Zealand Hot Springs Research Articles

A new symbiotic nanoarchaeote (Candidatus Nanoclepta minutus) and its host (Zestosphaera tikiterensis gen. nov., sp. nov.) from a New Zealand hot spring

Three thermophilic Nanoarchaeota-Crenarchaeota symbiotic systems have been described. We obtained another stable anaerobic enrichment culture at 80°C, pH 6.0 from a New Zealand hot spring. The nanoarchaeote (Ncl-1) and its host (NZ3T) were isolated in co-culture and their genomes assembled. The small (∼200nm) flagellated cocci were often attached to larger cocci. Based on 16S rRNA gene similarity (88.4%) and average amino acid identity (52%), Ncl-1 is closely related to Candidatus Nanopusillus acidilobi. Their genomes both encode for archaeal flagella and partial glycolysis and gluconeogenesis pathways, but lack ATP synthase genes. Like Nanoarchaeum equitans, Ncl-1 has a CRISPR-Cas system. Ncl-1 also relies on its crenarchaeotal host for most of its biosynthetic needs. The host NZ3T was isolated and grows on proteinaceous substrates but not on sugars, alcohols, or fatty acids. NZ3T requires thiosulfate and grows best at 82°C, pH 6.0. NZ3T is most closely related to the Desulfurococcaceae, Ignisphaera aggregans (∼92% 16S rRNA gene sequence similarity, 45% AAI). Based on phylogenetic, physiological and genomic data, Ncl-1 and NZ3T represent novel genera in the Nanoarchaeota and the Desulfurococcaceae, respectively, with the proposed names Candidatus Nanoclepta minutus and Zestosphaera tikiterensis gen. nov., sp. nov., type strain NZ3T (=DSMZ 107634T=OCM 1213T).

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring

ABSTRACTA draft genome of a novel Dictyoglomus sp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to other Dictyoglomus spp. Like its relatives, Dictyoglomus sp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism.

Draft Genome Sequence of a Novel Thermofilum sp. Strain from a New Zealand Hot Spring Enrichment Culture.

ABSTRACTA draft genome of a new Thermofilum sp. strain was obtained from an enrichment culture metagenome. Like its relatives, Thermofilum sp. strain NZ13 is adapted to organic-rich thermal environments and has to depend on other organisms and the environment for some key amino acids, purines, and cofactors.

The major lipid cores of the archaeon Ignisphaera aggregans: implications for the phylogeny and biosynthesis of glycerol monoalkyl glycerol tetraether isoprenoid lipids

The lipid cores from Ignisphaera aggregans, a hyperthermophilic Crenarchaeon recently isolated from New Zealand hot springs, have been profiled by liquid chromatography-tandem mass spectrometry. The distribution revealed includes relatively high proportions of monoalkyl (also known as H-shaped) tetraether cores which have previously been implicated as kingdom-specific biomarkers for the Euryarchaeota. Such high expression of monoalkyl tetraether lipids is unusual in the archaeal domain and may indicate that formation of these components is an adaptive mechanism that allows I. aggregans to regulate membrane behaviour at high temperatures. The observed dialkyl tetraether and monoalkyl tetraether lipid distributions are similar but not fully concordant, showing differences in the average number of incorporated rings. The similarity supports a biosynthetic route to the ring-containing dialkyl and monoalkyl tetraether lipids via a dialkyl tetraether core containing zero rings, or a closely related structural relative, as an intermediate. Currently, however, the precise nature of the biosynthetic route to these lipids cannot be deduced.

Cyanobacterial composition of microbial mats from an Australian thermal spring: a polyphasic evaluation

Cyanobacterial composition of microbial mats from an alkaline thermal spring issuing at 43-71 degrees C from tropical north-eastern Australia are described using a polyphasic approach. Eight genera and 10 species from three cyanobacterial orders were identified based on morphological characters. These represented taxa previously known as thermophilic from other continents. Ultrastructural analysis of the tower mats revealed two filamentous morphotypes contributed the majority of the biomass. Both types had ultrastructural characteristics of the family Pseudanabaenaceae. DNA extracts were made from sections of the tentaculiform towers and the microbial community analysed by 16S cyanobacteria-specific PCR and denaturing-gradient gel electrophoresis. Five significant bands were identified and sequenced. Two bands clustered closely with Oscillatoria amphigranulata isolated from New Zealand hot springs; one unique phylotype had only moderate similarity to a range of Leptolyngbya species; and one phylotype was closely related to a number of Geitlerinema species. Generally the approaches yielded complementary information, however the results suggest that species designation based on morphological and ultrastructural criteria alone often fails to recognize their true phylogenetic position. Conversely some molecular techniques may fail to detect rare taxa suggesting that the widest possible suite of techniques be applied when conducting analyses of cyanobacterial diversity of natural populations. This is the first polyphasic evaluation of thermophilic cyanobacterial communities from the Australian continent.

