The oxygen isotopic signature of soil‐ and plant‐derived sulphate is controlled by fertilizer type and water source

The oxygen isotope signature of sulphate (δ¹⁸O_sulphate) is increasingly used to study nutritional fluxes and sulphur transformation processes in a variety of natural environments. However, mechanisms controlling the δ¹⁸O_sulphate signature in soil–plant systems are largely unknown. The objective of this study was to determine key factors, which affect δ¹⁸O_sulphate values in soil and plants. The impact of an ¹⁸O‐water isotopic gradient and different types of fertilizers was investigated in a soil incubation study and a radish (Raphanus sativus L.) greenhouse growth experiment. Water provided 31–64% of oxygen atoms in soil sulphate formed via mineralization of organic residues (green and chicken manures) while 49% of oxygen atoms were derived from water during oxidation of elemental sulphur. In contrast, δ¹⁸O_sulphate values of synthetic fertilizer were not affected by soil water. Correlations between soil and plant δ¹⁸O_sulphate values were controlled by water δ¹⁸O values and fertilizer treatments. Additionally, plant δ³⁴S data showed that the sulphate isotopic composition of plants is a function of S assimilation. This study documents the potential of using compound‐specific isotope ratio analysis for investigating and tracing fertilization strategies in agricultural and environmental studies.     

Isolation of alkane‐degrading bacteria from deep‐sea Mediterranean sediments

Aims: To isolate and identify alkane‐degrading bacteria from deep‐sea superficial sediments sampled at a north‐western Mediterranean station. Methods and Results: Sediments from the water/sediment interface at a 2400 m depth were sampled with a multicorer at the ANTARES site off the French Mediterranean coast and were promptly enriched with Maya crude oil as the sole source of carbon and energy. Alkane‐degrading bacteria belonging to the genera Alcanivorax, Pseudomonas, Marinobacter, Rhodococcus and Clavibacter‐like were isolated, indicating that the same groups were potentially involved in hydrocarbon biodegradation in deep sea as in coastal waters. Conclusions: These results confirm that members of Alcanivorax are important obligate alkane degraders in deep‐sea environments and coexist with other degrading bacteria inhabiting the deep‐subsurface sediment of the Mediterranean. Significance and Impact of the Study: The results suggest that the isolates obtained have potential applications in bioremediation strategies in deep‐sea environments and highlight the need to identify specific piezophilic hydrocarbon‐degrading bacteria (HCB) from these environments.     

Life‐cycle kinetic model for endospore‐forming bacteria,including germination and sporulation

We develop a mechanistic life‐cycle model for endospore‐forming bacteria (EFB) and test the model with experiments with a Bacillus mixed culture. The model integrates and quantifies how sporulation and germination are triggered by depletion or presence of a limiting substrate, while both substrates affect the rate of vegetative growth by a multiplicative model. Kinetic experiments show the accumulation of small spherical spores after the triggering substrate is depleted, substantially more rapid decay during sporulation than for normal decay of vegetative cells, and a higher specific substrate utilization rate for the germinating cells than that for growth of vegetative cells. Model simulations capture all of these experimental trends. According to model predictions, when a batch reactor is started, seeding with EFB spores instead of active EFB delays the onset of rapid chemical oxygen demand (COD) utilization and biomass growth, but the end points are the same. Simulated results with low aeration intensity show that germination can consume some substrate without dissolved oxygen (DO) depletion. Biotechnol. Bioeng. 2009; 104: 1012–1024. © 2009 Wiley Periodicals, Inc.     

Antagonistic interactions subdue inter‐species green‐beard cooperation in bacteria

Cooperation can be favoured through the green‐beard mechanism, where a set of linked genes encodes both a cooperative trait and a phenotypic marker (green beard), which allows carriers of the trait to selectively direct cooperative acts to other carriers. In theory, the green‐beard mechanism should favour cooperation even when interacting partners are totally unrelated at the genome level. Here, we explore such an extreme green‐beard scenario between two unrelated bacterial species—Pseudomonas aeruginosa and Burkholderia cenocepacia, which share a cooperative locus encoding the public good pyochelin (an iron‐scavenging siderophore) and its cognate receptor (green beard) required for iron–pyochelin uptake. We show that pyochelin, when provided in cell‐free supernatants, can be mutually exchanged between species and provide fitness benefits under iron limitation. However, in co‐culture we observed that these cooperative benefits vanished and communities were dominated by P. aeruginosa, regardless of strain background and species starting frequencies. Our results further suggest that P. aeruginosa engages in interference competition to suppress B. cenocepacia, indicating that inter‐species conflict arising from dissimilarities at the genome level overrule the aligned cooperative interests at the pyochelin locus. Thus, green‐beard cooperation is subdued by competition, indicating that interspecific siderophore cooperation is difficult to evolve and to be maintained.     

