Identification of enzyme-producing thermophilic bacilli isolated from marine vents of Aeolian Islands (Italy)

Enzyme-producing thermophilic bacilli were isolated from different thermal sites located in and around Aeolian Islands (Italy) and characterised by both molecular and culture-based methods. Spore-forming bacteria with optimal growth from 45 to 70 °C were isolated from submarine vents and a geothermal well of Aeolian Islands (Italy). Eighteen selected strains were screened for extracellular enzyme presence by using nine substrates: Tween 20, Tween 80, tributyrin, soluble starch, xylan, dextran, carragenan, gelatine and casein. Sixteen isolates were able to grow at pH 9. The isolates were differentiated on the basis of restriction pattern of their amplified 16S rDNA (ARDRA) prior to 16S rRNA gene sequence analysis. On the basis of the most complete sequencing results strain V3 was identified asGeobacillus thermodenitrificans, most of isolates (10/14) was similar at high level (≥95%) to different reference strains of the speciesBacillus licheniformis. The remaining isolates, exhibiting sequence similarity below 95%, may represent novel species of the genusBacillus

The submarine hydrothermal system of Panarea (Southern Italy): biogeochemical processes at the thermal fluids-sea bottom interface

Among the submarine hydrothermal systems located off-shore the volcanic archipelago of the Eolian Islands (Southern Italy), the most active is located off the coasts of Panarea Island. Thermal waters, gases and sulphur deposits coexist at the sea bottom where hydrothermal fluids are released from both shallow and deep vents. The chemical and isotopic composition of the fluid phase shows the presence of a significant magmatic component and the physico-chemical conditions of the geothermal reservoir allow the release of reduced chemical species that are microbially mediated towards the production of organic carbon as a form of biochemical energy. Mi- croorganisms inhabiting this environment possess nutritional requirements and overall metabolic pathways ide- ally suited to such ecosystem that represents a clear example of the close connection between geosphere and biosphere. Microscopic examination of the white mat attached to rock surfaces showed the presence of Thio- thrix-like filamentous bacteria. Moderately thermophilic heterotrophic isolates were identified as strains of the genus Bacillus. Although the hydrothermal system of Panarea has to be considered a «shallow» system, it shows many characteristics that make it similar to the «deep» oceanic systems, giving a unique opportunity for improv- ing our knowledge on such an unexplored world by working at this easily accessible site

Phylogenetic Diversity of Archaea in Shallow Hydrothermal Vents of Eolian Islands, Italy

Shallow hydrothermal systems (SHS) around the Eolian Islands (Italy), related to both active and extinct volcanism, are characterized by high temperatures, high concentrations of CO2 and H2S, and low pH, prohibitive for the majority of eukaryotes which are less tolerant to the extreme conditions than prokaryotes. Archaea and bacteria are the key elements for the functioning of these ecosystems, as they are involved in the transformation of inorganic compounds released from the vent emissions and are at the basis of the hydrothermal system food web. New extremophilic archaea (thermophilic, hyperthermophilic, acidophilic, alkalophilic, etc.) have been isolated from vents of Vulcano Island, exhibiting interesting features potentially valuable in biotechnology. Metagenomic analyses, which mainly involved molecular studies of the 16S rRNA gene, provided different insights into microbial composition associated with Eolian SHS. Archaeal community composition at Eolian vent sites results greatly affected by the geochemistry of the studied vents, principally by hypersaline conditions and declining temperatures. Archaeal community in sediments was mostly composed by hyperthermophilic members of Crenarchaeota (class Thermoprotei) and Euryarchaeota (Thermococci and Methanococci) at the highest temperature condition. Mesophilic Euryarchaeota (Halobacteria, Methanomicrobia, and Methanobacteria) increased with decreasing temperatures. Eolian SHS harbor a high diversity of largely unknown archaea, and the studied vents may be an important source of new isolates potentially useful for biotechnological purposes

Diversity of prokaryotes at a shallow submarine vent of Panarea Island (Italy) by high-throughput sequencing

To determine microbial community composition and possible key microbial processes in the shallow-sea hydrothermal vent system off Panarea Island (Italy), we examined bacterial and archaeal communities of sediment and fluid samples from a hot vent by 16S rDNA Illumina sequencing technique. Both high abundant (>1% of total sequences), low abundant (from 0.1 to <1%) and rare (< 0.1%) phylogenetic groups were responsible for the distinct prokaryotic communities characterizing the heated sediment and fluid. The bacterial and archaeal communities from sediment were dominated by sequences affiliated with Rhodovulum genus (Alphaproteobacteria), including phototrophic ferrous-iron-oxidizing purple bacteria, Thiohalospira and Thiomicrospira (Gammaproteobacteria), typically involved in the sulphur cycle, and Methanococcus (Euryarchaeota). Fluid communities were dominated by anoxygenic phototrophic members of Chlorobium, followed by Thiomicrospira (Gammaproteobacteria), Sulfurimonas, Arcobacter and Sulfurospirillum (Epsilonproteobacteria), and Methanosarcina (Euryarchaeota). Obtained sequences were affiliated with prokaryotes taking a key part in the carbon, iron and sulphur cycling at the shallow hydrothermal system off Panarea Island. Despite the huge sequencing efforts, a great number of Bacteria and Archaea still remains unaffiliated at genus level, indicating that Black Point vent represents a hotspot of prokaryotic diversity

