Publications by authors named "Andrea Squartini"

71 Publications

Novel Effects of Leonardite-Based Applications on Sugar Beet.

Front Plant Sci 2021 18;12:646025. Epub 2021 Mar 18.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, Padua, Italy.

The present study aimed to explore the effects of foliar application of a leonardite-based product on sugar beet ( L.) plants grown in the field. The approach concerned the evaluation of the community compositional structure of plant endophytic bacteria through a metabarcoding approach, the expression level of a gene panel related to hormonal metabolism and signaling, and the main sugar beet productivity traits. Results indicated that plants treated with leonardite (dosage of 2,000 ml ha, dilution 1:125, 4 mg C l) compared with untreated ones had a significant increase ( < 0.05) in (i) the abundance of spp., recognized to be an endophyte bacterial genus with plant growth-promoting activity; (ii) the expression level of gene, coding for auxin transport proteins; and (iii) sugar yield. This study represents a step forward to advance our understanding of the changes induced by leonardite-based biostimulant in sugar beet.
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http://dx.doi.org/10.3389/fpls.2021.646025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8013720PMC
March 2021

Quantification of rhizomania virus by automated RNA isolation and PCR based methods in sugar beet.

Virusdisease 2021 Mar 19:1-6. Epub 2021 Mar 19.

DAFNAE, Dipartimento di Agronomia Animali Alimenti Risorse Naturali e Ambiente, Università Degli Studi Di Padova, Viale dell'Università 16, 35020 Legnaro, PD Italy.

Rhizomania is a grave disease affecting sugar beet ( L.). It is caused by the Beet Necrotic Yellow Vein Virus (BNYVV), an RNA virus transmitted by the plasmodiophorid vector . Genetic resistance to the virus has been accomplished mostly using phenotype-genotype association studies. As yet, the most convenient method to ascertain plant resistance has been the quantification of viral titer in roots through the ELISA test. This method is particularly time-consuming and clashes with the necessities of modern plant breeding. Here, we propose an alternative and successful phenotyping method based on the automatic extraction of the viral RNA from sugar beet roots and its relative and absolute quantification by quantitative real-time PCR (qRT-PCR) and digital PCR (dPCR), respectively. Such a method enables an improved standardization of the study, as well as an accurate quantification of the virus also in those samples presenting low virus titer, with respect to the ELISA test.

Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-021-00674-7.
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http://dx.doi.org/10.1007/s13337-021-00674-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7976688PMC
March 2021

Pangenomics of the Symbiotic Rhizobiales. Core and Accessory Functions Across a Group Endowed with High Levels of Genomic Plasticity.

Microorganisms 2021 Feb 16;9(2). Epub 2021 Feb 16.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, 35020 Legnaro, Italy.

Pangenome analyses reveal major clues on evolutionary instances and critical genome core conservation. The order Rhizobiales encompasses several families with rather disparate ecological attitudes. Among them, Rhizobiaceae, Bradyrhizobiaceae, Phyllobacteriacreae and Xanthobacteriaceae, include members proficient in mutualistic symbioses with plants based on the bacterial conversion of N into ammonia (nitrogen-fixation). The pangenome of 12 nitrogen-fixing plant symbionts of the Rhizobiales was analyzed yielding total 37,364 loci, with a core genome constituting 700 genes. The percentage of core genes averaged 10.2% over single genomes, and between 5% to 7% were found to be plasmid-associated. The comparison between a representative reference genome and the core genome subset, showed the core genome highly enriched in genes for macromolecule metabolism, ribosomal constituents and overall translation machinery, while membrane/periplasm-associated genes, and transport domains resulted under-represented. The analysis of protein functions revealed that between 1.7% and 4.9% of core proteins could putatively have different functions.
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http://dx.doi.org/10.3390/microorganisms9020407DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7920277PMC
February 2021

A multi-kingdom metabarcoding study on cattle grazing Alpine pastures discloses intra-seasonal shifts in plant selection and faecal microbiota.

Sci Rep 2021 Jan 13;11(1):889. Epub 2021 Jan 13.

Department of Agronomy Food Natural Resources Animals and Environment (DAFNAE), University of Padova, Campus of Agripolis, Viale dell'Università 16, 35020, Legnaro, Padova, Italy.

Diet selection by grazing livestock may affect animal performance as well as the biodiversity of grazed areas. Recent DNA barcoding techniques allow to assess dietary plant composition in faecal samples, which may be additionally integrated by the description of gut microbiota. In this high throughput metabarcoding study, we investigated the diversity of plant, fungal and bacterial taxa in faecal samples of lactating cows of two breeds grazing an Alpine semi-natural grassland during summer. The estimated plant composition of the diet comprised 67 genera and 39 species, which varied remarkably during summer, suggesting a decline of the diet forage value with the advancing of the vegetative season. The fungal community included Neocallimastigomycota gut symbionts, but also Ascomycota and Basidiomycota plant parasite and coprophilous taxa, likely ingested during grazing. The proportion of ingested fungi was remarkably higher than in other studies, and varied during summer, although less than that observed for plants. Some variation related to breed was also detected. The gut bacterial taxa remained stable through the summer but displayed a breed-specific composition. The study provided insights in the reciprocal organisms' interactions affecting, and being affected by, the foraging behaviour: plants showed a high temporal variation, fungi a smaller one, while bacteria had practically none; conversely, the same kingdoms showed the opposite gradient of variation as respect to the animal host breed, as bacteria revealed to be the group mostly characterized by host-specificity.
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http://dx.doi.org/10.1038/s41598-020-79474-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806629PMC
January 2021

Metabarcoding analysis of the bacterial succession during vermicomposting of municipal solid waste employing the earthworm Eisenia fetida.

Sci Total Environ 2021 Apr 25;766:144389. Epub 2020 Dec 25.