Inhibitory effect of iron‐oxidizing bacteria on ferrous‐promoted chalcopyrite leaching

It is generally accepted that iron‐oxidizing bacteria, Thiobacillus ferrooxidans, enhance chalcopyrite leaching. However, this article details a case of the bacteria suppressing chalcopyrite leaching. Bacterial leaching experiments were performed with sulfuric acid solutions containing 0 or 0.04 mol/dm³ ferrous sulfate. Without ferrous sulfate, the bacteria enhance copper extraction and oxidation of ferrous ions released from chalcopyrite. However, the bacteria suppressed chalcopyrite leaching when ferrous sulfate was added. This is mainly due to the bacterial consumption of ferrous ions which act as a promoter for chalcopyrite oxidation with dissolved oxygen. Coprecipitation of copper ions with jarosite formed by the bacterial ferrous oxidation also causes the bacterial suppression of copper extraction. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 64: 478–483, 1999.     

Microbial production of volatile sulphur compounds in the large intestine of pigs fed two different diets

Aims: To investigate the production of volatile sulphur compounds (VSC) in the segments of the large intestine of pigs and to assess the impact of diet on this production. Methods and Results: Pigs were fed two diets based on either wheat and barley (STD) or wheat and dried distillers grains with solubles (DDGS). Net production of VSC and potential sulphate reduction rate (SRR) (sulphate saturated) along the large intestine were determined by means of in vitro incubations. The net production rate of hydrogen sulphide and potential SRR increased from caecum towards distal colon and were significantly higher in the STD group. Conversely, the net methanethiol production rate was significantly higher in the DDGS group, while no difference was observed for dimethyl sulphide. The number of sulphate‐reducing bacteria and total bacteria were determined by quantitative PCR and showed a significant increase along the large intestine, whereas no diet‐related differences were observed. Conclusion: VSC net production varies widely throughout the large intestine of pigs and the microbial processes involved in this production can be affected by diet. Significance and Impact of the Study: This first report on intestinal production of all VSC shows both spatial and dietary effects, which are relevant to both bowel disease‐ and odour mitigation research.     

Degradation of milk‐based bioactive peptides by yogurt fermentation bacteria*

Aims: To analyse the effect of cell‐associated peptidases in yogurt starter culture strains Lactobacillus delbrueckii ssp. bulgaricus (LB) and Streptococcus thermophilus (ST) on milk‐protein‐based antimicrobial and hypotensive peptides in order to determine their survival in yogurt‐type dairy foods. Methods and Results: The 11mer antimicrobial and 12mer hypotensive milk‐protein‐derived peptides were incubated with mid‐log cells of LB and ST, which are required for yogurt production. Incubations were performed at pH 4·5 and 7·0, and samples removed at various time points were analysed by reversed‐phase high‐performance liquid chromatography (RP‐HPLC). The peptides remained mostly intact at pH 4·5 in the presence of ST strains and moderately digested by exposure to LB cells. Peptide loss occurred more rapidly and was more extensive after incubation at pH 7·0. Conclusions: The 11mer and 12mer bioactive peptides may be added at the end of the yogurt‐making process when the pH level has dropped to 4·5, limiting the overall extent of proteolysis. Significance and Impact of the Study: The results show the feasibility of using milk‐protein‐based antimicrobial and hypotensive peptides as food supplements to improve the health‐promoting qualities of liquid and semi‐solid dairy foods prepared by the yogurt fermentation process.     

