Genomic and phenotypic insights into the ecology of Arthrobacter from Antarctic soils.

BACKGROUND: Members of the bacterial genus Arthrobacter are both readily cultured and commonly identified in Antarctic soil communities. Currently, relatively little is known about the physiological traits that allow these bacteria to survive in the harsh Antarctic soil environment. The aim of this study is to investigate if Antarctic strains of Arthrobacter owe their resilience to substantial genomic changes compared to Arthrobacter spp. isolated from temperate soil environments. RESULTS: Quantitative PCR-based analysis revealed that up to 4% of the soil bacterial communities were comprised of Arthrobacter spp. at four locations in the Ross Sea Region. Genome analysis of the seven Antarctic Arthrobacter isolates revealed several features that are commonly observed in psychrophilic/psychrotolerant bacteria. These include genes primarily associated with sigma factors, signal transduction pathways, the carotenoid biosynthesis pathway and genes induced by cold-shock, oxidative and osmotic stresses. However, these genes were also identified in genomes of seven temperate Arthrobacter spp., suggesting that these mechanisms are beneficial for growth and survival in a range of soil environments. Phenotypic characterisation revealed that Antarctic Arthrobacter isolates demonstrate significantly lower metabolic versatility and a narrower salinity tolerance range compared to temperate Arthrobacter species. Comparative analyses also revealed fewer protein-coding sequences and a significant decrease in genes associated with transcription and carbohydrate transport and metabolism in four of the seven Antarctic Arthrobacter isolates. Notwithstanding genome incompleteness, these differences together with the decreased metabolic versatility are indicative of genome content scaling. CONCLUSIONS: The genomes of the seven Antarctic Arthrobacter isolates contained several features that may be beneficial for growth and survival in the Antarctic soil environment, although these features were not unique to the Antarctic isolates. These genome sequences allow further investigations into the expression of physiological traits that enable survival under extreme conditions and, more importantly, into the ability of these bacteria to respond to future perturbations including climate change and human impacts.

Paenibacillus darwinianus sp. nov., isolated from gamma-irradiated Antarctic soil.

A novel bacterium, strain Br(T), was isolated from gamma-irradiated soils of the Britannia drift, Lake Wellman Region, Antarctica. This isolate was rod-shaped, endospore forming, Gram-stain-variable, catalase-positive, oxidase-negative and strictly aerobic. Cells possessed a monotrichous flagellum. Optimal growth was observed at 18 °C, pH 7.0 in PYGV or R2A broth. The major cellular fatty acid was anteiso-C15 : 0 (63.4 %). Primary identified lipids included phosphatidylethanolamine, diphosphatidylglycerol and phosphatidylglycerol. Total phospholipid was 60 % (w/w) of the total lipid extract. MK-7 was the dominant isoprenoid quinone. The genomic DNA G+C content was 55.6 mol%. Based on 16S rRNA gene sequence similarity, strain Br(T) clusters within the genus Paenibacillus with similarity values ranging from 93.9 to 95.1 %. Phylogenetic analyses by maximum-likelihood, maximum-parsimony and neighbour-joining methods revealed that strain Br(T) clusters with Paenibacillus daejeonensis (AF290916), Paenibacillus tarimensis (EF125184) and Paenibacillus pinihumi (GQ423057), albeit with weak bootstrap support. On the basis of phenotypic, chemotaxonomic and phylogenetic characteristics, we propose that strain Br(T) represents a novel species, Paenibacillus darwinianus sp. nov. The type strain is Br(T) ( = DSM 27245(T) = ICMP 19912(T)).

Leaching of from stony soils after effluent application.