Department of Environment & Sustainable Development, Institute of Environment & Sustainable Development, Banaras Hindu University, Varanasi 221005, India. Electronic address:

A culture-independent DNA metabarcoding analysis of the bacterial communities was carried out throughout a complete vermicomposting cycle of municipal solid waste material using the earthworm Eisenia fetida. 16S rRNA amplicons from the initial material (0 days), an intermediate (42 days), and a final stage (84 days) were sequenced in an Illumina NGS platform and compared. A steady increase in community diversity was observed corresponding to a 2.5-fold higher taxa richness and correspondingly risen values of the Shannon and Simpson ecological indexes and the evenness parameter. A total of 49,665 operational taxonomic units (OTUs) were counted. From the qualitative standpoint, a clear successional shift was observed with an initial community dominated by putatively plant-associated groups belonging to the Rhizobiales order within the Alphaproteobacteria class, regressively leaving the scores of relative abundance (RA) to the Firmicutes phylum and in particular to the Bacilli. Vermistabilization of municipal solid waste (MSW) increased (p < 0.001) the TKN and total P content in the final vermicompost, while pH, TOC, and C/N ratio declined (p < 0.001) in the process. Likewise, a progressive decrease was noticed in β-glucosidase, acid phosphatase, and urease activity while protease and dehydrogenase showed a slight increase, followed by a steep fall. A strong positive correlation was observed among the canonical functions of physico-chemical attributes and enzyme activities. The canonical correspondence analysis (CCA) revealed that significant families did not change on the temporal scale; however, their abundance was influenced by the abiotic environmental factors. In comparison to prior studies on vermicomposting that used different earthworm species (Eisenia andrei) and different substrates, results reflect a considerable degree of substrate specificity for the earthworm species used. The results offer clues to optimize the vermistabilization of MSW along with its potential use in agriculture, to foster improved levels of the circular economy.
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http://dx.doi.org/10.1016/j.scitotenv.2020.144389DOI Listing
April 2021

Analysis of the Interaction between L. and Strains Nodulating This Legume in Northwest Spain.

Plants (Basel) 2020 Dec 11;9(12). Epub 2020 Dec 11.

Departamento de Microbiologíay Genética, Universidad de Salamanca, 37007 Salamanca, Spain.

L. (pea) is one of the most cultivated grain legumes in European countries due to the high protein content of its seeds. Nevertheless, the rhizobial microsymbionts of this legume have been scarcely studied in these countries. In this work, we analyzed the rhizobial strains nodulating the pea in a region from Northwestern Spain, where this legume is widely cultivated. The isolated strains were genetically diverse, and the phylogenetic analysis of core and symbiotic genes showed that these strains belong to different clusters related to sv. viciae. Representative strains of these clusters were able to produce cellulose and cellulases, which are two key molecules in the legume infection process. They formed biofilms and produced acyl-homoserine lactones (AHLs), which are involved in the quorum sensing regulation process. They also exhibited several plant growth promotion mechanisms, including phosphate solubilization, siderophore, and indole acetic acid production and symbiotic atmospheric nitrogen fixation. All strains showed high symbiotic efficiency on pea plants, indicating that strains of sv. viciae are promising candidates for the biofertilization of this legume worldwide.
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http://dx.doi.org/10.3390/plants9121755DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763339PMC
December 2020

Green walls to treat kitchen greywater in urban areas: Performance from a pilot-scale experiment.

Sci Total Environ 2021 Feb 4;757:144189. Epub 2020 Dec 4.

Department of Agronomy, Food, Natural resources, Animals and Environment, University of Padova, Viale dell'Universita' 16, 35020 Legnaro, Padova, Italy.

An increase in water use in urban areas is forcing scientists and policy makers to find alternative solutions for freshwater management, aimed at attaining integrated water resources management. Here, we tested in a 2-year experiment (June 2017-April 2019) the treatment performance of an innovative wall cascade constructed wetland (WCCW) system. The aim was to combine the multifunctional benefits of green walls (e.g. aesthetic, surface area requirements) with those of constructed wetland systems (e.g. high pollutants removal efficiencies, water recycling) to treat kitchen greywaters. The WCCW was a terraced system of six phytoremediation lines, each of which was composed of three plastic tanks (3 × 0.04 m), filled with lightweight porous media, and vegetated with different ornamental species, namely Mentha aquatica L., Oenanthe javanica (Blume) DC., and Lysimachia nummularia L. Physicochemical (temperature, pH, electrical conductivity, dissolved oxygen, turbidity) and chemical parameters (chemical oxygen demand, biochemical oxygen demand, anionic surfactants, Kjeldahl, ammonium and nitric nitrogen, total orthophosphate) were monitored at a frequency of at least 15 days, depending on the season and WCCW management. Results showed that the WCCW significantly reduced the main water pollutants (e.g. organic compounds, nutrients), suggesting its potential application in urban environments for water recycling in the context of green infrastructures and ecological sanitation. A culture-independent taxonomic assessment of suspended bacterial communities before and after the treatment showed clear treatment-related shifts, being the functional ecology attributes changed according to changes in greywater chemical parameters. Future research should attempt to optimize the WCCW system management by regulating the nutrients balance to avoid macronutrients deficiency, and setting the most suitable water flow dynamics (hydraulic retention time, saturation-desaturation cycles) to improve the greywater treatment.
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http://dx.doi.org/10.1016/j.scitotenv.2020.144189DOI Listing
February 2021

Intermittent aeration of landfill simulation bioreactors: Effects on emissions and microbial community.

Waste Manag 2020 Nov 19;117:146-156. Epub 2020 Aug 19.

Department of Agronomy, Food, Natural Resources, Animals and Environment, DAFNAE, University of Padua, viale dell'Università 16, 35020 Legnaro (PD), Italy.