The geomicrobiology of gold

Microorganisms capable of actively solubilizing and precipitating gold appear to play a larger role in the biogeochemical cycling of gold than previously believed. Recent research suggests that bacteria and archaea are involved in every step of the biogeochemical cycle of gold, from the formation of primary mineralization in hydrothermal and deep subsurface systems to its solubilization, dispersion and re-concentration as secondary gold under surface conditions. Enzymatically catalysed precipitation of gold has been observed in thermophilic and hyperthermophilic bacteria and archaea (for example, Thermotoga maritime, Pyrobaculum islandicum), and their activity led to the formation of gold- and silver-bearing sinters in New Zealand's hot spring systems. Sulphate-reducing bacteria (SRB), for example, Desulfovibrio sp., may be involved in the formation of gold-bearing sulphide minerals in deep subsurface environments; over geological timescales this may contribute to the formation of economic deposits. Iron- and sulphur-oxidizing bacteria (for example, Acidothiobacillus ferrooxidans, A. thiooxidans) are known to breakdown gold-hosting sulphide minerals in zones of primary mineralization, and release associated gold in the process. These and other bacteria (for example, actinobacteria) produce thiosulphate, which is known to oxidize gold and form stable, transportable complexes. Other microbial processes, for example, excretion of amino acids and cyanide, may control gold solubilization in auriferous top- and rhizosphere soils. A number of bacteria and archaea are capable of actively catalysing the precipitation of toxic gold(I/III) complexes. Reductive precipitation of these complexes may improve survival rates of bacterial populations that are capable of (1) detoxifying the immediate cell environment by detecting, excreting and reducing gold complexes, possibly using P-type ATPase efflux pumps as well as membrane vesicles (for example, Salmonella enterica, Cupriavidus (Ralstonia) metallidurans, Plectonema boryanum); (2) gaining metabolic energy by utilizing gold-complexing ligands (for example, thiosulphate by A. ferrooxidans) or (3) using gold as metal centre in enzymes (Micrococcus luteus). C. metallidurans containing biofilms were detected on gold grains from two Australian sites, indicating that gold bioaccumulation may lead to gold biomineralization by forming secondary 'bacterioform' gold. Formation of secondary octahedral gold crystals from gold(III) chloride solution, was promoted by a cyanobacterium (P. boryanum) via an amorphous gold(I) sulphide intermediate. 'Bacterioform' gold and secondary gold crystals are common in quartz pebble conglomerates (QPC), where they are often associated with bituminous organic matter possibly derived from cyanobacteria. This may suggest that cyanobacteria have played a role in the formation of the Witwatersrand QPC, the world's largest gold deposit.

Experimental studies on New Zealand hot spring sinters: rates of growth and textural development

To study the rate of growth of sinters in New Zealand hot springs, field experiments were conducted in seven geothermal areas. At Wairakei, fan-shaped subaqueous deposits of amorphous silica grow rapidly (350 kg a–1 m–2 of drain wall) and are composed of silicified filaments with subordinate bacillus and spirillium-shaped organisms. Characterization of bacteria revealed isolates sharing > 97% 16S rRNA gene sequence homologies affiliated with Thermus, Meiothermus,Bacillus, Tepidomonas, Thermomonas, Porphyrobacter, Thermonema, and, Hydrogenophilus spp., as well as previously uncultured bacteria. At Rotokawa, microstromatolites have a slow growth rate (0.004 mm day–1) that is attributed to low pH, capillary rise, and evaporation. At Champagne Pool, sinter growth (0.023 mm day–1) is dominated by wave action building alternating microbe-rich and microbe-poor layers. Silica sinter was not observed at Waikite, where slides developed a layer of calcite (0.026 mm day–1). Sinter growth at Ngatamariki (0.016 mm day–1) forms by capillary rise, evaporation, and diffusion and at Tokaanu, subaqueous growth is slow (0.002 mm day–1) and contains silicified microbes. Textures at Orakei Korako indicate similar mechanisms to Ngatamariki, except that growth is more rapid (0.023 mm day–1) due to a splash contribution. Silica and calcite saturation indices adequately explain the growth of the sinters and calcite, indicating that microbes are not inducing precipitation where it should not occur. The rate of precipitation is correlated with silica supersaturation, but pH effects can alter this relationship. The degree of preservation of microbial material is explained by the effect of Ostwald ripening on silica spherules. Subaqueous growth allows coarsening of spherules and poor preservation of smaller microbes while subaerial nucleation is rapid, Ostwald ripening is inactive, and better preservation can be expected.