Irrigation of dairy shed effluent (DSE) onto land is an integral part of New Zealand's farming practice. The use of inappropriate soils can result in contamination of ground waters with microbes and nutrients. A gap in our knowledge is the ability of stony soils to safely treat DSE. Replicates of four stony soils were collected from the Canterbury region of New Zealand as intact soil lysimeters 460 mm in diameter and up to 750 mm deep. The soils had either stones to the surface or 300 to 600 mm fines over stones. To determine breakthrough characteristics, a pulse of DSE (25 mm depth) spiked with bromide (2000 mg L) was applied to the soil cores followed by continuous artificial rainfall, for one pore volume, at 5 mm h. Leachate aliquots were analyzed for , bromide, and NH-N. The lysimeters were then subjected to hoof pugging using a mechanical hoof, and the leaching characteristics of the soil were determined again. breakthrough curves revealed that the potential for to leach through the soils was high for Selwyn very stony soil and low for other soils analyzed. After pugging, leaching of increased in Mackenzie soil with stones to the surface. For most other soil cores, concentrations in soil leachates were low. In soils where stones are close to the surface, especially where the soil matrix is sandy, we anticipate that shallow groundwater is vulnerable to microbial contamination under some land management practices.

Bacterial composition of soils of the Lake Wellman area, Darwin Mountains, Antarctica.

The aim of this study was to examine the bacterial composition of high latitude soils from the Darwin-Hatherton glacier region of Antarctica. Four soil pits on each of four glacial drift sheets were sampled for chemical and microbial analyses. The four drifts-Hatherton, Britannia, Danum, and Isca-ranged, respectively, from early Holocene (10 ky) to mid-Quaternary (ca 900 ky). Numbers of culturable bacteria were low, with highest levels detected in soils from the younger Hatherton drift. DNA was extracted and 16S rRNA gene clone libraries prepared from samples below the desert pavement for each of the four drift sheets. Between 31 and 262 clones were analysed from each of the Hatherton, Britannia, and Danum drifts. Bacterial sequences were dominated by members of the phyla Deinococcus-Thermus, Actinobacteria, and Bacteroidetes. Culturable bacteria, including some that clustered with soil clones (e.g., members of the genera Arthrobacter, Adhaeribacter, and Pontibacter), belonged to Actinobacteria and Bacteroidetes. The isolated bacteria are ideal model organisms for genomic and phenotypic investigations of those attributes that allow bacteria to survive and/or grow in Antarctic soils because they have close relatives that are not tolerant of these conditions.

Occurrence and distribution of capB in Antarctic microorganisms and study of its structure and regulation in the Antarctic biodegradative Pseudomonas sp. 30/3.

The analysis of the cold-shock domain (CSD)-encoding genes, capB and cspA, by PCR amplification showed presence of capB in all 18 Antarctic Pseudomonas isolates, but the absence of cspA. Nucleotide sequence analysis of capB ORF from a biodegradative Pseudomonas 30/3 and its regulatory sequences including the promoter and 5'-UTR was determined and compared with the other CSD-encoding genes. Expression analysis using translational gene fusion of the putative capB promoter and its flanking sequence from Pseudomonas sp. 30/3 with lacZ' exhibited a significant increase in beta-galactosidase activity at 15 and 6 degrees C. Unlike the expression of E. coli CspA, Pseudomonas sp. 30/3 showed a slow but steady increase of the CapB expression at 6 degrees C. Subcellular localization of CapB at 6 degrees C showed accumulation in and around the nucleoid whereas at 22 or 30 degrees C, it was identified around the nucleoid as well as in the cytosol. Our study attempts to elucidate the detailed structure of capB from Pseudomonas 30/3 and the role of 5'UTR in the transcriptional regulation along with the possible role of CapB in transcription and translation suited for the cold adaptation of this bacterium in Antarctic environment.

Detection, expression and quantitation of the biodegradative genes in Antarctic microorganisms using PCR.