Landfill simulation experiments were run at lab-scale to compare the effects of intermittent and continuous aeration on the evolution of leachate composition and biogas production. The experiments were carried out using six reactors; two of them under continuous aeration, two under intermitted aeration and two anaerobic as a control. Different aeration regimes produced different effects on reactors. As expected, carbon discharge via biogas was higher in reactors under continuous aeration than under intermittent aeration. The evolution of leachate quality was affected by the aeration regimes; however, at test end very similar concentration were ascertained for relevant leachate parameters in all aerated reactors. A comprehensive description of the aerobic and anaerobic landfill microbiome is provided, using a metagenomic approach focused on the microbial genome reconstruction. A time course investigation evidenced the modification of the microbiome and revealed taxa and specific microbes more strictly connected to the environmental parameters of the reactors. Methanoculleus, Syntrophomonas and Parabacteroides were identified as the genera more strictly connected to biogas production, while numerous species belonging to Thiomonas, Nitrosomonas, Xanthomonadaceae, Myxococcales and Alcaligenaceae were found to be connected with NH oxidation.
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http://dx.doi.org/10.1016/j.wasman.2020.08.010DOI Listing
November 2020

The distribution of functional N-cycle related genes and ammonia and nitrate nitrogen in soil profiles fertilized with mineral and organic N fertilizer.

PLoS One 2020 2;15(6):e0228364. Epub 2020 Jun 2.

Gruppo Ricicla labs., DiSAA, Università degli Studi di Milano, Milan, Italy.

Nitrogen transformation in soil is a complex process and the soil microbial population can regulate the potential for N mineralization, nitrification and denitrification. Here we show that agricultural soils under standard agricultural N-management are consistently characterized by a high presence of gene copies for some of the key biological activities related to the N-cycle. This led to a strong NO3- reduction (75%) passing from the soil surface (15.38 ± 11.36 g N-NO3 kg-1 on average) to the 1 m deep layer (3.92 ± 4.42 g N-NO3 kg-1 on average), and ensured low nitrate presence in the deepest layer. Under these circumstances the other soil properties play a minor role in reducing soil nitrate presence in soil. However, with excessive N fertilization, the abundance of bacterial gene copies is not sufficient to explain N leaching in soil and other factors, i.e. soil texture and rainfall, become more important in controlling these aspects.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0228364PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7266355PMC
August 2020

The hidden layers of microbial community structure: extracting the concealed diversity dimensions from our sequencing data.

FEMS Microbiol Lett 2020 06;367(11)

Department of Agronomy, Animals, Food, Natural Resources and Environment DAFNAE, University of Padova, Viale dell'Università, 16, 35020 Legnaro, PD, Italy.

Microbial metabarcoding is the standard approach to assess communities' diversity. However reports are often limited to simple OTU abundances for each phylum, giving rather one-dimensional views of microbial assemblages, overlooking other accessible aspects. The first is masked by databases incompleteness; OTU picking involves clustering at 97% (near-species) sequence identity, but different OTUs regularly end up under a same taxon name. When expressing diversity as number of obtained taxonomical names, a large portion of the real diversity lying within the data remains underestimated. Using the 16S sequencing results of an environmental transect across a gradient of 17 coastal habitats we first extracted the number of OTUs hidden under the same name. Further, we observed which was the deepest rank yielded by annotation, revealing for which microbial groups are we missing most knowledge. Data were then used to infer an evolutionary aspect: what is, in each phylum the success of the present time individuals (abundances for each OTU) in relation to their prior evolutionary success in differentiation (number of OTUs). This information reveals whether the past speciation/diversification force is matched by the present competitiveness in reproduction/persistence. The final layer explored is functional diversity, i.e. abundances of groups involved in specific environmental processes.
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http://dx.doi.org/10.1093/femsle/fnaa086DOI Listing
June 2020

Weed Seed Decay in No-Till Field and Planted Riparian Buffer Zone.

Plants (Basel) 2020 Mar 1;9(3). Epub 2020 Mar 1.

Department of Agronomy, Food, Natural resources, Animals and Environment, University of Padova, 35020 Legnaro (PD), Italy.

Field management practices can alter the physical and chemical properties of the soil, also causing changes to the seed bank. Alterations can also occur to the soil microbial community, which in turn can increase or diminish the process of weed seed decay. In this research, the issue of seed degradation was studied in an undisturbed and a no-till soil, trying not only to uncover where seeds are more degraded, but also to investigate the microbial activities that could be involved in this process. Six different weed species, commonly found in northern Italy, were used: and . Seed decay was tested in two different sites, a no-till field and the adjacent buffer zone. Soil microbial activity was also measured using the Fertimetro, an approach based on the degradation of cotton and silk threads buried in the soil for one week. Degradation of the buried seeds was higher in the no-till field soil than in the buffer strip for all the studied species as was the microbial cellulolytic activity. Even though the buffer strip soil is an undisturbed habitat and resulted as having higher organic matter, the no-till soil conditions appeared more unfavourable to seed viability. Our findings suggest that no-till management can improve weed seed suppression in the soil. Moreover, cellulolytic microorganisms play an important role in seedbank longevity, so cellulolytic activity surveys could be used as an early monitoring bioindicator for weed seed suppression in soil.
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http://dx.doi.org/10.3390/plants9030293DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7154824PMC
March 2020

The Role of the Endophytic Microbiome in the Grapevine Response to Environmental Triggers.

Front Plant Sci 2019 9;10:1256. Epub 2019 Oct 9.

Department of Biology, University of Padova, Padova, Italy.

