Phage vB_Tsc2631 infects the extremophilic bacterium Thermus scotoductus MAT2631 and uses the Ts2631 endolysin for the release of its progeny. The Ts2631 endolysin is the first endolysin from ...thermophilic bacteriophage with an experimentally validated catalytic site. In silico analysis and computational modelling of the Ts2631 endolysin structure revealed a conserved Zn2+ binding site (His30, Tyr58, His131 and Cys139) similar to Zn2+ binding site of eukaryotic peptidoglycan recognition proteins (PGRPs). We have shown that the Ts2631 endolysin lytic activity is dependent on divalent metal ions (Zn2+ and Ca2+). The Ts2631 endolysin substitution variants H30N, Y58F, H131N and C139S dramatically lost their antimicrobial activity, providing evidence for the role of the aforementioned residues in the lytic activity of the enzyme. The enzyme has proven to be not only thermoresistant, retaining 64.8% of its initial activity after 2 h at 95°C, but also highly thermodynamically stable (Tm = 99.82°C, ΔHcal = 4.58 × 10(4) cal mol(-1)). Substitutions of histidine residues (H30N and H131N) and a cysteine residue (C139S) resulted in variants aggregating at temperatures ≥75°C, indicating a significant role of these residues in enzyme thermostability. The substrate spectrum of the Ts2631 endolysin included extremophiles of the genus Thermus but also Gram-negative mesophiles, such as Escherichia coli, Salmonella panama, Pseudomonas fluorescens and Serratia marcescens. The broad substrate spectrum and high thermostability of this endolysin makes it a good candidate for use as an antimicrobial agent to combat Gram-negative pathogens.
Hot springs are natural habitats for thermophilic Archaea and Bacteria. In this paper, we present the metagenomic analysis of eight globally distributed terrestrial hot springs from China, Iceland, ...Italy, Russia, and the USA with a temperature range between 61 and 92 ∘C and pH between 1.8 and 7. A comparison of the biodiversity and community composition generally showed a decrease in biodiversity with increasing temperature and decreasing pH. Another important factor shaping microbial diversity of the studied sites was the abundance of organic substrates. Several species of the Crenarchaeal order Thermoprotei were detected, whereas no single bacterial species was found in all samples, suggesting a better adaptation of certain archaeal species to different thermophilic environments. Two hot springs show high abundance of Acidithiobacillus, supporting the idea of a true thermophilic Acidithiobacillus species that can thrive in hyperthermophilic environments. Depending on the sample, up to 58 % of sequencing reads could not be assigned to a known phylum, reinforcing the fact that a large number of microorganisms in nature, including those thriving in hot environments remain to be isolated and characterized.
Hydrothermal systems form at divergent and convergent boundaries of lithospheric plates and within plates due to weakened crust and mantle plumes, playing host to diverse microbial ecosystems. Little ...is known of how differences in tectonic setting influence the geochemical and microbial compositions of these hydrothermal ecosystems. Here, coordinated geochemical and microbial community analyses were conducted on 87 high‐temperature (>65°C) water and sediment samples from hot springs in Yellowstone National Park, Wyoming, USA (n = 41; mantle plume setting), Iceland (n = 41, divergent boundary), and Japan (n = 5; convergent boundary). Region‐specific variation in geochemistry and sediment‐associated 16S rRNA gene amplicon sequence variant (ASV) composition was observed, with 16S rRNA gene assemblages being nearly completely distinguished by region and pH being the most explanatory parameter within regions. Several low abundance ASVs exhibited cosmopolitan distributions across regions, while most high‐abundance ASVs were only identified in specific regions. The presence of some cosmopolitan ASVs across regions argues against dispersal limitation primarily shaping the distribution of taxa among regions. Rather, the results point to local tectonic and geologic characteristics shaping the geochemistry of continental hydrothermal systems that then select for distinct microbial assemblages. These results provide new insights into the co‐evolution of hydrothermal systems and their microbial communities.
Globally distributed hot springs hosted distinct microbial community cohorts that were broadly structured by a similar suite of environmental parameters. Nonetheless, subtle variation in the geochemistry of hot springs in different geologic settings was associated with population‐level differences among regions. These results argue against dispersal limitation as the primary factor shaping hot spring microbial biogeography and suggest that local geologic and hydrologic characteristics dictate geochemical variation among hydrothermal systems that select for region‐specific microbial cohorts.
The microbial diversity of intertidal hot springs on the seashore of northwest Iceland was examined by combining directed in situ enrichments, artificial support colonization, and mat sampling. ...Analysis of 16S rRNA genes revealed the presence of clones related to both marine and terrestrial, thermophilic, mesophilic, and psychrophilic microorganisms scattered among 11 bacterial divisions. No archaea were found. The species composition of the enrichments was affected by the length of the hot periods experienced at low tide and was very different from those found in the biomass. A total of 36 chitinase genes were detected by molecular screening of the samples with degenerate primers for glycoside hydrolase family 18. The chitinase gene diversity was at least twofold higher in the enrichment samples than in the controls, indicating that a much higher diversity of hydrolytic genes can be accessed with this approach.