Partial purification of a thermostable dextranase using Sephacryl S-300 adsorption

Thermostable dextranase (1,6-α-d-glucan 6-glucanohydrolysase) from a thermophilic anaerobic bacterium strain Rt364, isolated from a New Zealand hot spring, was partially purified from the cell-free supernatant fluid by adsorption onto Sephacryl S-300, a dextran-based chromatographic resin. It was competitively eluted with 2% T10 dextran, dialysed, concentrated and examined by SDS–PAGE. The overall recovery was 47% and the increase in specific activity by this procedure was 25-fold. The Rt364 dextranase had previously been found to have an optimum temperature of 80 °C and hydrolysed both α-1,6 and α-1,4 glucosidic bonds. Sephacryl S-300 adsorption is a simple, useful step with general application for concentrating and purifying bacterial enzymes that hydrolyse dextrans.

DNA: DNA homology and cellular components of Thermus filiformis and other strains of Thermus from New Zealand hot springs

DNA: DNA homology and cellular components of Thermus filiformis and other strains of Thermus from New Zealand hot springs

DNA: DNA homology and cellular components ofThermus filiformisand other strains ofThermusfrom New Zealand hot springs

Cells of isolates of Thermus from hot springs in New Zealand were tested for the composition of peptidoglycan, the occurrence of respiratory quinones and the mean base composition of DNA. The DNA: DNA homology was tested by the filter hybridisation and spectrophotometric reassociation rate methods. Thermus filiformis and non-filamentous strains isolated from New Zealand hot springs show great homogeneity, and have low DNA: DNA homology with the species Thermus aquaticus, ‘Thermus thermophilus’, and a new genospecies, Thermus brockianus.

A thermophilic green sulfur bacterium from New Zealand hot springs, Chlorobium tepidum sp. nov.

Thermophilic green sulfur bacteria of the genus Chlorobium were isolated from certain acidic high sulfide New Zealand hot springs. Cells were Gram-negative nonmotile rods of variable length and contained bacteriochlorophyll c and chlorosomes. Cultures of thermophilic chlorobia grew only under anaerobic, phototrophic conditions, either photoautotrophically or photoheterotrophically. The optimum growth temperature for the strains of thermophilic green sulfur bacteria isolated was 47–48°C with generation times of about 2 h being observed. The upper temperature limit for growth was about 52°C. Thiosulfate was a major electron donor for photoautotrophic growth while sulfide alone was only poorly used. N2 fixation was observed at 48°C and cell suspensions readily reduced acetylene to ethylene. The G+C content of DNA from strains of thermophilic chlorobia was 56.5–58.2 mol% and the organisms positioned phylogenetically within the green sulfur bacterial branch of the domain Bacteria. The new phototrophs are described as a new species of the genus Chlorobium, Chlorobium tepidum.

Cellulolytic properties of an extremely thermophilic anaerobe

An extremely thermophilic anaerobe was isolated from a New Zealand hot spring by incubating bacterial mat strands in a medium containing xylan. The Gramreaction-negative organism that was subsequently purified had a temperature optimum of 70° C and a pH optimum of 7.0. The isolate, designated strain H173, grew on a restricted range of carbon sources. In batch culture H173 could degrade Avicel completely when supplied at 5 or 10 g l−1. There was an initial growth phase, during which a cellulase complex was produced and carbohydrates fermented to form acetic and lactic acids, followed by a phase where cells were not metabolising but the cellulase complex actively converted cellulose to glucose. When co-cultured with strain Rt8.B1, an ethanologenic extreme thermophile, glucose was fermented to ethanol and acetate, and no reducing sugars accumulated in the medium. In pH controlled batch culture H173 produced an increased amount of lactate and acetate but there was again a phase when reducing sugars accumulated in the medium, and these were converted to ethanol by co-culture with Rt8.B1.

In Situ Morphologies of Some Bacteria from New Zealand Hot Springs

Summary Conventional and unconventional microbial morphotypes were observed in geothermal springs when in situ electron microscopy methods were used. The conventional morphologies observed were rod, coccal and spiral shaped cells. In certain cases, phage and flagellated cells were also observed. The unconventional morphotypes consisted of cells with “flap” like structures or filamentous cells which possessed swollen sac-like structures. Based on the typical cell wall architecture some of the microorganisms could tentatively be assigned to archaebacterial species or Thermus species. The methods described in the paper have aided in the identification and thereby the isolation of some of the unique morphotypes whereas some other types still elude isolation.

