Antagonistic spectrum of action of saprophite microorganisms on phytopathogenic bacteria

. The antagonistic activity of 41 strains of various physiological groups of saprophytic soil spore-bearing and non-spore-bearing microorganisms to seven species of phytopathogenic bacteria was studied. It was found that out of 40 strains, only 10 did not have this property, the remaining strains showed an antimicrobial spectrum to phytopathogenic bacteria, but their spectrum of action was not the same. It is known that representatives of useful soil microflora are nitrogen-fixing microorganisms capable of fixing molecular nitrogen, as well as contributing to the enrichment of nitrogen forms digestible for plants, thereby increasing soil fertility. Nitrogen-fixing microorganisms include free-living bacteria of the genus Azotobacter , which most intensively develop in the rhizosphere zone of plant roots and on their surface, as well as symbiotic nodule bacteria of the genus Rhizobium penetrating into the root system of leguminous plants and forming nodules on the bottom. Studies have been carried out to study the effect of metabolites in the studied antagonists on the survival of Azotobacter and nodule bacteria. It has been established that the culture liquid of the studied antagonists does not have a detrimental effect on the studied bacteria of the genus Azotobacter and the genus Rhizobium .


Introduction
One of the most important tasks of raising and increasing agricultural production is to find measures to combat plant diseases [1].At present, due to the growing danger of the negative impact of the applied chemical plant protection products on human health and the environment, the attention of scientists is directed to the search for biological means of combating phytopathogens [2].
An evaluation of recent results on the use of biological agents to control phytopathogens by microorganisms shows that biological methods have a great future in agriculture in terms of environmental protection, replacing pesticides currently used to protect crops [3].
The essence of biological control methods is the use of other microorganisms against pathogens, as well as their metabolic products, or the use of antagonistic relationships between pathogens and saprophytic microorganisms [4].
Studies in the field of using microbes -antagonists and products of their metabolism as a biological control against phytopathogenic microorganisms indicate the prospects for searching for antagonistic microorganisms and studying their effect on phytopathogenic microorganisms [5][6][7][8].
In this regard, the main goal of this work was to find active microorganisms that inhibit the development of some phytopathogenic bacteria and to study their effect on representatives of beneficial soil microflora.

Materials and methods
The objects of research were 41 strains of microbes -antagonists and 7 species of phytopathogenic bacteria.The studied microorganisms were kindly presented to us by the Institute of Microbiology of the Academy of Sciences of Uzbekistan, the Department of Agricultural Phytopathology and Agrobiotechnology of Tashkent State Agrarian University, the Department of Microbiology of National University of Uzbekistan.
The study of the antagonistic properties of microorganisms to phytopathogenic bacteria was carried out [9][10][11].
Phytopathogenic bacteria included the following species [12] Antagonist bacteria were grown on peptone agar for 48 hours, then antagonist agar blocks were placed on freshly seeded zones of test cultures.The growth inhibition of phytopathogenic microorganisms around the agar blocks was determined 48 hours after the start of the experiment.Determination of the antimicrobial activity of antagonists on the survival of beneficial soil microflora (azotobacter and nodule bacteria) was carried out using the cylinder (block) method [13].
For this purpose, the culture liquid of the latter was centrifuged for 20 minutes at 8000 rpm, then the sedimentary liquid obtained was filtered through a Seitz filter using a GF/A glass fiber bacterial filter (Whatman Jiternational ltd, England) and a drop of cell suspension was applied to the surface of the nutrient medium [14].Test microorganism and evenly rub with a spatula over the entire surface.After that, 0.1 ml of the antagonist filtrate is applied with a sterile pipette into the previously made agar hole with a diameter of 0.5.The Petri dishes inoculated in this way were placed in a thermostat for 48 hours at a temperature of 28 0 C.After 48 hours, the Petri dishes were inspected and the zones of absence of growth of bacteria of the genus Azotobacter and Rhizobium were measured.