Several bacteriophages that infect different strains of the thermophilic bacterium
Rhodothermus marinus
were isolated and their infection pattern was studied. One phage, named RM378 was cultivated ...and characterized. The RM378 genome was also sequenced and analyzed. The phage was grouped as a member of the Myoviridae family with A2 morphology. It had a moderately elongated head, with dimensions of 85 and 95 nm between opposite apices and a 150 nm long tail, attached with a connector to the head. RM378 showed a virulent behavior that followed a lytic cycle of infection. It routinely gave lysates with 10
11
pfu/ml, and sometimes reached titers as high as 10
13
pfu/ml. The titer remained stable up to 65 °C but the phage lost viability when incubated at higher temperatures. Heating for 30 min at 96 °C lowered the titer by 10
4
. The RM378 genome consisted of ds DNA of 129.908 bp with a GC ratio of 42.0 % and contained about 120 ORFs. A few structural proteins, such as the major head protein corresponding to the gp23 in T4, could be identified. Only 29 gene products as probable homologs to other proteins of known function could be predicted, with most showing only low similarity to known proteins in other bacteriophages. These and other studies based on sequence analysis of a large number of phage genomes showed RM378 to be distantly related to all other known T4-like phages.
Nine thermophilic strains of aerobic, non-sporulating, heterotrophic bacteria were isolated after enrichment of chimney material sampled from a deep-sea hydrothermal field at a depth of 2634m on the ...East-Pacific Rise (1 °N). The bacteria stained Gram-negative. They were rod-shaped and measured approximately 0.5μm in width and 1.5-3.5μm in length. They grew at 55-80°C, pH 6-8 and 1-6 % NaCl. Optimal growth was observed at 70-75°C, pH7.0 and 1-3 % NaCl. The organisms were identified as members of the genus Rhodothermus, having a 16S rRNA gene similarity of 98.1 % with Rhodothermus marinus DSM 4252(T). The novel isolates differed morphologically, physiologically and chemotaxonomically from R. marinus, e.g. in lack of pigmentation, response to hydrostatic pressure, maximum growth temperature and DNA G+C content. DNA-DNA hybridization revealed a reassociation value of 37.2 % between strain PRI 2902(T) and R. marinus DSM 4252(T), which strongly suggested that they represent different species. Furthermore, AFLP fingerprinting separated the novel strains from R. marinus reference strains. It is therefore concluded that the strains described here should be classified as representatives of a novel species for which the name Rhodothermus profundi sp. nov. is proposed; the type strain is PRI 2902(T) (=DSM 22212(T) =JCM 15944(T)).
Recently, we have shown that glycoside hydrolases enzymes of family GH17 from proteobacteria (genera Pseudomonas, Azotobacter) catalyze elongation transfer reactions with laminari-oligosaccharides ...generating (β1→3) linkages preferably and to a lesser extent (β1→6) or (β1→4) linkages. In the present study, the cloning and characterization of the gene encoding the structurally very similar GH17 domain of the NdvB enzyme from Bradyrhizobium diazoefficiens, designated Glt20, as well as its catalytic properties are described. The Glt20 enzyme was strikingly different from the previously investigated bacterial GH17 enzymes, both regarding substrate specificity and product formation. The Azotobacter and Pseudomonas enzymes cleaved the donor laminari-oligosaccharide substrates three or four moieties from the non-reducing end, generating linear oligosaccharides. In contrast, the Glt20 enzyme cleaved donor laminari-oligosaccharide substrates two glucose moieties from the reducing end, releasing laminaribiose and transferring the remainder to laminari-oligosaccharide acceptor substrates creating only (β1→3)(β1→6) branching points. This enables Glt20 to transfer larger oligosaccharide chains than the other type of bacterial enzymes previously described, and helps explain the biologically significant formation of cyclic β-glucans in B. diazoefficiens.
Strains PRI 2268 and PRI 3838(T) were isolated from two separate hot springs in the Torfajokull geothermal area of South Iceland. The cells were non-motile rods, approximately 0.3 microm in width and ...1.5-2.5 microm in length. Electron microscopy revealed a Gram-negative cell-wall structure. The strains grew at 45-79 degrees C (optimum, 65 degrees C) and pH 5.5-10.5 (optimum, pH 6.0-7.0). 16S rRNA gene sequence analysis indicated that they formed a separate branch within the genus Thermus with 'Thermus kawarayensis' KW11 as their closest cultured relative (96.5 % similarity). The gene sequence similarities of both new isolates to Thermus aquaticus YT-1(T) and Thermus igniterrae RF-4(T) were 96.1 % and 95.5 %, respectively. DNA-DNA relatedness between strain PRI 3838(T) and 'T. kawarayensis' was 46.1 %. The DNA G+C content of strain PRI 3838(T) was 69.0 mol%. The predominant menaquinones, pigmentation, fatty acid profiles and phospholipid profiles of the novel strains were similar to those of other members of the genus Thermus. However, the new strains could be differentiated from the type strains of all other species of the genus Thermus by their lack of catalase activity and their utilization of only a few carbon sources. Furthermore, the novel strains exhibited mixotrophic growth with sulfur oxidation. On the basis of 16S rRNA gene sequence comparisons, DNA-DNA hybridization and physiological and biochemical characteristics, the new isolates represent a novel species. Since the species appears to be ubiquitous in Icelandic hot springs, the name Thermus islandicus sp. nov. is proposed. The type strain is PRI 3838(T) (=DSM 21543(T)=ATCC BAA-1677(T)).
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