Endophytism within represents a topic of critical relevance due to the multiple standpoints from which it can be approached and considered. From the biological and botanical perspectives, the interaction between microorganisms and perennial woody plants falls within the category of stable relationships from which the plants can benefit in multiple ways. The life cycle of the host ensures persistence in all seasons, repeated chances of contact, and consequent microbiota accumulation over time, leading to potentially high diversity compared with that of herbaceous short-lived plants. Furthermore, grapevines are agriculturally exploited, highly selected germplasms where a profound man-driven footprint has indirectly and unconsciously shaped the inner microbiota through centuries of cultivation and breeding. Moreover, since endophyte metabolism can contribute to that of the plant host and its fruits' biochemical composition, the nature of grapevine endophytic taxa identities, ecological attitudes, potential toxicity, and clinical relevance are aspects worthy of a thorough investigation. Can endophytic taxa efficiently defend grapevines by acting against pests or confer enough fitness to the plants to endure attacks? What are the underlying mechanisms that translate into this or other advantages in the hosting plant? Can endophytes partially redirect plant metabolism, and to what extent do they act by releasing active products? Is the inner microbial colonization necessary priming for a cascade of actions? Are there defined environmental conditions that can trigger the unleashing of key microbial phenotypes? What is the environmental role in providing the ground biodiversity by which the plant can recruit microsymbionts? How much and by what practices and strategies can these symbioses be managed, applied, and directed to achieve the goal of a better sustainable viticulture? By thoroughly reviewing the available literature in the field and critically examining the data and perspectives, the above issues are discussed.
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http://dx.doi.org/10.3389/fpls.2019.01256DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6794716PMC
October 2019

Expression Profiling of Candidate Genes in Sugar Beet Leaves Treated with Leonardite-Based Biostimulant.

High Throughput 2019 Oct 11;8(4). Epub 2019 Oct 11.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, 35020 Legnaro (PD), Italy.

Leonardite-based biostimulants are a large class of compounds, including humic acid substances. Foliar application of biostimulants at field level improves plant growth, yield and quality through metabolic changes and stimulation of plant proton pumps. The present study aimed at identifying optimum dosage of BLACKJAK, a humic acid-based substance, which is able to modify genes involved in sugar beet growth. Thirty-three genes belonging to various biochemical pathway categories were tested in leaves of treated sugar beet ( L.) samples to assess gene expression profiling in response to BLACKJAK. Seedlings of a diploid and multigerm variety were grown in plastic pots and sprayed with two dilutions of BLACKJAK (dilution 1:500-1.0 mg C L and dilution 1:1000-0.5 mg C L). Leaf samples were collected after 24, 48, and 72 h treatment with BLACKJAK for each dilution. RNA was extracted and the quantification of gene expression was performed while using an OpenArray platform. Results of analysis of variance demonstrated that, 15 genes out of a total of 33 genes tested with OpenArray qPCR were significantly affected by treatment and exposure time. Analysis for annotation of gene products and pathways revealed that genes belonging to the mitochondrial respiratory pathways, nitrogen and hormone metabolisms, and nutrient uptake were up-regulated in the BLACKJAK treated samples. Among the up-regulated genes, Bv_PHT2;1 and Bv_GLN1 expression exerted a 2-fold change in 1:1000 and 1:500 BLACKJAK concentrations. Overall, the gene expression data in the BLACKJAK treated leaves demonstrated the induction of plant growth-related genes that were contributed almost to amino acid and nitrogen metabolism, plant defense system, and plant growth.
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http://dx.doi.org/10.3390/ht8040018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6970231PMC
October 2019

Molecular and Morphological Changes Induced by Leonardite-based Biostimulant in L.

Plants (Basel) 2019 Jun 18;8(6). Epub 2019 Jun 18.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, Viale Università, 16, 35020 Legnaro (PD), Italy.

Humic substances extracted from leonardite are widely considered to be bioactive compounds, influencing the whole-plant physiology and the crop yield. The aim of this work was to evaluate the effect of a new formulate based on leonardite in the early stage of growth of sugar beet ( L.). A commercial preparation of leonardite (BLACKJAK) was characterized by ionomic analysis, solid-state C MAS NMR spectroscopy. Seedlings of sugar beet were grown in Hoagland's solution under controlled conditions. After five days of growth, an aliquot of the concentrated BLACKJAK was added to the solution to obtain a final dilution of 1:1000 (0.5 mg C L). The sugar beet response in the early stage of growth was determined by evaluating root morphological traits as well as the changes in the expression of 53 genes related to key morphophysiological processes. Root morphological traits, such as total root length, fine root length (average diameter < 0.5 mm), and number of root tips, were significantly < 0.001) increased in plants treated with BLACKJAK, compared to the untreated plants at all sampling times. At the molecular level, BLACKJAK treatment upregulated many of the evaluated genes. Moreover, both Real Time PCR and digital PCR showed that genes involved in hormonal response, such as PIN, ARF3, LOGL 10, GID1, and BRI1, were significantly 0.05) upregulated by treatment with BLACKJAK. Our study provides essential information to understand the effect of a leonardite-based formulate on plant growth hormone metabolism, although the molecular and physiological basis for these complicated regulatory mechanisms deserve further investigations.
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http://dx.doi.org/10.3390/plants8060181DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6630732PMC
June 2019

Nodule-associated microbiome diversity in wild populations of Sulla coronaria reveals clues on the relative importance of culturable rhizobial symbionts and co-infecting endophytes.

Microbiol Res 2019 Apr 18;221:10-14. Epub 2019 Jan 18.

