated by testing onions in a low-temperature storage house in Muan, Jeollanam-do. Based on these results, the present study suggests that fumigation of the thymol solution as a natural preservative and fungicide can be used as an eco-friendly substitute for existing methods to control postharvest disease in long-term storage crops on a commercial scale.

Inactivation of Escherichia coli and Staphylococcus aureus on contaminated perilla leaves by Dielectric Barrier Discharge (DBD) plasma treatment.

This study focused on sterilization methods for the reduction of microorganisms on perilla leaves by cylinder type Dielectric Barrier Discharge (DBD) plasma with underwater bubbler treatment. S. aureus and E. coli in a suspension were reduced to less than 3.4 and 0.5 log CFU/ml after the plasma treatment for 3 min, respectively. On the perilla leaves, they were also reduced to 4.8 and 1.6 log CFU/ml after the plasma treatment, respectively. The S. aureus and E. coli bacterial cell wall was damaged by the plasma treatment evident by scanning electron microscopic analysis. The observed infrared bands of the FTIR spectra demonstrated changes in protein, lipid, polysaccharide, polyphosphate group and other carbohydrate functionalities of plasma treated bacteria and untreated bacterial cell membranes. The degradation of the constituent bonds of the bacterial cell membrane by RONS generated from plasma destroys the DNA, RNA, and proteins within the cell, and may eventually cause cell death. In this study, H2O2 (13.68 µM) and NO3 (138 µM), which are the main factors generated by plasma, proved to have a bactericidal effect by inducing lipid peroxidation of bacterial cell membranes. In conclusion, cylinder type DBD plasma with underwater bubbler can be used as an environmentally friendly food disinfection device in cleaning processes of the food industry.

Effects of high voltage nanosecond pulsed plasma and micro DBD plasma on seed germination, growth development and physiological activities in spinach.

In this study, we analyzed seed germination, seedling growth, and physiological aspects after treatment with high voltage nanosecond pulsed plasma and micro DBD plasma in spinach (Spinacia oleracea L.), a green leafy vegetable known to have low germination rate. Both germination and dry weight of seedlings increased after high voltage pulse shots were applied to spinach seeds. However seeds treated with many shots (10 shots) showed a decrease in germination rate and seedling growth. Seeds treated with air DBD plasma exhibited slightly higher germination and subsequent seedling growth than those treated with N2 plasma. Seed surface was degenerated after treated with high voltage pulsed plasma and micro DBD plasma but no significant difference in the degree of degeneration was observed among micro DBD plasma treatment time. Level of GA3 hormone and mRNA expression of an amylolytic enzyme-related gene in seeds were elevated 1 day after treatment with high voltage pulsed plasma. The relative amount of chlorophyll and total polyphenols in spinach seedlings grown from seeds treated with air DBD plasma was increased in 30 s, 1 min, and 3 min treatments. Taken together, our results suggest a possibility that plasma can enhance seed germination by triggering biochemical processes in seeds.

Expression analysis of rice pathogenesis-related proteins involved in stress response and endophytic colonization properties of gfp-tagged Bacillus subtilis CB-R05.

Bacillus subtilis CB-R05, possessing antagonistic effects against several fungal pathogens, is a diazotrophic plant growth-promoting bacteria marked with the green fluorescent protein (gfp) gene. To confirm the expression level of the pathogenesis-related (PR) proteins in rice inoculated with CB-R05, the expressions of four pathogenesis-related (PR) proteins (PR2, PR6, PR15, and PR16) were examined in the rice leaves treated with wounding stress over a time period. The PR proteins were generally more strongly expressed in the rice leaves inoculated with CB-R05 compared with the untreated control. The marked gfp-tagged B. subtilis CB-R05 strain was inoculated onto the rice seedlings under axenic conditions. Under the confocal laser scanning microscope (CLSM), the gfp-tagged CB-R05 bacterial cells were observed to penetrate the rhizoplane, especially in the elongation and differentiation zones of the rice roots, and colonize the root intracellularly. The bacteria, 24 h after the gfp-tagged CB-R05 inoculation, were seen to penetrate into the cell wall, cortex, xylem, and concentrate mainly in the vascular bundle. Numerous bacteria were observed within the intercellular spaces, root cortical cells, and xylem vessels. Over time, these bacteria dispersed to the lateral root junctions and propagated slowly from the roots to the stems and leaves. The B. subtilis CB-R05 population in the rice root rhizosphere was also monitored. These results show a very widespread colonization of the B. subtilis CB-R05 in the rice rhizosphere. Further attempts are under way to investigate the competition between the CB-R05 bacteria and the fungal pathogen in vivo.

Isolation and characterization of plant growth promoting endophytic diazotrophic bacteria from Korean rice cultivars.

We have isolated 576 endophytic bacteria from the leaves, stems, and roots of 10 rice cultivars and identified 12 of them as diazotrophic bacteria using a specific primer set of nif gene. Through 16S rDNA sequence analysis, nifH genes were confirmed in the two species of Penibacillus, three species of Microbacterium, three Bacillus species, and four species of Klebsiella. Rice seeds treated with these plant growth-promoting bacteria (PGPB) showed improved plant growth, increased height and dry weight and antagonistic effects against fungal pathogens. In addition, auxin and siderophore producing ability, and phosphate solubilizing activity were studied for the possible mechanisms of plant growth promotion. Among 12 isolates tested, 10 strains have shown higher auxin producing activity, 6 isolates were confirmed as strains with high siderophore producing activity while 4 isolates turned out to have high phosphate-solubilizing activity. These results strongly suggest that the endophytic diazotrophic bacteria characterized in this study could be successfully used to promote plant growth and inducing fungal resistance in plants.