Potential of root nodule nonrhizobial endophytic bacteria for growth promotion of Lotus corniculatus L. and Dactylis glomerata L.

AIMS: This research aimed to isolate and characterize nonrhizobial endophytic bacteria from root nodules of Medicago sativa L. and Lotus corniculatus L. with plant growth-promoting characteristics and to test its activity in a pot experiment with acid soil. METHODS AND RESULTS: Out of 44 nonrhizobial isolates, the majority exhibited indole-3-acetic acid (IAA) production; 29 produced siderophores, few isolates performed phosphate solubilization and/or produced lytic enzymes, while 30% of isolates showed notable antifungal activity. The most promising strains were identified as members of Bacillus, Pseudomonas and Serratia genera, based on 16S rRNA. Bacillus megaterium DZK1BH exhibited the overall best attributes for plant growth promotion and positively influenced the growth of L. corniculatus and Dactylis glomerata. CONCLUSIONS: Root nodule endophytic B. megaterium DZK1BH could potentially be used as a biofertilizer for growing L. corniculatus L. and D. glomerata L. in acid soils, while Bacillus mojavensis L3 is a candidate for further antifungal potential investigation. SIGNIFICANCE OF IMPACT OF THE STUDY: The use of root nodule endophytic bacteria with PGP traits may find its future application in organic agriculture, as their utilization could decrease the use of chemical fertilizers and pesticides and simultaneously promote plant growth, especially in soils with low production quality.

Phenolic composition and biological activities of geographically different type of propolis and black cottonwood resins against oral streptococci, vaginal microbiota and phytopathogenic Fusarium species.

AIMS: A multidisciplinary approach was used to compare phenolic composition, radical scavenging and antimicrobial activity of propolis samples from different geographical localities, and plant resin against various microorganisms. METHODS AND RESULTS: Using UHPLC-qqqMS quantitative analysis, 28 phenolic compounds were determined. Caffeic and p-coumaric acids were identified as main phenolic acids in poplar propolis samples, except samples from Russia (P6) and China (P7). Radical scavenging activity (applying DPPH spectrophotometric assay) showed the highest activity of Serbian (40·51%) and Chinese (53·21%) propolis samples. Broth microdilution method was used for the oral cavity, fungal phytopathogenic and human vaginal isolates which have been identified at a molecular level. The most sensitive bacterial isolates were Lactobacillus acidophilus (MIC of 0·03-0·13 mg ml-1 ) and the oral streptococci isolates (MIC values of 0·19-0·13 mg ml-1 ). The most sensitive fungal phytopathogenic isolate was Fusarium oxysporum (MIC 0·003 mg ml-1 ). All samples, except propolis from Serbia (P4) and Turkey (P5), showed a strong antifungal activity against Fusarium sporotrichioides, Fusarium subglutinans and Fusarium proliferatum. CONCLUSION: The results of various tests indicate good radical scavenging and antimicrobial activity against important human and plant pathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: A detailed propolis analysis is important when proposing a preparation of new biological antimicrobial products which have a positive impact on human health and reduce antibacterial resistance.

Bacterial communities of plum phyllosphere and characterization of indigenous antagonistic Bacillus thuringiensis R3/3 isolate.

AIMS: The characterization of bacterial communities diversity on four local plum cultivars in two phenological stages using culture-dependent and culture-independent methods and screening among culturable plum community for indigenous bacteria active against phytopathogens. METHODS AND RESULTS: The bacterial communities associated with leaves and fruits of four local Serbian plum cultivars (Pozegaca, Ranka, Cacanska Lepotica and Cacanska Rodna) were investigated in two phenological stages during early (May) and late (July) fruit maturation. Metagenomic approach revealed Methylobacterium, Sphingomonas and Hymenobacter as dominant genera. The most frequently isolated representatives with cultivable approach were pseudomonads with Pseudomonas syringae and Pseudomonas graminis, the most likely resident species of plum community. Antagonistic Bacillus thuringiensis R3/3 isolate from plum phyllosphere had ability to produce exoenzymes, reduce the growth of phytopathogenic bacteria in co-culture environment and show quorum quenching activity. CONCLUSIONS: Plum cultivar and growth season contribute to the structure of the bacterial community associated with plum. Plum phyllosphere is good source of antagonists effective against phytopathogens. SIGNIFICANCE AND IMPACT OF STUDY: Knowledge of bacterial communities on plum will have an impact on studies related to phyllosphere ecology and biocontrol. The indigenous antagonistic isolate, B. thuringiensis R3/3, from plum could be further investigated for its potential use in biological control of plum diseases.