Department of Agronomy, Food, Natural Resources, Animals and Environment, DAFNAE, University of Padova, Viale dell'Università 16, Legnaro, Padova, Italy. Electronic address:

The culturable bacteria from root nodules of Sulla coronaria growing in spontaneous conditions in Sardinia were characterized. This plant's peculiarity is to represent a legume still found in both wild and cropped statuses. We tested whether culturable bacteria would differ from those commonly isolated from its field-cropped varieties, to date exclusively represented by Rhizobium sullae. 63 isolates from 60 surface-sterilized nodules were analyzed by ARDRA and 16S rDNA sequencing. The official nitrogen-fixing symbiont Rhizobium sullae was found only in 25 nodules out of 60. The remaining nodules did not yield culturable rhizobia but a number of different endophytic genera including Pseudomonas sp. (17 nodules), Microbacterium sp. (15 nodules), Pantoea agglomerans (5 nodules). The situation appears therefore a hybrid between what is commonly observed in other Mediterranean legumes occurring only in wild status (featuring non-culturable rhizobia and arrays of culturable endophytes within nodules), as opposed to cropped legumes (endowed with fully culturable rhizobia and minimal endophytic occurrence). These findings, within a species bridging the ecology between native and cropped conditions, suggest insights on the relative importance of endophytic co-occupancy vs. true N-fixing symbiont culturability within nodules. The latter trait thus appears to accompany the domestication path of plants with a main trade-off of renouncing to interactions with a diversity of endophytic co-invaders; the relationships with those being critical in the non-cropped status. In fact, endophytes are known to promote plant growth in harsh conditions, which can be particularly stressful in the Mediterranean due to drought, highly calcareous soils, and pathogens outbreaks.
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http://dx.doi.org/10.1016/j.micres.2019.01.004DOI Listing
April 2019

Biocontrol traits of Bacillus licheniformis GL174, a culturable endophyte of Vitis vinifera cv. Glera.

BMC Microbiol 2018 10 16;18(1):133. Epub 2018 Oct 16.

Botanical Garden and Department of Biology, University of Padova, Padova, Italy.

Background: Bacillus licheniformis GL174 is a culturable endophytic strain isolated from Vitis vinifera cultivar Glera, the grapevine mainly cultivated for the Prosecco wine production. This strain was previously demonstrated to possess some specific plant growth promoting traits but its endophytic attitude and its role in biocontrol was only partially explored. In this study, the potential biocontrol action of the strain was investigated in vitro and in vivo and, by genome sequence analyses, putative functions involved in biocontrol and plant-bacteria interaction were assessed.

Results: Firstly, to confirm the endophytic behavior of the strain, its ability to colonize grapevine tissues was demonstrated and its biocontrol properties were analyzed. Antagonism test results showed that the strain could reduce and inhibit the mycelium growth of diverse plant pathogens in vitro and in vivo. The strain was demonstrated to produce different molecules of the lipopeptide class; moreover, its genome was sequenced, and analysis of the sequences revealed the presence of many protein-coding genes involved in the biocontrol process, such as transporters, plant-cell lytic enzymes, siderophores and other secondary metabolites.

Conclusions: This step-by-step analysis shows that Bacillus licheniformis GL174 may be a good biocontrol agent candidate, and describes some distinguished traits and possible key elements involved in this process. The use of this strain could potentially help grapevine plants to cope with pathogen attacks and reduce the amount of chemicals used in the vineyard.
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http://dx.doi.org/10.1186/s12866-018-1306-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6192205PMC
October 2018

Dynamic membrane bioreactor (DMBR) for the treatment of landfill leachate; bioreactor's performance and metagenomic insights into microbial community evolution.

Environ Pollut 2018 Dec 31;243(Pt A):326-335. Epub 2018 Aug 31.

Department of Agronomy, Food, Natural Resources, Animals and Environment (DAFNAE), Legnaro, Padova, Italy.

The use of dynamic membranes as a low-cost alternative for conventional membrane for the treatment of landfill leachate (LFL) was investigated in this study. For this purpose a lab-scale, submerged pre-anoxic and post-aerobic bioreactor configuration was used with nylon mesh as dynamic membrane support. The study was conducted at ambient temperature and LFL was fed to the bioreactor in gradually increasing concentration mixed with tap water (from 20% to 100%). The results of this study demonstrated that lower mesh pore size of 52 μm achieved better results in terms of solid-liquid separation performance (turbidity <10 NTU) of the formed dynamic membrane layer as compared to 200 and 85 μm meshes while treating LFL. Consistently high NH-N conversion efficiency of more than 98% was achieved under all nitrogen loading conditions, showing effectiveness of the formed dynamic membrane in retaining slow growing nitrifying species. Total nitrogen removal reached more than 90% however, the denitrification activity showed a fluctuating profile and found to be inhibited by elevated concentrations of free nitrous acid and NO-N at low pH values inside the anoxic bioreactor. A detailed metagenomic analysis allowed a taxonomic investigation over time and revealed the potential biochemical pathways involved in NH-N conversion. This study led to the identification of a dynamic system in which nitrite concentration is determined by the contribution of NH oxidizers (Nitrosomonas), and by a competition between nitrite oxidizers (Nitrospira and Nitrobacter) and reducers (Thauera).
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http://dx.doi.org/10.1016/j.envpol.2018.08.090DOI Listing
December 2018

Application of anaerobic dynamic membrane bioreactor (AnDMBR) for the successful enrichment of Anammox bacteria using mixed anaerobic and aerobic seed sludge.

Bioresour Technol 2018 Oct 4;266:532-540. Epub 2018 Jul 4.

Laboratory of Technologies for Waste, Wastewater and Raw Materials Management, Italian National Agency for New Technologies, Energy and Sustainable Economic Development (ENEA), via M.M. Sole 4, 40129 Bologna, Italy.

This study investigated a novel bioreactor configuration coupled with a side-stream dynamic membrane (DM) for Anammox enrichment as an alternative for conventional membrane. Bioreactor was fed with synthetic feed and seeded with a mix of anaerobic and aerobic sludge. In situ mechanical cleaning was employed for DM cleaning. DM development and performance was analysed over two polyamide-nylon meshes (200 and 52 µm). Solid-liquid separation of 52 µm mesh outperformed 200 µm with an average effluent turbidity of 2.4 ± 0.1 NTU. The system was operated at a maximum nitrogen loading rate of 696 mg-N L d and achieved a maximum nitrogen removal rate of 611.6 mg-N L d. At steady state, the average ammonium, nitrite and total nitrogen removal efficiencies were 87 ± 0.6%, 98.5 ± 0.15% and 87.5 ± 0.56% respectively. Digital realtime PCRSequence analysis showed that Planctomycetales belonging to ascertained Anammox-specific genera progressively increased their presence in the reactor consistently with its nitrogen removal performance.
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http://dx.doi.org/10.1016/j.biortech.2018.06.100DOI Listing
October 2018

Quorum sensing in rhizobia isolated from the spores of the mycorrhizal symbiont Rhizophagus intraradices.