In this study, 28 hydrocarbon-degrading bacterial isolates from oil-contaminated Antarctic soils were screened for the presence of biodegradative genes such as alkane hydroxylase (alks), the ISPalpha subunit of naphthalene dioxygenase (ndoB), catechol 2,3-dioxygenase (C23DO) and toluene/biphenyl dioxygenase (todC1/bphA1) by using polymerase chain reaction (PCR) methods. All naphthalene degrading bacterial isolates exhibited the presence of a 648 bp amplicon that shared 97% identity to a known ndoB sequence from Pseudomonas putida. Twenty-two out of the twenty-eight isolates screened were positive for one, two or all three different regions of the C23DO gene. For alkane hydroxylase, all 6 Rhodococcus isolates were PCR-positive for a 194 bp and a 552 bp segment of the alkB gene, but exhibited variable results with primers located at different segments of this gene. Three Pseudomonas spp. 4/101, 19/1, 30/3 amplified 552 bp segment of alkB. Only two Pseudomonas sp. 7/156 and 4/101 amplified a segment of alkB exhibiting 89-94% nucleotide sequence identity with the existing sequence of alkB in the GenBank sequence database. Transcripts of three genes, alkB2, C23DO and ndoB, that were amplified by DNA-PCR in three different bacterial isolates also exhibited positive amplification by reverse transcriptase PCR (RT-PCR) method confirming that these genes are functional. A competitive PCR (cPCR) method was developed for a quantitative estimation of ndoB from pure cultures of the naphthalene-degrading Pseudomonas sp. 30/2. A minimum of 1 x 10(7) copies of the ndoB gene was detected based on the comparison of the intensities of the competitor and target DNA bands. It is expected that the identification and characterization of the biodegradative genes will provide a better understanding of the catabolic pathways in Antarctic psychrotolerant bacteria, and thereby help support bioremediation strategies for oil-contaminated Antarctic soils.

Regionalizing potential for microbial bypass flow through New Zealand soils.

Microbial breakthrough curves of 12 soils, generated by the application of dairy shed effluent followed by continuous artificial rainfall for one pore volume at 5 mm h(-1) onto large undisturbed soil cores, have been ranked as high, medium, or low potential for microbial bypass flow. The ranking is based on the position of the peak in the breakthrough curve. Knowledge of soil properties that affect microbial transport through soil gained from the microbial breakthrough curves was linked to soil classes, or to their accessory properties, of the New Zealand Soil Classification. Spatial depiction of the ratings has been achieved via the national 1:50,000 scale soil map. Soils with a drainage impediment or those with well developed soil structure have a high potential for microbial bypass flow, whereas soils from tephra and Recent Soils with less developed, porous, soil structure have a low potential for microbial bypass flow. The risk rankings should be considered as maxima because management may change some rankings.

Characterization of Arthrobacter nicotinovorans HIM, an atrazine-degrading bacterium, from agricultural soil New Zealand.

Arthrobacter nicotinovorans HIM was isolated directly from an agricultural sandy dune soil 6 months after a single application of atrazine. It grew in minimal medium with atrazine as sole nitrogen source but was unable to mineralize 14C-ring-labelled atrazine. Atrazine was degraded to cyanuric acid. In addition to atrazine the bacterium degraded simazine, terbuthylazine, propazine, cyanazine and prometryn but was unable to grow on terbumeton. When added to soil, A. nicotinovorans HIM did enhance mineralization of 14C-ring-labelled atrazine and simazine, in combination with naturally occurring cyanuric acid degrading microbes resident in the soil. Using PCR, the atrazine-degradation genes atzABC were identified in A. nicotinovorans HIM. Cloning of the atzABC genes revealed significant homology (>99%) with the atrazine degradation genes of Pseudomonas sp. strain ADP. The atrazine degradation genes were held on a 96 kbp plasmid.

Hydrocarbon contamination changes the bacterial diversity of soil from around Scott Base, Antarctica.

A combination of culture-independent and culturing methods was used to determine the impacts of hydrocarbon contamination on the diversity of bacterial communities in coastal soil from Ross Island, Antarctica. While numbers of culturable aerobic heterotrophic microbes were 1-2 orders of magnitude higher in the hydrocarbon-contaminated soil than control soil, the populations were less diverse. Members of the divisions Fibrobacter/Acidobacterium, Cytophaga/Flavobacterium/Bacteroides, Deinococcus/Thermus, and Low G+C gram positive occurred almost exclusively in control soils whereas the contaminated soils were dominated by Proteobacteria; specifically, members of the genera Pseudomonas, Sphingomonas and Variovorax, some of which degrade hydrocarbons. Members of the Actinobacteria were found in both soils.