Results and discussion
To identify antagonistic properties, the studied microorganisms were grown on glucose agar, and the culture test was grown on potato agar.The growth retardation of phytopathogenic bacteria was determined after 48 hours.
The results obtained on the effect of antagonist microorganisms on phytopathogenic bacteria are presented in Table 1.From the data obtained, it can be seen that not all microorganisms have an antimicrobial spectrum in relation to phytopathogenic bacteria.1).
It is known that the Bacillus genus combines spore-forming bacteria that are highly viable and live in the soil as saprophytes, while members of the Pseudomonas genus are non-spore-forming bacteria that can live in the soil and protect plants from pathogenic microorganisms.Bacteria of the genus Pseudomonas sp are of particular interest as antagonists of many phytopathogenic bacteria of the genus Erwinia, Pseudomonas, Xanthomonas and Agrobacterium.
Thus, the study of the antimicrobial effect of the studied microorganisms to 7 species of phytopathogenic bacteria showed that out of 41 species of soil bacteria, only 10 did not have this property.The rest of the strains showed an antimicrobial spectrum of action against phytopathogens, but their spectrum of action and antibiotic activity were not the same.
Data on the antibiotic activity of microorganisms indicate the possibility of using some of them to limit the infection caused by phytopathogenic bacteria.Based on the data obtained, it can be concluded that antagonism in the studied microorganisms in relation to phytopathogenic bacteria was observed actively in some cases, and weakly in others.
Currently, there is information in the literature that in soils where antagonists develop, microbes sensitive to them develop weaker or do not develop at all.It is known that representatives of useful soil microflora are nitrogen-fixing microorganisms capable of fixing molecular nitrogen, as well as contributing to the enrichment of nitrogen forms digestible for plants, thereby increasing soil fertility.Nitrogen-fixing microorganisms include free-living bacteria of the genus Azotobacter, which most intensively develop in the rhizosphere zone of plant roots and on their surface, as well as symbiotic nodule bacteria of the genus Rhizobium penetrating into the root system of leguminous plants and forming nodules on the bottom.A number of researchers who have studied the effect of antagonists on microorganisms indicate that soil antagonists can greatly affect the abundance of Azotobacter and nodule bacteria.
Sharma et al. [11] studied the in vitro interaction of microorganisms and two species of azotobacter (A.vinelandii and Chrococcum) isolated from the wheat rhizosphere.Experiments have established that bacteria Ps. putida, Bac.subtilis, Xanthomonas oryzae suppressed the germination of cysts Azotobacter sp.
Shumilina et al. [14], studying the interaction of antagonist Ps.mycophaga with some soil microorganisms Azotobacter, nodule and phosphorus bacteria, found that the antagonist did not inhibit the bacteria used.
Thus, the literature data indicate that some soil antagonists have a detrimental effect on beneficial microflora, while others have no effect.In this regard, we were interested in studying the effect of metabolites in the studied antagonists on the survival of Azotobacter and nodule bacteria.Our studies on the study of antagonistic properties in the studied bacteria showed that the Ps.fluorescens culture has the highest antagonistic activity and a wide spectrum of action.
We studied the effect of metabolites Ps.fluorescens, B.subtilies 23 and B.megaterium 26 on the survival of beneficial microflora, which occurs in the soil and plays an important role in plant nutrition processes.Bacteria of the genus Rhizobium and Azotobacter were used for the work.
The study of the effect of culture liquid Ps.fluorescens, B.subtilies and B.megaterium on the survival of beneficial microflora -Azotobacter and nodule bacteria was carried out by the method of cylinders (blocks).
The results of the studies showed that all the studied antagonists only in some variants had a weak antibiotic activity in relation to nitrogen fixers (Table 2).Azotobacter chroococcum 19 8±2.5 --
A.agila 7 ---Thus, bacterium B.subtilis 23 had weak antagonistic activity against only nodule bacterium R.trifolii 329 Ps.fluorescens (41) and showed antimicrobial activity against two nitrogen fixers of nodule bacteria R.meliloti D and free-living bacteria R.meliloti D.

Table 1 .
Antagonistic spectrum of the studied microorganisms to phytopathogenic bacteria.

Table 2 .
Effect of antagonists on nitrogen-fixing bacteria.