Mycorrhiza 2018 Nov 24;28(8):773-778. Epub 2018 Jun 24.

Department of Agriculture, Food and Environment, University of Pisa, Via del Borghetto 80, 56124, Pisa, Italy.

Most beneficial services provided by arbuscular mycorrhizal fungi (AMF), encompassing improved crop performance and soil resource availability, are mediated by AMF-associated bacteria, showing key-plant growth-promoting (PGP) traits, i.e., the production of indole acetic acid, siderophores and antibiotics, and activities increasing the availability of plant nutrients by nitrogen fixation and phosphate mobilization. Such functions may be affected by the ability of AMF-associated bacteria to communicate through the production and secretion of extracellular small diffusible chemical signals, N-acyl homoserine lactone signal molecules (AHLs), that regulate bacterial behavior at the community level (quorum sensing, QS). This work investigated the occurrence and extent of QS among rhizobia isolated from AMF spores, using two different QS reporter strains, Agrobacterium tumefaciens NTL4 pZRL4 and Chromobacterium violaceum CV026. We also assessed the quorum quenching (QQ) activity among Bacillus isolated from the same AMF spores. Most rhizobia were found to be quorum-signaling positive, including six isolates producing very high levels of AHLs. The results were confirmed by microtiter plate assay, which detected 65% of the tested bacteria as medium/high AHL producers. A 16S rDNA sequence analysis grouped the rhizobia into two clusters, consistent with the QS phenotype. None of the tested bacteria showed QQ activity able to disrupt the QS signaling, suggesting the absence of antagonism among bacteria living in AMF sporosphere. Our results provide the first evidence of the ability of AMF-associated rhizobia to communicate through QS, suggesting further studies on the potential importance of such a behavior in association with key-plant growth-promoting functions.
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http://dx.doi.org/10.1007/s00572-018-0847-7DOI Listing
November 2018

Biodiversity of fungi in hot desert sands.

Microbiologyopen 2019 01 5;8(1):e00595. Epub 2018 Mar 5.

Department of Infection and Immunity, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia.

The fungal community of six sand samples from Saudi Arabia and Jordan deserts was characterized by culture-independent analysis via next generation sequencing of the 18S rRNA genes and by culture-dependent methods followed by sequencing of internal transcribed spacer (ITS) region. By 18S sequencing were identified from 163 to 507 OTUs per sample, with a percentage of fungi ranging from 3.5% to 82.7%. The identified fungal Phyla were Ascomycota, Basal fungi, and Basidiomycota and the most abundant detected classes were Dothideomycetes, Pezizomycetes, and Sordariomycetes. A total of 11 colonies of filamentous fungi were isolated and cultured from six samples, and the ITS sequencing pointed toward five different species of the class Sordariomycetes, belonging to genera Fusarium (F. redolens, F. solani, F. equiseti), Chaetomium (C. madrasense), and Albifimbria (A. terrestris). The results of this study show an unexpectedly large fungal biodiversity in the Middle East desert sand and their possible role and implications on human health.
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http://dx.doi.org/10.1002/mbo3.595DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6341031PMC
January 2019

Innovative Approaches to Evaluate Sugar Beet Responses to Changes in Sulfate Availability.

Front Plant Sci 2018 31;9:14. Epub 2018 Jan 31.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, Padova, Italy.

In this study, a system based on omics profiling was set-up for sugar beet ( L. subsp. ) evaluation after changes in sulfate availability. Seedlings were grown on sulfate-deprived Hoagland solution. Six days after germination, 100 μM MgSO was added to the solution. Root samples were collected 36 h after treatments. WinRHIZO root-scanning approach was used for the automated image analysis of plant root morphology. Inductively Coupled Plasma Spectrometry (ICP-OES) and quadrupole-time-of-flight mass spectrometry (Q-TOF) were used for ionomic and metabolic analysis, respectively. Nanofluidic real-time PCR (OpenArray system) was used for molecular profiling. OpenArray chips were designed with TaqMan probes for 53 sugar beet genes putatively involved in sulfate nutrition. At morphological level treated seedlings showed significantly higher values ( < 0.01) than untreated plants for root traits related to soil exploration and nutrient uptake, such as total root length, fine roots length and root tips number. ICP-OES, Q-TOF and transcriptomic data revealed changes due to sulfate availability in sugar beet samples. Two key results are highlighted in sulfate-supplied roots and leaves. Firstly, high expression levels of auxin efflux carrier component 1 (PIN) and 5-phosphoribosyl-anthranilate, precursor of tryptophan and auxin synthesis, were observed in roots. Secondly, high levels of 2-Cys peroxiredoxin BAS1, chloroplastic, thioredoxin reductase (NADPH) and cysteine synthase, chloroplastic/chromoplastic, acetylserine sulfhydrylase, involved in protection against oxidative stress and cysteine synthase activity, respectively, were observed in leaves. Based on our findings, the combination of evaluated omics approaches could become a key system for the evaluation of the nutritional status of sugar beet under different nutrient availability conditions.
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http://dx.doi.org/10.3389/fpls.2018.00014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5797807PMC
January 2018

Genome Sequence of EM01, Isolated from Midgut and Responsible for Flacherie Disease in Silkworms Reared on an Artificial Diet.

