N. Kozyrovska

Correlation between pectate lyase activity and ability of diazotrophic Klebsiella oxytoca VN 13 to penetrate into plant tissues

Diazotrophic Klebsiella oxytoca VN 13 was able to lyse pectate, but the lytic activity of cultures grown on non-selective media was weak, and the enzyme was located mainly inside the cells. A small fraction of the population (10-6 to 10-5) was able to grow in a selective medium with sodium polygalacturonate as sole carbon source, and produced increased amounts of the pectinolytic enzyme pectate lyase. When wheat seeds were inoculated with these bacteria, increased levels of this enzyme correlated with a higher rate of internal colonization of wheat roots and with stimulation of wheat growth resulting in higher biomass. This suggests that colonization occurs via lyzed pectin layers.

Identification of Klebsiella oxytoca using a specific PCR assay targeting the polygalacturonase pehX gene

Bacteria of the genus Klebsiella are important opportunistic pathogens responsible for nosocomial infections that are increasingly resistant to antimicrobial agents. Distinctive identification of the species K. oxytoca, K. pneumoniae, K. planticola, K. ornithinolytica and K. terrigena is difficult based on phenotypic tests and misidentifications are frequent in routine clinical microbiology. We developed a specific method to discriminate K. oxytoca from the other species of the genus Klebsiella, based on the PCR amplification of the polygalacturonase (pehX) gene. A PCR amplicon of 344 bp was obtained in all 35 K. oxytoca strains tested, but in none of the 29 K. pneumoniae, 12 K. planticola/K. ornithinolytica and 7 K. terrigena strains tested. The test was also negative for polygalacturonate-degrading species of the genus Erwinia. Analysis of 24 strains designated as K. pneumoniae from international collections (NCTC, PZH) revealed previous misidentification of six K. oxytoca strains. Key biochemical tests fully confirmed the pehX PCR results. The new K. oxytoca identification assay should be useful for both clinical and ecological monitoring of K. oxytoca strains, as well as for controlling the previous identification of collection strains.

Novel inoculants for an environmentally-friendly crop production

Abstract The high input of agrochemicals into soils is a major agricultural technique for enhancement of plant crop production, but it is also an environmental hazard. Concern over health hazards related to agrochemicals, especially on radioactively polluted territories, as well as economic problems, have promoted fundamental research to search for new agrobiotechnologies. Nonsymbiotic plant growth-promoting Rhizobacteria (PGPR) are often used as inoculants; however, they are not as effective as endophytic bacteria. These bacteria have the advantage of living within the plant tissue, protecting the plant from superinfection by soil bacteria, and recolonizing the plant surface after some stress situations in the soil. Although the use of beneficial microorganisms is not a new idea, it is an idea that is not heavily utilized. Application of inoculants designed on the base of competitive endophytic bacteria may be a full or partial alternative to agrochemicals, and can diminish the level of penetration and accumulation of heavy metals and radionuclides inside plants. This study evaluates the input of the association of the bacteria, isolated from the plant interior, in the crop of corn ( Zea mays ) and develops inoculants on their base. Nitrogen-fixing Klebsiella oxytoca VN13 along with Xanthomonas maltophilia VN12 were found to be useful bacteria, the former capable of excreting auxins and antimicrobial substances, the latter promoting assimilation of soil phosphorous by the plants. As the association of the two bacteria has been chosen as an effective endophytic system for the inoculant development, the two are mixed to form an inoculant ‘Duet’ which is directly inoculated into seeds. Corn inoculated with this ‘Duet’ produced greater yields, and possessed a greater percentage of protein. In addition, experiments performed in Chernobyl, showed that the ‘Duet’ could protect the plant from radionuclides penetration. It is suggested that a novel inoculant, ‘Kleps’, be applied, as the formulation is simple, it is inexpensive, it can be produced on a large scale, and it can be stored for a long period of time in a relatively small volume. It is clear that simple formulations of novel inoculants can be designed on the base of the competitive endophytic bacteria for an environmentally friendly crop production on poor and polluted territories as an alternative to the use of agrochemicals.Abstract The high input of agrochemicals into soils is a major agricultural technique for enhancement of plant crop production, but it is also an environmental hazard. Concern over health hazards related to agrochemicals, especially on radioactively polluted territories, as well as economic problems, have promoted fundamental research to search for new agrobiotechnologies. Nonsymbiotic plant growth-promoting Rhizobacteria (PGPR) are often used as inoculants; however, they are not as effective as endophytic bacteria. These bacteria have the advantage of living within the plant tissue, protecting the plant from superinfection by soil bacteria, and recolonizing the plant surface after some stress situations in the soil. Although the use of beneficial microorganisms is not a new idea, it is an idea that is not heavily utilized. Application of inoculants designed on the base of competitive endophytic bacteria may be a full or partial alternative to agrochemicals, and can diminish the level of penetration and accumulation of heavy metals and radionuclides inside plants. This study evaluates the input of the association of the bacteria, isolated from the plant interior, in the crop of corn ( Zea mays ) and develops inoculants on their base. Nitrogen-fixing Klebsiella oxytoca VN13 along with Xanthomonas maltophilia VN12 were found to be useful bacteria, the former capable of excreting auxins and antimicrobial substances, the latter promoting assimilation of soil phosphorous by the plants. As the association of the two bacteria has been chosen as an effective endophytic system for the inoculant development, the two are mixed to form an inoculant ‘Duet’ which is directly inoculated into seeds. Corn inoculated with this ‘Duet’ produced greater yields, and possessed a greater percentage of protein. In addition, experiments performed in Chernobyl, showed that the ‘Duet’ could protect the plant from radionuclides penetration. It is suggested that a novel inoculant, ‘Kleps’, be applied, as the formulation is simple, it is inexpensive, it can be produced on a large scale, and it can be stored for a long period of time in a relatively small volume. It is clear that simple formulations of novel inoculants can be designed on the base of the competitive endophytic bacteria for an environmentally friendly crop production on poor and polluted territories as an alternative to the use of agrochemicals.

Heavy Metal Resistance in Plants: A Putative Role of Endophytic Bacteria

Heavy metals and metalloids have become one of the major environmental concerns which pose a serious threat to plants and animal health. In this context, endophytic bacteria could play an important role in understanding the uptake mechanism of heavy metal ions and providing immunity to plant against metal toxicity. The defensive effects of certain elements in plants are known, but the role of endophytes in providing protection to plants has poorly been investigated. Endophytic bacteria, originating from hyperaccumulator plants, exhibit a comparatively higher level of resistance to heavy metals than the soil and the rhizosphere bacteria. Among bacteria, Methylobacterium spp., as well as the representatives of Gram-positive bacteria, are the most widespread bacterial species in both the hyperaccumulator endosphere and endorhizae. The endophytic microbial populations enhance the resistance capacity of the host plants, which, however, depends on the structure and activity of the community. Moreover, endophytic bacteria including those of legume endophytes are considered a promising biological material for improving the efficiency of phytoremediation and, consequently, growing of clean and safe crops including legumes in metal polluted soils. The recent developments in the putative mechanisms by which endophytic microorganisms affect the plant resistance to heavy metals and how they could affect phytoextraction of metals from contaminated soils are highlighted.