Gram‐positive bacteria produce membrane vesicles: Proteomics‐based characterization of Staphylococcus aureus‐derived membrane vesicles

Although archaea, Gram‐negative bacteria, and mammalian cells constitutively secrete membrane vesicles (MVs) as a mechanism for cell‐free intercellular communication, this cellular process has been overlooked in Gram‐positive bacteria. Here, we found for the first time that Gram‐positive bacteria naturally produce MVs into the extracellular milieu. Further characterizations showed that the density and size of Staphylococcus aureus‐derived MVs are both similar to those of Gram‐negative bacteria. With a proteomics approach, we identified with high confidence a total of 90 protein components of S. aureus‐derived MVs. In the group of identified proteins, the highly enriched extracellular proteins suggested that a specific sorting mechanism for vesicular proteins exists. We also identified proteins that facilitate the transfer of proteins to other bacteria, as well to eliminate competing organisms, antibiotic resistance, pathological functions in systemic infections, and MV biogenesis. Taken together, these observations suggest that the secretion of MVs is an evolutionally conserved, universal process that occurs from simple organisms to complex multicellular organisms. This information will help us not only to elucidate the biogenesis and functions of MVs, but also to develop therapeutic tools for vaccines, diagnosis, and antibiotics effective against pathogenic strains of Gram‐positive bacteria.     

Biology and epidemics of Candidatus Liberibacter species,psyllid‐transmitted plant‐pathogenic bacteria

Candidatus Liberibacter species are Gram‐negative bacteria that live as phloem‐limited obligate parasites in plants, and are associated with several plant diseases. These bacteria are transmitted by insects called psyllids, or jumping plant lice, which feed on plant phloem sap. Citrus huanglongbing (yellow shoot) or citrus greening disease is associated with three different species of Ca. Liberibacter – Ca. L. asiaticus, Ca. L. africanus and Ca. L. americanus – all originally found on different continents. Ca. L. asiaticus is the most severe pathogen, spread by Asian citrus psyllid Diaphorina citri and causing devastating epidemics in several countries. Ca. L. africanus occurs in Africa where it is spread by the African citrus psyllid Trioza erytreae. Ca. Liberibacter solanacearum is associated with diseases in several solanaceous plants, and transmitted by potato psyllid Bactericera cockerelli. Zebra chip disease is causing large damage in potato crops in North America. In Europe Ca. Liberibacter solanacearum is associated with diseases of the Apiaceae family of plants, carrot and celery, and transmitted by psyllids Trioza apicalis and Bactericera trigonica. When Ca. Liberibacter is suspected as the disease agent, the diagnosis is confirmed by DNA‐based detection methods. Ca. Liberibacter‐associated plant diseases can be controlled by using healthy plant propagation material, eradicating symptomatic plants, and by controlling the psyllid populations spreading the disease.     

Dissolved organic sulphur in soil water under <Emphasis Type="Italic">Pinus sylvestris</Emphasis> L. and <Emphasis Type="Italic">Fagus sylvatica</Emphasis> L. stands in northeastern Bavaria,Germany variations with seasons and soil depth

Organically bound species have been identified as prominent and mobile forms of nitrogen and phosphorus in soils. Since a large portion of sulphur (S) in soil is bonded to carbon (C) also dissolved organic S likely is a significant constituent in soil water. To investigate the role of dissolved organic forms in leaching and cycling of S in forest soils, we examined concentrations, fluxes, and chemical composition of organic S in forest floor leachates and in soil solutions of Rendzic Leptosols under 90-year-old European beech (Fagus sylvatica L.) and Haplic Arenosols under 160-year-old Scots pine (Pinus sylvestris L.) for 27 months. These soils are low in adsorbed SO₄^2- and receive little atmospheric S depositions at present. The chemical composition of organic S was estimated by fractionation with XAD-8 and wet-chemical characterisation (HI reduction) of binding forms. Although not as prominent as the organic forms of other nutrient elements, organic S proved to be an important contributor to S dissolved in forest floor leachates and in mineral soil solutions. Dissolved organic matter contained on average 29% of total S in forest floor leachates at the pine site and 34% at the beech site. The largest portion of organic S occurred in the subsoil solutions under beech in summer and autumn (up to 53%). Mean concentrations of organic S peaked (up to 1.1 mg l^-1) in summer after rainstorms that followed dry periods. Fluxes with forest floor leachates and at 90 cm soil depth were largest in autumn because of huge amounts of rainfall. Organic S contributed significantly to the fluxes of S in the subsoils under beech comprising on average 39% of total dissolved S at 90 cm depth. Organic S produced in the forest floor layers was mainly in the hydrophilic fraction of dissolved organic matter (62 ± 6% at the pine site, 85 ± 4% at the beech site). The major binding form of organic S in the hydrophobic fraction was C-bonded S while in the hydrophilic fraction ester sulphate S, possibly associated with carbohydrates, was more prominent. Since the hydrophobic fraction increased in summer and autumn, C-bonded S was of greater importance during that time of the year than in winter and spring. With depth, concentrations and composition of organic S (and also of C) hardly changed at the pine site because of little retention of dissolved organic matter, presumably because of the small sorption capability of that soil. At the beech where organic C showed a marked decrease with depth, only a slight decrease in organic S, exclusively from the hydrophobic fraction, was found indicating that organic S was mobile compared with organic C. This was probably due to the concentration of S in the hydrophilic fraction of dissolved organic matter. Because of being concentrated in the mobile hydrophilic fraction, ester sulphate S was more mobile in the soil under beech than C-bonded S.     