Draft Genome Sequence of Sphingomonas sp. Strain Ant20, Isolated from Oil-Contaminated Soil on Ross Island, Antarctica.

Here, we present the draft genome of Sphingomonas sp. strain Ant20, isolated from oil-polluted soil near Scott Base, Ross Island, Antarctica. The genome of this aromatic hydrocarbon-degrading bacterium provides valuable information on the microbially mediated biodegradation of aromatic compounds in cold-climate systems.

Genome-based comparative analyses of Antarctic and temperate species of Paenibacillus.

Antarctic soils represent a unique environment characterised by extremes of temperature, salinity, elevated UV radiation, low nutrient and low water content. Despite the harshness of this environment, members of 15 bacterial phyla have been identified in soils of the Ross Sea Region (RSR). However, the survival mechanisms and ecological roles of these phyla are largely unknown. The aim of this study was to investigate whether strains of Paenibacillus darwinianus owe their resilience to substantial genomic changes. For this, genome-based comparative analyses were performed on three P. darwinianus strains, isolated from gamma-irradiated RSR soils, together with nine temperate, soil-dwelling Paenibacillus spp. The genome of each strain was sequenced to over 1,000-fold coverage, then assembled into contigs totalling approximately 3 Mbp per genome. Based on the occurrence of essential, single-copy genes, genome completeness was estimated at approximately 88%. Genome analysis revealed between 3,043-3,091 protein-coding sequences (CDSs), primarily associated with two-component systems, sigma factors, transporters, sporulation and genes induced by cold-shock, oxidative and osmotic stresses. These comparative analyses provide an insight into the metabolic potential of P. darwinianus, revealing potential adaptive mechanisms for survival in Antarctic soils. However, a large proportion of these mechanisms were also identified in temperate Paenibacillus spp., suggesting that these mechanisms are beneficial for growth and survival in a range of soil environments. These analyses have also revealed that the P. darwinianus genomes contain significantly fewer CDSs and have a lower paralogous content. Notwithstanding the incompleteness of the assemblies, the large differences in genome sizes, determined by the number of genes in paralogous clusters and the CDS content, are indicative of genome content scaling. Finally, these sequences are a resource for further investigations into the expression of physiological attributes that enable survival under extreme conditions and selection processes that affect prokaryotic genome evolution.

Draft Genome Sequence of Pseudomonas sp. Strain Ant30-3, a Psychrotolerant Bacterium with Biodegradative Attribute Isolated from Antarctica.

Pseudomonas sp. strain Ant30-3, isolated from fuel-contaminated Antarctic soil, exhibited distinctive psychrotolerant attributes and the potential for degrading aromatic hydrocarbon compounds at cold temperatures. We report here the 6.14-Mb draft genome of Ant30-3, which will provide insights into the genomic basis for the psychrotolerant and biodegradative properties of this bacterium.

Draft Genome Sequence of the Aromatic Hydrocarbon-Degrading Bacterium Sphingobium sp. Strain Ant17, Isolated from Antarctic Soil.

Here, we present the draft genome sequence of Sphingobium sp. strain Ant17, an aromatic hydrocarbon-degrading bacterium that was isolated from Antarctic oil-contaminated soil. An analysis of this genome can lead to insights into the mechanisms of xenobiotic degradation processes at low temperatures and potentially aid in bioremediation applications.

Draft Genome Sequence of Williamsia sp. Strain D3, Isolated From the Darwin Mountains, Antarctica.

Actinobacteria are the dominant taxa in Antarctic desert soils. Here, we describe the first draft genome of a member of the genus Williamsia (strain D3) isolated from Antarctic soil. The genome of this psychrotolerant bacterium may help to elucidate crucial survival mechanisms for organisms inhabiting cold desert soil systems.

Draft Genome Sequence of Microbacterium sp. Strain CH12i, Isolated from Shallow Groundwater in Cape Hallett, Antarctica.

