Genome Analysis of Two Pseudomonas syringae pv. aptata Strains with Different Virulence Capacity Isolated from Sugar Beet: Features of Successful Pathogenicity in the Phyllosphere Microbiome

Members of the Pseudomonas syringae species complex are heterogeneous bacteria that are the most abundant bacterial plant pathogens in the plant phyllosphere, with strong abilities to exist on and infect different plant hosts and survive in/outside agroecosystems. In this study, the draft genome sequences of two pathogenic P. syringae pv. aptata strains with different in planta virulence capacities isolated from the phyllosphere of infected sugar beet were analyzed to evaluate putative features of survival strategies and to determine the pathogenic potential of the strains. The draft genomes of P. syringae pv. aptata strains P16 and P21 are 5,974,057 bp and 6,353,752 bp in size, have GC contents of 59.03% and 58.77%, respectively, and contain 3,439 and 3,536 protein-coding sequences, respectively. For both average nucleotide identity and pangenome analysis, P16 and P21 largely clustered with other pv. aptata strains from the same isolation source. We found differences in the repertoire of effectors of the type III secretion system among all 102 selected strains, suggesting that the type III secretion system is a critical factor in the different virulent phenotypes of P. syringae pv. aptata. During genome analysis of the highly virulent strain P21, we discovered genes for T3SS effectors (AvrRpm1, HopAW1, and HopAU1) that were not previously found in genomes of P. syringae pv. aptata. We also identified coding sequences for pantothenate kinase, VapC endonuclease, phospholipase, and pectate lyase in both genomes, which may represent novel effectors of the type III secretion system. IMPORTANCE Genome analysis has an enormous effect on understanding the life strategies of plant pathogens. Comparing similarities with pathogens involved in other epidemics could elucidate the pathogen life cycle when a new outbreak happens. This study represents the first in-depth genome analysis of Pseudomonas syringae pv. aptata, the causative agent of leaf spot disease of sugar beet. Despite the increasing number of disease reports in recent years worldwide, there is still a lack of information about the genomic features, epidemiology, and pathogenic life strategies of this particular pathogen. Our findings provide advances in disease etiology (especially T3SS effector repertoire) and elucidate the role of environmental adaptations required for prevalence in the pathobiome of the sugar beet. From the perspective of the very heterogeneous P. syringae species complex, this type of analysis has specific importance in reporting the characteristics of individual strains

Occurrence of plant pathogenic Pseudomonas syringae in the Danube River Basin: abundance and diversity assessment

Abstract Plant pathogenic strains of Pseudomonas syringae (Psy) spp. have been detected in nonagricultural habitats, including those associated with the water cycle. Their presence in aquatic systems allows dissemination over long distances, especially with irrigation practices. In May 2021, we sampled 15 sites along the Danube River Basin in Serbia to gain insight into P. syringae abundance and diversity. We identified 79 Psy strains using Psy-specific primers, and a partial sequence of the citrate synthase (cts) house-keeping gene has served for phylogenetic diversity assessments. Phenotypic diversity determination included characterizing features linked with survival and pathogenic lifestyle. The ice nucleation activity, pectinolytic activity, swimming and swarming assays, and hypersensitive reaction on plants were tested. Psy was detected at ten of 15 sites examined at abundance ranging from 1.0 × 102 to 1.2 × 104 CFU/L. We discovered the presence of four phylogroups, with phylogroup 2 being the most abundant, followed by phylogroups 7, 9, and 13. The hypersensitive reaction was induced by 68.63% of the isolates from the collection. A partial sequence comparison of the cts gene showed 100% similarity between isolates from cherry plants epidemics in Serbia caused by Psy and isolates from the Danube River. Our results suggest that the Danube River, extensively used for irrigation of agricultural fields, harbors diverse strains of Psy, which possess various features that could lead to potential disease outbreaks on crops. This study represents the first in-depth analysis of Psy abundance and diversity in the Danube River Basin. It sets the ground for future pre-epidemic studies and seasonal monitoring of Psy population dynamics