Genome Announc 2018 Jan 18;6(3). Epub 2018 Jan 18.

Department of Agronomy, Food, Natural Resources, Animals and Environment (DAFNAE), Legnaro, PD, Italy.

The whole genome sequence of strain EM01 is reported here. The isolate proved to be the cause of flacherie in To date, the genomes of 11 other strains have been sequenced. EM01 is the only strain that displayed active pathological effects on its associated animal species.
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http://dx.doi.org/10.1128/genomeA.01495-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5773739PMC
January 2018

Rapid peat accumulation favours the occurrence of both fen and bog microbial communities within a Mediterranean, free-floating peat island.

Sci Rep 2017 08 17;7(1):8511. Epub 2017 Aug 17.

Department of Agronomy, Animals, Natural Resources and Environment - DAFNAE, University of Padua, Viale dell'Università 16, 35020, Legnaro (Padova), Italy.

The unique environment of a 4m-thick, free-floating peat island within the Posta Fibreno lake (Central Italy) was analyzed using DNA-based techniques to assess bacterial and fungal community members identity and abundance. Two depths were sampled at 41 and 279 cm from the surface, the former corresponding to an emerged portion of Sphagnum residues accumulated less than 30 yrs ago, and the latter mainly consisting of silty peat belonging to the deeply submerged part of the island, dating back to 1520-1660 AD. The corresponding communities were very diverse, each of them dominated by a different member of the Delta-proteobacteria class for prokaryotes. Among Eukaryotes, Ascomycota prevailed in the shallow layer while Basidiomycota were abundant in the deep sample. The identity of taxa partitioning between acidic surface layer and neutral core is very reminiscent of the differences reported between bogs and fens respectively, supporting the view of Posta Fibreno as a relic transitional floating mire. Moreover, some microbial taxa show an unusual concurrent species convergence between this sub-Mediterranean site and far Nordic or circumpolar environments. This study represents the first report describing the biotic assemblages of such a peculiar environment, and provides some insights into the possible mechanisms of its evolution.
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http://dx.doi.org/10.1038/s41598-017-08662-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5561117PMC
August 2017

Draft Genome Sequence of the Nitrogen-Fixing Type Strain IS123 Focusing on the Key Genes for Symbiosis with its Host L.

Front Microbiol 2017 26;8:1348. Epub 2017 Jul 26.

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of PadovaLegnaro, Italy.

The prominent feature of rhizobia is their molecular dialogue with plant hosts. Such interaction is enabled by the presence of a series of symbiotic genes encoding for the synthesis and export of signals triggering organogenetic and physiological responses in the plant. The genome of the type strain IS123 nodulating the legume , was sequenced and resulted in 317 scaffolds for a total assembled size of 7,889,576 bp. Its features were compared with those of genomes from rhizobia representing an increasing gradient of taxonomical distance, from a conspecific isolate ( WSM1592), to two congeneric cases ( bv. and ) and up to different genera within the legume-nodulating taxa. The host plant is of agricultural importance, but, unlike the majority of other domesticated plant species, it is able to survive quite well in the wild. Data showed that that the type strain of , isolated from a wild host specimen, is endowed with a richer array of symbiotic genes in comparison to other strains, species or genera of rhizobia that were rescued from domesticated plant ecotypes. The analysis revealed that the bacterium by itself is incapable of surviving in the extreme conditions that its host plant can tolerate. When exposed to drought or alkaline condition, the bacterium depends on its host to survive. Data are consistent with the view of the plant phenotype as the primary factor enabling symbiotic nitrogen fixing bacteria to survive in otherwise limiting environments.
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http://dx.doi.org/10.3389/fmicb.2017.01348DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5526965PMC
July 2017

Differential sensitivity to infections and antimicrobial peptide-mediated immune response in four silkworm strains with different geographical origin.

Sci Rep 2017 04 21;7(1):1048. Epub 2017 Apr 21.

Department of Biology, University of Padova, Padova, Italy.

The domesticated silkworm Bombyx mori has an innate immune system, whose main effectors are the antimicrobial peptides (AMPs). Silkworm strains are commonly grouped into four geographical types (Japanese, Chinese, European and Tropical) and are generally characterised by a variable susceptibility to infections. To clarify the genetic and molecular mechanisms on which the different responses to infections are based, we exposed one silkworm strain for each geographical area to oral infections with the silkworm pathogens Enterococcus mundtii or Serratia marcescens. We detected a differential susceptibility to both bacteria, with the European strain displaying the lowest sensitivity to E. mundtii and the Indian one to S. marcescens. We found that all the strains were able to activate the AMP response against E. mundtii. However, the highest tolerance of the European strain appeared to be related to the specific composition of its AMP cocktail, containing more effective variants such as a peculiar Cecropin B6 isoform. The resistance of the Indian strain to S. marcescens seemed to be associated with its prompt capability to activate the systemic transcription of AMPs. These data suggest that B. mori strains with distinct genetic backgrounds employ different strategies to counteract bacterial infections, whose efficacy appears to be pathogen-dependent.
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http://dx.doi.org/10.1038/s41598-017-01162-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5430696PMC
April 2017

Metagenomic analysis reveals changes of the Drosophila suzukii microbiota in the newly colonized regions.

Insect Sci 2018 Oct 9;25(5):833-846. Epub 2017 Jun 9.

Department of Agronomy, Food, Natural resources, Animals and Environment (DAFNAE), University of Padova, Legnaro, (PD), Italy.

