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

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.

Response of Coccomyxa cimbrica sp.nov. to Increasing Doses of Cu(II) as a Function of Time: Comparison between Exposure in a Microfluidic Device or with Standard Protocols.

In this study, we explore how the in vitro conditions chosen to cultivate and observe the long-term (up to 72 h) toxic effect of Cu(II) on the freshwater microalga Coccomyxa cimbrica sp.nov. can affect the dose response in time. We test three different cultivation protocols: (i) under static conditions in sealed glass cells, (ii) in a microfluidic device, where the sample is constantly circulated with a peristaltic pump, and (iii) under continuous agitation in plastic falcons on an orbital shaker. The advantage and novelty of this study resides in the fact that each condition can mimic different environmental conditions that alga cells can find in nature. The effect of increasing dose of Cu(II) as a function of time (24, 48, and 72 h) is monitored following chlorophyll a fluorescence intensity from single cells. Fluorescence lifetime imaging experiments are also explored to gain information on the changes induced by Cu(II) in the photosynthetic cycle of this microalga.

Characterization of bacterial communities isolated from municipal waste compost and screening of their plant-interactive phenotypes.

Four batches of commercial compost obtained from the organic fraction of municipal solid waste were analyzed from chemical and microbiological standpoints. The working hypothesis was that, being this type of compost derived partly from plant waste, it could contain plant-growth promoting bacterial endophytes, prone to be active again upon its usual delivery as fertilizer. Culturable bacteria were isolated at different temperatures, quantified by colony morphology, identified taxonomically by 16S sequencing and screened for plant-growth promoting phenotypes including auxin and siderophore production, phosphate solubilization and peptide mineralization to ammonia. In parallel, the total community was assessed by culture independent DNA metabarcoding. The capability of plants to select, uptake and internally multiply bacteria from these compost samples was analyzed using grapevine in-vitro rooting cuttings from which acquired bacteria were reisolated, quantified and their identities determined as above. Major differences in compost bacterial composition were observed as function of the season, with the winter sample being rather distinct from the summer ones. Bacillales and Actinomycetales dominated the culturable communities while Alteromonadales, Oceanospirillales and Flavobacteriales prevailed in the total community. In spite of the challenging composting cycle conditions, the plant nature of the main input substrates appeared determinant in guaranteeing that 82% of the culturable bacteria were found endowed with one or more of the plant growth-promoting phenotypes tested. Beside its fertilization role, compost proved to be also a potential inoculant carrier for the in-soil delivery of plant beneficial microorganisms. Furthermore, upon an in vitro passage through grapevine plants under axenic conditions, the subsequently recoverable endophyte community yielded also members of the Rhizobiales order which had not been detectable when culturing directly from compost. This observation further suggests that compost-borne plant-interacting taxa could be also rescued from non-culturable states and/or enriched above detectability levels by a contact with their potential host plants.

Population structure, demographic history, and selective processes: contrasting evidences from mitochondrial and nuclear markers in the European spiny lobster Palinurus elephas (Fabricius, 1787).

The European spiny lobster Palinurus elephas (Fabricius, 1787) is an ecologically and economically important species inhabiting a wide geographic range that extends from the North-east Atlantic and Azores to the Eastern Mediterranean. We investigated the population structure and evolutionary history of this species by both mitochondrial and microsatellite markers. Ten population samples covering a large part of the species distribution range (three samples from the Atlantic Ocean and seven from the Mediterranean Sea) were analyzed for a portion of the mitochondrial control region and seven polymorphic microsatellite loci. Both markers rejected the hypothesis of panmixia identifying two differentiated gene pools. The control region clearly distinguished the Atlantic and Mediterranean populations in two genetically separated groups. Microsatellites, also revealed two groups roughly associated to the Atlantic-Mediterranean separation, however, the Azores sample did not conform to this geographic scheme. Discrepancy between mitochondrial and nuclear markers emerged also when reconstructing the history of the species. Neutrality tests of the mitochondrial sequences indicated a departure from mutation-drift equilibrium that, combined to the mismatch analysis, pointed toward a sudden population expansion in both Atlantic and Mediterranean gene pools. Unexpectedly, microsatellites did not identify any signal of population expansion neither in the Atlantic pool nor in the Mediterranean one.

Phylogeography of the Chionodraco genus (Perciformes, Channichthydae) in the Southern Ocean.

Antarctic fish of the suborder Notothenioidei represent one of the most notable examples of adaptive radiation in the marine environment. The evolutionary relationships between and within the eight families of this suborder have been well established by numerous studies, whereas the microevolutionary processes of notothenioid species remain largely unexplored. In the present paper we investigated the evolutionary relationships between three closely related species of the genus Chionodraco (family Channichthyidae), namely Chionodraco hamatus, Chionodraco rastrospinosus, and Chionodraco myersi by analysing portions of the mitochondrial genome (D-loop and 16S rRNA). The taxonomic status of C. hamatus and C. rastrospinosus as separate species has been questioned because of the limited number of key morphological characters that distinguish these two taxa. Our results, based on the analysis of several specimens belonging to both morphological groups revealed a small genetic differentiation among haplotypes, however, a clear separation between the two nominal species emerged since all individuals of each of the two taxa clustered together in distinct monophyletic groups. C. myersi appeared more distantly related in the phylogenetic analysis. For one species, C. hamatus, sampling was carried out at three different geographic locations in the area of the Ross Sea and Weddell Sea. The results showed that the partition of the genetic variation within this species is not compatible with the hypothesis of panmixia as gene flow between populations was significantly reduced.

A new L1446P mutation is responsible for impaired von Willebrand factor synthesis, structure, and function.

We report on a new mutation (4337T-->C) in exon 28 of the von Willebrand factor (VWF) gene, resulting in a substitution of L with P at residue 1446 (L1446P) of pre-pro-VWF. The defect is transmitted as a dominant trait and induces a reduced VWF synthesis, an abnormal VWF multimer pattern and a deficient VWF-platelet glycoprotein Ib interaction. The proband had low plasma and platelet VWF antigen levels, a reduced VWF collagen-binding capacity, and a disproportionately low VWF ristocetin cofactor activity, associated with the absence of ristocetin-induced platelet aggregation. Multimer analysis showed that the smaller multimers were slightly low, whereas the larger ones were significantly reduced or absent, with a clear cutoff between the two patterns. Similar hemostatic findings were observed in the proband's sister and nephew. Desmopressin administration restored VWF levels to near normal, but this was not so for VWF ristocetin cofactor activity or ristocetin-induced platelet aggregation. VWF multimers improved after desmopressin, moreover, with the larger forms restored and the smaller ones still relatively more represented. Recombinant P1446 VWF synthesis was reduced at heterozygous level, and its multimer pattern was similar to that observed in plasma VWF. These findings confirm the role of L1446P mutation in determining the von Willebrand disease (VWD) phenotype observed in our patients. Given the lack of large and intermediate VWF multimers, and the fact that the VWF-platelet interaction defect appears to be partially independent of multimer pattern, the VWD associated with L1446P mutation may belong to the type 2A/2M VWD variant.