Identification of the selected soil bacteria genera based on their geometric and dispersion features.

The visual analysis of microscopic images is often used for soil bacteria recognition in microbiology. Such task can be automated with the aid of machine learning and digital image processing techniques. The best results for soil microorganism identification usually rely on extracting features based on color. However, accommodating in the latter an extra impact of lighting conditions or sample's preparation on classification accuracy is often omitted. In contrast, this research examines features which are insensitive to the above two factors by focusing rather on bacteria shape and their specific group dispersion. In doing so, the calculation of layout features resorts to k-means and mean shift methods. Additionally, the dependencies between specific distances determined from bacteria cells and the curvature of interpolated bacteria boundary are computed to extract vital geometric shape information. The proposed bacteria recognition tool involves testing four different classification methods for which the parameters are iteratively adjusted. The results obtained here for five selected soil bacteria genera: Enterobacter, Rhizobium, Pantoea, Bradyrhizobium and Pseudomonas reach 85.14% classification accuracy upon combining both geometric and dispersion features. The latter forms a promising result as a substitutive tool for color-based feature classification.

Agrobacterium cucumeris sp. nov. isolated from crazy roots on cucumber (Cucumis sativus).

Three plant rhizogenic strains O132T, O115 and O34 isolated from Cucumis sp. L. were assessed for taxonomic affiliation by using polyphasic taxonomic methods. Based on the results of the sequence analysis of the 16S rRNA and multilocus sequence analysis (MLSA) of the three housekeeping genes atpD, recA and rpoB, all the strains were clustered within the genus Agrobacterium where they form a novel branch. Their closest relative was Agrobacterium tomkonis (genomospecies G3). Moreover, digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) comparisons between strains O132T and O34 and their closest relatives provided evidence that they constitute a new species, because the obtained values were significantly below the threshold considered as a borderline for the species delineation. Whole-genome phylogenomic analysis also indicated that the cucumber strains are located within the separate, well-delineated biovar 1 sub-clade of the genus Agrobacterium. Furthermore, the physiological and biochemical properties of these strains allowed to distinguish them from their closest related species of the genus Agrobacterium. As a result of the performed overall characterization, we propose a new species as Agrobacterium cucumeris sp. nov., with O132T (=CFBP 8997T = LMG 32451T) as the type strain.

Microbial-Based Products to Control Soil-Borne Pathogens: Methods to Improve Efficacy and to Assess Impacts on Microbiome.

Microbial-based products (either as biopesticide or biofertilizers) have a long history of application, though their use is still limited, mainly due to a perceived low and inconsistent efficacy under field conditions. However, their efficacy has always been compared to chemical products, which have a completely different mechanism of action and production process, following the chemical paradigm of agricultural production. This paradigm has also been applied to regulatory processes, particularly for biopesticides, making the marketing of microbial-based formulations difficult. Increased knowledge about bioinocula behavior after application to the soil and their impact on soil microbiome should foster better exploitation of microbial-based products in a complex environment such as the soil. Moreover, the multifunctional capacity of microbial strains with regard to plant growth promotion and protection should also be considered in this respect. Therefore, the methods utilized for these studies are key to improving the knowledge and understanding of microbial-based product activity and improving their efficacy, which, from farmers' point of view, is the parameter to assess the usefulness of a treatment. In this review, we are thus addressing aspects related to the production and formulation process, highlighting the methods that can be used to evaluate the functioning and impact of microbial-based products on soil microbiome, as tools supporting their use and marketing.

Agrobacterium vaccinii sp. nov. isolated from galls on blueberry plants (Vaccinium corymbosum).

Four non-pathogenic strains isolated from the galls on blueberry plants (Vaccinium corymbosum) were characterized by using polyphasic taxonomic methods. Based on 16S rRNA gene phylogeny, strains were clustered within the genus Agrobacterium. Furthermore, multilocus sequence analysis (MLSA) based on the partial sequences of atpD, recA and rpoB housekeeping genes and whole-genome-based phylogeny indicated that the strains studied form a novel Agrobacterium species. Analyses showed that the strains belong to "rubi" sub-clade of Agrobacterium genus and their closest relatives are Agrobacterium rubi and "Agrobacterium bohemicum". Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) comparisons between genome sequences of representative strains B7.6T and B19.1.4, and their closest relatives, confirmed the distinct phylogenetic position of studied strains, because obtained values were considerably below the proposed thresholds for the species delineation. The four strains studied were phenotypically distinguishable from other species of the genus Agrobacterium. Overall, polyphasic characterization showed that the strains studied represent a novel species of the genus Agrobacterium, for which the name Agrobacterium vaccinii sp. nov. is proposed. The type strain of A. vaccinii is B7.6T (=CFBP 8740T = LMG 31849T).

The Use of Interactions Between Microorganisms in Strawberry Cultivation (Fragaria x ananassa Duch.).