The Antarctic continent is largely covered by an expansive ice sheet, but it harbors diverse terrestrial and aquatic habitats in the coastal ice-free continental margins. Here we present the draft genome of Microbacterium sp. CH12i, which was isolated from hypersaline, alkaline, and nutrient-rich groundwater from Cape Hallett, northern Victoria Land, Antarctica.

Bioremediation of hydrocarbon-contaminated polar soils.

Bioremediation is increasingly viewed as an appropriate remediation technology for hydrocarbon-contaminated polar soils. As for all soils, the successful application of bioremediation depends on appropriate biodegradative microbes and environmental conditions in situ. Laboratory studies have confirmed that hydrocarbon-degrading bacteria typically assigned to the genera Rhodococcus, Sphingomonas or Pseudomonas are present in contaminated polar soils. However, as indicated by the persistence of spilled hydrocarbons, environmental conditions in situ are suboptimal for biodegradation in polar soils. Therefore, it is likely that ex situ bioremediation will be the method of choice for ameliorating and controlling the factors limiting microbial activity, i.e. low and fluctuating soil temperatures, low levels of nutrients, and possible alkalinity and low moisture. Care must be taken when adding nutrients to the coarse-textured, low-moisture soils prevalent in continental Antarctica and the high Arctic because excess levels can inhibit hydrocarbon biodegradation by decreasing soil water potentials. Bioremediation experiments conducted on site in the Arctic indicate that land farming and biopiles may be useful approaches for bioremediation of polar soils.

Toluene-degrading Antarctic Pseudomonas strains from fuel-contaminated soil.

Two psychrotolerant toluene-degrading Pseudomonas spp. were isolated from JP8 jet-fuel-contaminated soils, Scott Base, Antarctica. Isolates metabolized meta-toluate as sole carbon source at temperatures ranging from 6 to 30 degrees C. Large plasmids (>64kb) were isolated from both isolates. Sequence analysis of PCR products amplified using xylB (the gene encoding benzyl alcohol dehydrogenase) primers revealed that isolates 7/167 and 8/46 were 100% and 92% homologous, respectively, to the xylB gene of the meta-cleavage toluene degradative pathway encoded by the TOL plasmid (pWWO) of Pseudomonas putida mt-2. Assays of cell-free extracts of 7/167 and 8/46 demonstrated activity of catechol 2,3-dioxygenase, benzyl alcohol dehydrogenase, and benzaldehyde dehydrogenase, indicating that the isolates use the meta-cleavage pathway enzymes of toluene degradation typical of TOL type plasmids. As both isolates are able to grow at 6 degrees C ex situ it is feasible that they would be able to metabolize toluene in the Antarctic soils from where they were originally isolated.

Hydrocarbon spills on Antarctic soils: effects and management.

Antarctic exploration and research have led to some significant although localized impacts on the environment. Human impacts occur around current or past scientific research stations, typically located on ice-free areas that are predominantly soils. Fuel spills, the most common occurrence, have the potential to cause the greatest environmental impact in the Antarctic through accumulation of aliphatic and aromatic compounds. Effective management of hydrocarbon spills is dependent on understanding how they impact soil properties such as moisture, hydrophobicity, soil temperature, and microbial activity. Numbers of hydrocarbon-degrading bacteria, typically Rhodococcus, Sphingomonas, and Pseudomonas species for example, may become elevated in contaminated soils, but overall microbial diversity declines. Alternative management practices to the current approach of "dig it up and ship it out" are required but must be based on sound information. This review summarizes current understanding of the extent and effects of hydrocarbon spillage on Antarctic soils; the observed physical, chemical, and biological responses of such soils; and current gaps in knowledge.

Free-living heterotrophic nitrogen-fixing bacteria isolated from fuel-contaminated antarctic soils.

Five bacterial isolates enriched from fuel-contaminated Antarctic soils fixed nitrogen in the dark heterotrophically and nonsymbiotically. Two isolates utilized jet fuel vapors and volatile hydrocarbons for growth but not in N-deficient medium. Bacteria such as these may contribute to in situ biodegradation of hydrocarbons in Antarctic soils.