A polyphasic taxonomic study of thermophilic bacilli from shallow, marine vents

Summary Eighty-seven thermophilic, aerobic, spore-forming bacteria were isolated from shallow, marine, thermal vents of the Eolian Islands (Italy) and tested for a broad spectrum of phenotypic characteristics. A numerical taxonomy study was performed on these isolates and 8 thermophilic Bacillus and Geobacillus reference strains by 89 selected features. Results from cluster analysis showed the formation of nine clusters. Most of the isolates (83%) fell into several phenetically well distinguished clusters, loosely related to Geobacillus thermodenitrificans. The remaining isolates grouped together with different reference strains. Eighteen isolates, representative of the different clusters, were selected for subsequent genotypic characterisation, including partial 16S rDNA sequence analysis of 18 strains and almost complete 16S rDNA sequences of 9 strains. Subsequent DNA/DNA reassociation studies and determination of the base composition of DNA identified seven isolates as Geobacillus thermodenitrificans, two isolates as G. thermoleovorans and one isolate as Bacillus pallidus. Four isolates represented two novel species of Bacillus. The remaining four represented novel Geobacillus species, one of which has recently been described as Bacillus vulcani DSMZ 13174 T

Role of Bacterial Exopolysaccharides as Agents in Counteracting Immune Disorders Induced by Herpes Virus

Extreme marine environments, such as the submarine shallow vents of the Eolian Islands (Italy), offer an almost unexplored source of microorganisms producing unexploited and promising biomolecules for pharmaceutical applications. Thermophilic and thermotolerant bacilli isolated from Eolian vents are able to produce exopolysaccharides (EPSs) with antiviral and immunomodulatory effects against Herpes simplex virus type 2 (HSV-2). HSV-2 is responsible for the most common and continuously increasing viral infections in humans. Due to the appearance of resistance to the available treatments, new biomolecules exhibiting different mechanisms of action could provide novel agents for treating viral infections. The EPSs hinder the HSV-2 replication in human peripheral blood mononuclear cells (PBMC) but not in WISH (Wistar Institute Susan Hayflic) cells line, indicating that cell-mediated immunity was involved in the antiviral activity. High levels of Th1-type cytokines were detected in PBMC treated with all EPSs, while Th2-type cytokines were not induced. These EPSs are water soluble exopolymers able to stimulate the immune response and thus contribute to the antiviral immune defense, acting as immunomodulators. As stimulants of Th1 cell-mediated immunity, they could lead to the development of novel drugs as alternative in the treatment of herpes virus infections, as well as in immunocompromised host

Poly-gamma-Glutamic Acid from Bacillus Horneckiae Strain APA of Shallow Marine Vent Origin with Antiviral and Immunomodulatory Effects against Herpes Simplex Virus Type-2

The shallow vents of the Eolian Islands (Italy) are sources of almost unexplored microorganisms able to produce biomolecules with promising pharmaceutical applications. This work describes the characterization of an extracellular poly-γ-glutamic acid (γ-PGA) from the new Bacillus horneckiae strain APA, and its antiviral and immunomodulatory effects against Herpes simplex virus type 2 (HSV-2). The γ-PGA-APA was able to hinder HSV-2 replication in human peripheral blood mononuclear cells (PBMC), involving the cell-mediated immunity. The exopolymer induced the expression of high levels of Th1–type cytokines in PBMC, while it had no effects on Th2-type cytokines production. This is the first report on a γ-PGA produced by a marine thermotolerant B. horneckiae strain with antiviral and immunomodulator activities. As stimulator of Th1 cell-mediated immunity, this biopolymer could be potentially used in the treatment of immunocompromised hosts as well as in patients affected by herpes virus infections

Comparing Spore Resistance of Bacillus Strains Isolated from Hydrothermal Vents and Spacecraft Assembly Facilities to Environmental Stressors and Decontamination Treatments

Submarine hydrothermal vents are inhabited by a variety of microorganisms capable of tolerating environmental extremes, making them ideal candidates to further expand our knowledge of the limitations for terrestrial life, including their ability to survive the exposure of spaceflight-relevant conditions. The spore resistance of two Bacillus spp. strains, APA and SBP3, isolated from two shallow vents off Panarea Island (Aeolian Islands, Italy), to artificial and environmental stressors (i.e., UVC radiation, X-rays, heat, space vacuum, hydrogen peroxide [H₂O₂], and low-pressure plasma), was compared with that of two close phylogenetic relatives (Bacillus horneckiae and Bacillus oceanisediminis). Additional comparisons were made with Bacillus sp. isolated from spacecraft assembly facilities (B. horneckiae, Bacillus pumilus SAFR-032, and Bacillus nealsonii) and the biodosimetry strain and space microbiology model organism Bacillus subtilis. Overall, a high degree of spore resistance to stressors was observed for the strains isolated from spacecraft assembly facilities, with an exceptional level of resistance seen by B. pumilus SAFR-032. The environmental isolate SBP3 showed a more robust spore resistance to UVC, X-rays, H₂O₂, dry heat, and space vacuum than the closely related B. horneckiae. Both strains (SBP3 and APA) were more thermotolerant than their relatives, B. horneckiae and B. oceanisediminis, respectively. SBP3 may have a novel use as a bacterial model organism for future interrogations into the potential of forward contamination in extraterrestrial environments (e.g., icy moons of Jupiter or Saturn), spacecraft sterilization and, broadly, microbial responses to spaceflight-relevant environmental stressors