As the market indicates a growing interest in organically grown fruit, there is a need for biostimulants to counter the adverse effects of pathogenic fungi and fungal-like-pathogens. Four microbial pathogens (Botrytis cinerea, Verticillium sp., Phytophthora sp., and Colletotrichum sp.) which are the most often causes of strawberry diseases were selected. Five kinds of biostimulants (C1, C2, C3, C4, and C5) containing bacterial consortia were developed to combat the pathogens. The antagonistic effect of selected microorganisms against strawberry pathogens was observed. The effectiveness of various beneficial bacteria in combating fungal pathogens of cv. Honeoye strawberries was compared and the impact of their activity on fruit quality was assessed. The most significant effect on the strawberry firmness was found for the C2 consortium, which provided the strawberries infected with the pathogens group (MIX: B. cinerea, Verticillium sp., Phytophthora sp., and Colletotrichum sp.) with a 140% increase in maximum load in a puncture test compared to the positive control (C0). Strawberries contaminated with Phytophthora sp. after the application of Consortium C4 (C4) showed the largest increase (127%) in soluble solid content (SSC) when compared to the C0. Fruit contaminated with Colletotrichum sp. and B. cinerea after the application of C2 and Consortium 5 (C5), respectively, had the highest levels of anthocyanins and total phenolic content, when compared to C0. The largest increase, which reached as high as 25%, in D-galacturonic acid content was observed for the group of pathogens after Consortium 1 (C1) application. The extraction of strawberry pectin allowed for the study of the rheological properties of pectin solutions; on this basis, strawberry pectin from the control (NC) was distinguished as it showed the highest viscosity (0.137-0.415 Pas). Taking into account the individual effects of bacteria on strawberry pathogenic fungi and fungal-like-pathogens, it is possible to reduce the adverse effects of fungal disease and to improve the properties of strawberries by selecting the appropriate bacterial consortium. Interactions between microorganisms are often complex and not fully understood, which suggests the need for further research in this direction.

Fermentation Strategies to Improve Soil Bio-Inoculant Production and Quality.

The application of plant beneficial microorganisms has been widely accepted as an efficient alternative to chemical fertilizers and pesticides. Isolation and selection of efficient microorganisms, their characterization and testing in soil-plant systems are well studied. However, the production stage and formulation of the final products are not in the focus of the research, which affects the achievement of stable and consistent results in the field. Recent analysis of the field of plant beneficial microorganisms suggests a more integrated view on soil inoculants with a special emphasis on the inoculant production process, including fermentation, formulation, processes, and additives. This mini-review describes the different groups of fermentation processes and their characteristics, bearing in mind different factors, both nutritional and operational, which affect the biomass/spores yield and microbial metabolite activity. The characteristics of the final products of fermentation process optimization strategies determine further steps of development of the microbial inoculants. Submerged liquid and solid-state fermentation processes, fed-batch operations, immobilized cell systems, and production of arbuscular mycorrhiza are presented and their advantages and disadvantages are discussed. Recommendations for further development of the fermentation strategies for biofertilizer production are also considered.

Integrated multi-channel optical system for bacteria characterization and its potential use for monitoring of environmental bacteria.

The potential use of a novel multichannel optical system towards fast and non-destructive bacteria identification and its application for environmental bacteria characterisation on the strain level is presented. It is the first attempt to use the proposed optical method to study various bacteria species (Gram-negative, Gram-positive) commonly present in the environment. The novel configuration of the optical system enables multichannel examination of bacterial colonies and provides additional functionality such as registration of two-dimensional (2D) distribution of monochromatic transmission coefficient of examined colonies, what can be used as a novel optical signature for bacteria characterization. Performed statistical analysis indicates that it is possible to identify representatives of environmental soil bacteria on the species level with the 98.51% accuracy and in case of two strains of Rahnella aquatilis bacteria on the strain level with the 98.8% accuracy. The proposed method is an alternative to the currently used preliminary bacteria examination in environment safety control with the advantage of being fast, reliable, non-destructive and requiring minimal sample preparation.

Microbial Products and Biofertilizers in Improving Growth and Productivity of Apple - a Review.

The excessive use of mineral fertilizers causes many negative consequences for the environment as well as potentially dangerous effects of chemical residues in plant tissues on the health of human and animal consumers. Bio-fertilizers are formulations of beneficial microorganisms, which upon application can increase the availability of nutrients by their biological activity and help to improve soil health. Microbes involved in the formulation of bio-fertilizers not only mobilize N and P but mediate the process of producing crops and foods naturally. This method avoids the use of synthetic chemical fertilizers and genetically modified organisms to influence the growth of crops. In addition to their role in enhancing the growth of the plants, biofertilizers can act as biocontrol agents in the rhizosphere at the same time. Biofertilizers are very safe for human, animal and environment. The use of Azotobacter, Azospirillum, Pseudomonas, Acetobacter, Burkholderia, Bacillus, Paenibacillus and some members of the Enterobacteriaceae is gaining worldwide importance and acceptance and appears to be the trend for the future.

Identification and assessment of genetic similarity of soil bacterial isolates of Pseudomonas spp. using molecular techniques.

Bacteria of the genus Pseudomonas are often components of bioproducts designed to enhance the condition of the soil and plants. The use of Pseudomonas bacteria in bioproducts must be preceded by the acquisition, characterization and selection of beneficial strains living in the soil. A prerequisite for the selection of bacterial strains for use in bioproducts is to be able to identify the isolates rapidly and accurately. To identify and differentiate 15 bacterial isolates obtained from the soil surrounding the roots of sour cherry trees and to assess their genetic similarity, the rep-PCR technique and restriction analysis of the 16S rRNA gene and the 16S-ITS-23S rRNA operon were used. In addition, a sequence analysis of the 16S rRNA gene was performed. The analyses made it possible to divide the isolates into four clusters and to confirm their affiliation with the Pseudomonas species. RFLP analysis of the 16S-ITS-23S rRNA operon enabled greater differentiation of the isolates than RFLP of the 16S rRNA gene. The greatest differentiation of isolates within the clusters was obtained after using the rep-PCR technique. However, none of the techniques was able to discriminate all the isolates, which indicates very high genetic similarity of the Pseudomonas isolates found in the same sample of soil from around the roots of sour cherry trees. The tests performed will find application for distinguishing and identifying Pseudomonas strains collected from the soil in order to select the most valuable bacterial strains that produce beneficial effects on plants.