Non-diauxic fermentation of acid hydrolysed pine by a strain of Clostridium thermohydrosulfuricum isolated from a New Zealand hot spring

Fermentation of dilute acid hydrolysed pine could be achieved by all of 18 thermophilic glycolytic anaerobic bacteria tested, while only seven isolates representing the species Clostridium thermohydrosulfuricum, Thermoanaerobium brockii and Thermoanaerobacter ethanolicus were able to ferment undiluted hydrolysate. A strain of C. thermohydrosulfuricum, designated Rt8.B1, was notably better at fermenting the hydrolysate and was studied further. Studies showed that glucose and xylose present in the hydrolysate were utilized simultaneously, a phenomenon which also occurred in a medium containing glucose and xylose as the added carbohydrates.

Non-diauxic fermentation of acid hydrolysed pine by a strain ofClostridium thermohydrosulfuricumisolated from a New Zealand hot spring

Fermentation of dilute acid hydrolysed pine could be achieved by all of 18 thermophilic glycolytic anaerobic bacteria tested, while only seven isolates representing the species Clostridium thermohydrosulfuricum, Thermoanaerobium brockii and Thermoanaerobacter ethanolicus were able to ferment undiluted hydrolysate. A strain of C. thermohydrosulfuricum, designated Rt8.B1, was notably better at fermenting the hydrolysate and was studied further. Studies showed that glucose and xylose present in the hydrolysate were utilized simultaneously, a phenomenon which also occurred in a medium containing glucose and xylose as the added carbohydrates.

Thermus filiformis sp. nov., a Filamentous Caldoactive Bacterium

In a preliminary investigation the isolation of a caldoactive filamentous microorganism from a New Zealand hot spring was reported. This organism is described here as a new species belonging to the genus Thermus, namely, Thermus filiformis, based on ultrastructural, phenotypic, and anomalous Gram type characteristics. The cell wall of T. filiformis resembles that of Thermus aquaticus apart from the presence of an extra layer. The Thermus species tested, including T. filiformis, are negative for the aminopeptidase test, which is unusual for a gram-negative genus. T. filiformis is nonproteolytic, unlike most other Thermus strains, and also differs radically from other strains in morphology when it is observed by using phase-contrast microscopy. The single strain of the species has been deposited with the American Type Culture Collection as strain ATCC 43280T (T = type strain).

A thermophilic gliding bacterium from New Zealand hot springs

Abstract 5 isolates of a thermophilic filamentous gliding bacterium have been isolated from 2 New Zealand thermal springs. The isolates were obligately aerobic and grew optimally at 60°C. Growth on media containing single carbon sources has not been achieved, and peptone-containing media have been used to grow the isolates. Carotenoids could be extracted and had an absorbance peak at 450 nm. The organism was Gram-negative and had a G + C mol% of 47. The isolates had a filamentous morphology and exhibited gliding at room temperature. The organism grew rapidly (μmax = 0.64 h−1), first as a pellicle, then as clumps in broth culture. This organism apparently represents a new taxon of thermophilic bacteria.

A thermophilic gliding bacterium from New Zealand hot springs

A thermophilic gliding bacterium from New Zealand hot springs

Isolation of an extremely thermophilic chemoorganotrophic anaerobe similar to Dictyoglomus thermophilum from new zealand hot springs

A strain similar to Dictyoglomus thermophilum, isolated from a New Zealand hot spring, is described. This strictly anaerobic, Gram-negative, non-motile and nonsporulating bacterium usually exists as long thin filaments of 5 to 25 μm by 0.35 to 0.45 μm. Rotund bodies are commonly observed. Thin sections of the cells revealed a two-layered cell wall. The optimum temperature and pH for growth was 70°C and 7.0 and 7.5 respectively. No growth was observed at 40°C and 85°C or at pH 4.5 to pH 9.0. The organism fermented glucose, maltose, mannose, xylose, lactose, cellobiose, galactose and sucrose and produced acetate as the major end-product with significant amounts of lactate, H2 and CO2 and only traces of ethanol. The doubling time on glucose was 10 h. The DNA base composition was 29.5% guanine plus cytosine as determined by the thermal denaturation method. Growth was inhibited by penicillin, tetracycline and chloramphenicol indicating that the organism was a eubacterium. These features are in common with the newly described species Dictyoglomus thermophilum to which the New Zealand isolate belongs.

Isolation of anaerobic, extremely thermophilic, sulphur metabolising archaebacteria from New Zealand hot springs

Enrichments of New Zealand geo‐thermal samples, initiated in anaerobic sulphur‐containing media and incubated at temperatures above 85°C, yielded rod and coccal shaped organisms which possessed archaebacterial characteristics. Pure cultures were isolated and characterised. Five of the seven isolates, which were rod‐shaped organisms and did not have an obligate requirement for sulphur respiration, were similar to Thermoproteus sp. but had more neutral pH optima for growth. Three of these five Thermoproteus sp. were obligate heterotrophs, which has not previously been reported. The two coccal isolates had an obligate requirement for sulphur as an electron acceptor and were similar to Desulfurococcus sp. but again with more neutral pH optima for growth.