Biological control of major pathogenic bacteria of potato by Bacillus amyloliquefaciens strains SS-12.6 and SS-38.4

Potato is ranked as one of the most important food crops. Bacterial wilt caused by Ralstonia solanacearum and blackleg and soft rot caused by different species from genera Pectobacterium and Dickeya are considered two of the most important diseases of the potato. Biological control is the optimal strategy for controlling pathogens in crops generally, including bacteria. The present study aimed to evaluate the antagonistic effects of two Bacillus amyloliquefaciens strains, SS-12.6 and SS-38.4, against bacterial pathogens isolated from the potato in Serbia, such as R. solanacearum, Pectobacterium carotovorum, Pectobacterium brasiliense, and Dickeya dianthicola. The diameter of the inhibition zones formed by ethyl-acetate extracts of SS-12.6 and SS-38.4 strains show much higher values than the inhibition zones of supernatants, which implies the main power of these antagonists' potential lies in lipopeptides. The effectiveness of the treatment (19.7–44.5%), based on the difference in weight of potato tubers on the fifth and 15th day after treatment (DAT), showed that the antagonistic strains were almost equally effective in the suppression of P. carotovorum, P. brasiliense, and D. dianthicola strains. Strains SS-12.6 and SS-38.4 exhibited the efficacy in the suppression of R. solanacearum wilt from 28.64 to 60.22%. The analysis of the area under the disease progress (AUDPC) confirmed differences among pathogen control treatments and biocontrol treatments with B. amyloliquefaciens strains in all trials. This study shows that the two B. amyloliquefaciens strains, SS-12.6 and SS-38.4, can potentially be used as biocontrol agents against potato pathogens

Suppression of <i>Monilinia</i> Brown Rot by <i>Bacillus</i> spp. Strains

Brown rot caused by Monilinia spp. is one of the main causes of pre- and postharvest losses in stone and pome fruit production. The use of beneficial microorganisms is considered one of the most promising, safe and effective alternative methods for controlling these pathogens. This study aimed to investigate the antagonistic potential of 33 Bacillus spp. strains, in order to identify the best candidate for brown rot biocontrol. Strains identified as Bacillus amyloliquefaciens B-241 and Bacillus subtilis B-313 and B-358 were chosen for further ex situ studies on detached apple fruit. The efficacy of B-241 (87.1–93.7%) did not differ significantly from a commercially available synthetic fungicide (p > 0.05). The putative mode of action of B. amyloliquefaciens B-241 against Monilinia species is competition for nutrients and antibiosis. The ethyl acetate extract of the strain, applied at 5 and 12.5 mg/mL, was bioactive in vitro and ex situ. A HPLC analysis confirmed the presence of surfactin and bacillomycin D in the extract. However, before developing a shelf-stable product and commercial production, the spectrum and efficacy on a larger scale of the B-241 strain should be determined, and its efficacy in combination with commercial biofungicides and fungicides tested in vivo

Spatio-Temporal Dynamics in Physico-Chemical Properties, Phytoplankton and Bacterial Diversity as an Indication of the Bovan Reservoir Water Quality

The study aimed to investigate the physico-chemical properties as well as phytoplankton and bacterial community diversity of Bovan Lake reservoir in Serbia to gain insight into the seasonal dynamic of water quality. All analyses were performed at three localities and water depths in spring, summer, autumn, and winter 2019. Seven phytoplankton phyla comprising 139 taxa were detected at all three localities (Chlorophyta 58%, Bacillariophyta 14%, Cyanobacteria 9%, Chrysophyta 5%, Dinophyta 5%, Euglenophyta 5%, and Cryptophyta 4%). Winter 2019 was characterized by the presence of 36 unique species in all phyla except Euglenophyta. Bacterial diversity analyses showed that Proteobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, and Verrucomicrobia dominated the water intake locality at all three water depths (0.5, 10, and 20 m below the surface). In general, the physico-chemical parameters, phytoplankton, and bacterial community composition depended on the season and the water depth and showed that Bovan Lake was of satisfactory ecological status and water quality at all localities, and meets the needs for all intended purposes