Isolation of dextran‐hydrolyzing intestinal bacteria and characterization of their dextranolytic activities

The aim of this study was to isolate dextran‐hydrolyzing bacteria from the human intestines and to identify their dextranolytic enzymes. For this, dextranase‐producing microorganisms were screened from fecal samples by using blue dextran‐containing media. Colonies producing a decolorized zone were isolated and they were grouped using RAPD‐PCR. 16S rRNA gene sequencing analysis revealed the isolates were Bacteroides (B.) thetaiotaomicron, B. ovatus, B. vulgatus, B. dorei, B. xylanisolvens, B. uniformis, and Veillonella (V.) rogosae. Thin layer chromatography analysis showed that the dextranases exhibit mainly endo‐type activity and produce various oligosaccharides including isomaltose and isomaltotriose. Zymogram analysis demonstrated that enzymes localized mainly in the cell membrane fraction and the molecular weight was 50–70 kDa. When cultured in a dextran‐containing medium, all strains isolated in this study produced short‐chain fatty acids, with butyric acid as the major compound. This is the first study to report that human intestinal B. xylanisolvens, B. dorei, and V. rogosae metabolize dextran utilizing dextranolytic enzymes. © 2015 Wiley Periodicals, Inc. Biopolymers 103: 321–327, 2015.     

Community structures of ammonia‐oxidising archaea and bacteria in high‐altitude lakes on the Tibetan Plateau

1. Community structures of planktonic ammonia‐oxidising archaea (AOA) and bacteria (AOB) were investigated for five high‐altitude Tibetan lakes, which could be classified as freshwater, oligosaline or mesosaline, to develop a general view of the AOA and AOB in lakes on the Tibetan Plateau. 2. Based on PCR screening of the ammonia monooxygenase α‐subunit (amoA) gene, AOA were present in 14 out of 17 samples, whereas AOB were detected in only four samples. Phylogenetic analyses indicated that the AOB communities were dominated by a unique monophylogenetic lineage within Nitrosomonas, which may represent a novel cluster of AOB. AOA, on the other hand, were distinct among lakes with different salinities. 3. Multivariate statistical analyses indicated a heterogeneous distribution of the AOA communities among lakes largely caused by lake salinity, whereas the uniform chemical properties within lakes and their geographical isolation may favour relatively homogeneous AOA communities within lakes. 4. Our results suggest a wide occurrence of AOA in Tibetan lakes and provide the first evidence of salinity‐related differentiation of AOA community composition as well as potential geographical isolation of AOA in inland aquatic environments.     

STED nanoscopy of KK114‐stained pathogenic bacteria

Super‐resolution microscopy techniques can provide answers to still pending questions on prokaryotic organisms but are yet to be used at their full potential for this purpose. To address this, we evaluate the ability of the rhodamine‐like KK114 dye to label various types of bacteria, to enable imaging of fine structural details with stimulated emission depletion microscopy (STED). We assessed fluorescent labeling with KK114 for eleven Gram‐positive and Gram‐negative bacterial species and observed that this contrast agent binds to their cell membranes. Significant differences in the labeling outputs were noticed across the tested bacterial species, but importantly, KK114‐staining allowed the observation of subtle nanometric cell details in some cases. For example, a helix pattern resembling a cytoskeleton arrangement was detected in Bacillus subtilis. Furthermore, we found that KK114 easily penetrates the membrane of bacterial microorganism that lost their viability, which can be useful to discriminate between living and dead cells.