The spotted wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae) is a highly polyphagous pest of a wide variety of wild or cultivated berry and stone fruit. Originating from Southeast Asia, it has recently invaded a wide range of regions in Europe and North America. It is well known that insect microbiotas may significantly influence several aspects of the host biology and play an important role in invasive species introduction into new areas. However, in spite of the great economic importance of D. suzukii, a limited attention has been given so far to its microbiota. In this study, we present the first in-depth characterization of gut bacterial diversity from field (native and invasive range) and lab-reared populations of this insect. The gut bacterial communities of field insects were dominated, regardless of their origin, by 2 families of the phylum Proteobacteria: Acetobacteraceae and Enterobacteriaceae, while Firmicutes, mainly represented by the family Staphylococcaceae, prevailed in lab-reared population. Locality was the most significant factor in shaping the microbiota of wild flies. Moreover, a negative correlation between diversity and abundance of Enterobacteriaceae and the time elapsed since the establishment of D. suzukii in a new region was observed. Altogether our results indicate that habitat, food resources as well as the colonization phase of a new region contribute to shape the bacterial communities of the invasive species which, in turn, by evolving more quickly, could influence host adaptation in a new environment.
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http://dx.doi.org/10.1111/1744-7917.12458DOI Listing
October 2018

Modeling quorum sensing trade-offs between bacterial cell density and system extension from open boundaries.

Sci Rep 2016 12 14;6:39142. Epub 2016 Dec 14.

Department of Agronomy, Animals, Food, Natural resources and Environment DAFNAE, Università di Padova, Viale dell'Università 16, 35020 Legnaro (Padova) Italy.

Bacterial communities undergo collective behavioural switches upon producing and sensing diffusible signal molecules; a mechanism referred to as Quorum Sensing (QS). Exemplarily, biofilm organic matrices are built concertedly by bacteria in several environments. QS scope in bacterial ecology has been debated for over 20 years. Different perspectives counterpose the role of density reporter for populations to that of local environment diffusivity probe for individual cells. Here we devise a model system where tubes of different heights contain matrix-embedded producers and sensors. These tubes allow non-limiting signal diffusion from one open end, thereby showing that population spatial extension away from an open boundary can be a main critical factor in QS. Experimental data, successfully recapitulated by a comprehensive mathematical model, demonstrate how tube height can overtake the role of producer density in triggering sensor activation. The biotic degradation of the signal is found to play a major role and to be species-specific and entirely feedback-independent.
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http://dx.doi.org/10.1038/srep39142DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5155435PMC
December 2016

Direct 16S rRNA-seq from bacterial communities: a PCR-independent approach to simultaneously assess microbial diversity and functional activity potential of each taxon.

Sci Rep 2016 08 31;6:32165. Epub 2016 Aug 31.

Department of Agronomy Animals, Food, Natural Resources and Environment, DAFNAE, University of Padova, Legnaro (Padova) Italy.

The analysis of environmental microbial communities has largely relied on a PCR-dependent amplification of genes entailing species identity as 16S rRNA. This approach is susceptible to biases depending on the level of primer matching in different species. Moreover, possible yet-to-discover taxa whose rRNA could differ enough from known ones would not be revealed. DNA-based methods moreover do not provide information on the actual physiological relevance of each taxon within an environment and are affected by the variable number of rRNA operons in different genomes. To overcome these drawbacks we propose an approach of direct sequencing of 16S ribosomal RNA without any primer- or PCR-dependent step. The method was tested on a microbial community developing in an anammox bioreactor sampled at different time-points. A conventional PCR-based amplicon pyrosequencing was run in parallel. The community resulting from direct rRNA sequencing was highly consistent with the known biochemical processes operative in the reactor. As direct rRNA-seq is based not only on taxon abundance but also on physiological activity, no comparison between its results and those from PCR-based approaches can be applied. The novel principle is in this respect proposed not as an alternative but rather as a complementary methodology in microbial community studies.
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http://dx.doi.org/10.1038/srep32165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5006002PMC
August 2016

Characterization of lipopeptides produced by Bacillus licheniformis using liquid chromatography with accurate tandem mass spectrometry.

Rapid Commun Mass Spectrom 2016 Oct;30(20):2237-52

Department of Biology, University of Padova, via Ugo Bassi 58/B, 35131, Padova, Italy.

Rationale: The plant endophyte Bacillus licheniformis, isolated from leaves of Vitis vinifera, was studied to individuate and characterize the presence of bioactive lipopeptides having amino acidic structures.

Methods: Crude extracts of liquid cultures were analyzed by ultra-high-performance liquid chromatography (UHPLC) coupled to a quadrupole time-of-flight (QTOF) mass analyzer. Chromatographic conditions were optimized in order to obtain an efficient separation of the different isobaric lipopeptides, avoiding merged fragmentations of co-eluted isomeric compounds and reducing possible cross-talk phenomena. Composition of the amino acids was outlined through the interpretation of the fragmentation behavior in tandem high-resolution mass spectrometry (HRMS/MS) mode, which showed both common-class and peculiar fragment ions. Both [M + H](+) and [M + Na](+) precursor ions were fragmented in order to differentiate some isobaric amino acids, i.e. Leu/Ile. Neutral losses characteristic of the iso acyl chain were also evidenced.

Results: More than 90 compounds belonging to the classes of surfactins and lichenysins, known as biosurfactant molecules, were detected. Sequential LC/HRMS/MS analysis was used to identify linear and cyclic lipopeptides, and to single out the presence of a large number of isomers not previously reported. Some critical issues related to the simultaneous selection of different compounds by the quadrupole filter were highlighted and partially solved, leading to tentative assignments of several structures. Linear lichenysins are described here for the first time.

Conclusions: The approach was proved to be useful for the characterization of non-target lipopeptides, and proposes a rationale MS experimental scheme aimed to investigate the difference in amino acid sequence and/or in the acyl chain of the various congeners, when standards are not available. Results expanded the knowledge about production of linear and cyclic bioactive compounds from Bacillus licheniformis, clarifying the structures of isomeric forms, and enabling the use of selected endophytes to produce fungicides for eco-friendly biocontrol. Copyright © 2016 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/rcm.7705DOI Listing
October 2016