Biological control of Pseudomonas syringae pv. aptata on sugar beet with Bacillus pumilus SS-10.7 and Bacillus amyloliquefaciens (SS-12.6 and SS-38.4) strains.

AIM: Assessment of biological control of Pseudomonas syringae pv. aptata using crude lipopeptide extracts (CLEs) of two Bacillus amyloliquefaciens strains (SS-12.6 and SS-38.4) and one Bacillus pumilus strain (SS-10.7). METHODS AND RESULTS: The minimum inhibitory concentration (MIC) of CLEs and their combinations against the pathogen and potential interaction between the extracts were determined in vitro. The most effective antibacterial activity was achieved with the CLE from B. amyloliquefaciens SS-12.6, with an MIC value of 0·63 mg ml-1 . Interactions between CLE combinations were mostly indifferent. The biocontrol potential of CLEs, mixtures of CLEs, and cell culture of B. amyloliquefaciens SS-12.6 was tested on sugar beet plants inoculated with P. syringae pv. aptata P53. The best result in inhibiting the appearance of tissue necrosis (up to 92%) was achieved with B. amyloliquefaciens SS-12.6 cell culture. CONCLUSION: This work demonstrated significant biocontrol potential of the CLE and cell culture of B. amyloliquefaciens SS-12.6 which successfully suppress leaf spot disease severity on sugar beet plants. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of biocontrol of sugar beet emerging pathogen will contribute to growers in terms of alternative disease control management. This study represents first assessment of biological control of P. syringae pv. aptata.

Microbiota associated with pollen, bee bread, larvae and adults of solitary bee Osmia cornuta (Hymenoptera: Megachilidae).

Using cultivation-dependant method, we isolated 184 strains from fresh and old bee bread, pollen, larvae and adults of solitary bee Osmia cornuta. The 16S rDNA sequencing of 79 selected isolates gave the final species-specific identification of strains. Phylogenetic analysis indicated that microbiota isolated from five different sources were represented with 29 species within three different phyla, Firmicutes with 25 species, Actinobacteria with only one species and Proteobacteria with three species of Enterobacteriaceae. Bacterial biodiversity presented with Shannon-Wiener index (H') was highest in the alimentary tract of adults and old bee bread (H' = 2.43 and H' = 2.53, respectively) and in the same time no dominance of any species was scored. On the contrary, results obtained for Simpson index (D) showed that in pollen samples the dominant species was Pantoea agglomerans (D = 0.42) while in fresh bee bread that was Staphylococcus sp. (D = 0.27). We assume that microbial diversity detected in the tested samples of solitary bee O. cornuta probably come from environment.

Novel antilisterial bacteriocin licheniocin 50.2 from Bacillus licheniformis VPS50.2 isolated from soil sample.

AIM: To isolate and characterize bacteriocin, licheniocin 50.2, from soil bacteria identified as Bacillus licheniformis. METHODS AND RESULTS: The strain B. licheniformis VPS50.2 was identified as bacteriocin producer, effective against Gram-positive bacteria, including Listeria monocytogenes, methicillin-resistant Staphylococcus aureus (MRSA) and ß-haemolytic streptococci. The start of bacteriocin production coincides with the beginning of sporulation. Ammonium sulfate precipitation, chloroform extraction and ultrafiltration were used for bacteriocin purification. MALDI TOF/TOF mass spectrometry of purified sample detected the protein with molecular mass of 3253·209 Da. N-terminal sequencing recognized first 15 amino acids with the sequence: W E E Y N I I X Q L G N K G Q. We named the newly characterized bacteriocin as subclass II.3 bacteriocin, licheniocin 50·2. The bacteriocin activity was insensitive to lysozyme and proteinase K, heat stable after incubation at 100°C for 30 min and over wide range of pH (2-12). MICs of crude bacteriocin extract were determined for L. monocytogenes and MRSA. Time-kill study showed that licheniocin had bactericidal effect to L. monocytogenes. CONCLUSION: A novel, thermostable, pH-tolerant bacteriocin active against Gram-positive bacteria was isolated. SIGNIFICANCE AND IMPACT OF THE STUDY: Attributes of new, stable licheniocin 50.2 make it a promising agent for application as biopreservative in food industry.