Variation of near surface atmosphere microbial communities at an urban and a suburban site in Philadelphia, PA, USA.

Microorganisms are abundant in the near surface atmosphere and make up a significant fraction of organic aerosols with implications on both human health and ecosystem services. Despite their importance, studies investigating biogeographical patterns of the atmospheric microbiome between urban and suburban areas are limited. Urban and suburban locations (including their microbial communities) vary considerably depending on climate, topography, industrial activities, demographics and other socio-economic factors. Hence, we need more location-specific data to make informed decision affecting air quality, human health, and the implication of a changing climate and policy decisions. The objective of this study was to describe how the atmospheric microbiome varies in composition and function between urban and suburban sites. We used high-throughput sequencing to analyze microbial communities collected at different times from PM2.5 samples collected by active sampling method (using a pump and an impactor) and dust settling of TSP collected by passive sampling method (no pump and no impactor) from an urban and suburban site. We found diverse communities unique in composition at both sites with equivalent functional potential. Taxonomic composition varied significantly with Proteobacteria, Firmicutes, Actinobacteria, Bacteroidetes, and Other phyla in greater relative abundance at the urban site. In contrast, Cyanobacteria, Tenericutes, Fusobacteria, and Deinococcus, were enriched at the suburban site. Community diversity also demonstrated a high degree of temporal variation within site. We identified over one-third of the communities as potentially pathogenic taxa (urban: 47.52% ± 14.40%, suburban: 34.53% ± 14.60%) and determined the majority of organisms come from animal-associated host or are environmental non-specific. Potentially pathogenic taxa and source environments were similar between active- and passive- sampling method results. Our research is novel it adds to the underrepresented set of studies on atmospheric microbial structure and function across land types and is the first to compare suburban and urban atmospheric communities.

Acrolein toxicity involves oxidative stress caused by glutathione depletion in the yeast Saccharomyces cerevisiae.

Exposure of yeast cells to allyl alcohol results in intracellular production of acrolein. The toxicity of so formed acrolein involves oxidative stress, as (1) strains deficient in antioxidant defense are hypersensitive to allyl alcohol, (2) exposure to allyl alcohol increases the level of thiobarbituric-acid-reactive substances and decreases glutathione level in the cells, (3) hypoxic and anoxic atmosphere and antioxidants protect against allyl alcohol toxicity, and (4) allyl alcohol causes activation of Yap1p. No increased formation of reactive oxygen species was detected in cells exposed to allyl alcohol, so oxidative stress is due to depletion of cellular thiols and thus alteration in the redox state of yeast cells.

Effect of stress on the life span of the yeast Saccharomyces cerevisiae.

A correlation is known to exist in yeast and other organisms between the cellular resistance to stress and the life span. The aim of this study was to examine whether stress treatment does affect the generative life span of yeast cells. Both heat shock (38 degrees C, 30 min) and osmotic stress (0.3 M NaCl, 1 h) applied cyclically were found to increase the mean and maximum life span of Saccharomyces cerevisiae. Both effects were more pronounced in superoxide dismutase-deficient yeast strains (up to 50% prolongation of mean life span and up to 30% prolongation of maximum life span) than in their wild-type counterparts. These data point to the importance of the antioxidant barrier in the stress-induced prolongation of yeast life span.

Reactive oxygen species as second messengers? Induction of the expression of yeast catalase T gene by heat and hyperosmotic stress does not require oxygen.

It is shown that oxygen is not absolutely needed for stress-induced synthesis of catalase T in the yeast Saccharomyces cerevisiae. Yeast cells develop heat resistance after exposure to elevated temperatures in anoxia. The levels of catalase activity and thermotolerance are comparable to those in aerobically stressed cells. While these results obviously do not exclude a stress signaling role of reactive oxygen species in some systems, as postulated by other authors, they suggest that the question of the obligatory requirement for reactive oxygen species in other stress signaling systems should be rigorously re-investigated.

Oxidative stress during aging of stationary cultures of the yeast Saccharomyces cerevisiae.

Comparison of 5 d old stationary cultures of Saccharomyces cerevisiae and of cultures aged for 3 months revealed increased generation of reactive oxygen species assessed by 2', 7'-dichlorofluorescin oxidation, decreased activity of superoxide dismutase, decreased content of glutathione and increased protein carbonyl content during prolonged incubation of stationary yeast cultures. These results point to the occurrence of oxidative stress during aging of stationary cultures of the yeast. The magnitude of this stress was augmented in antioxidant-deficient strains, devoid of superoxide dismutases and catalases, and of decreased glutathione content.

Heme synthesis in yeast does not require oxygen as an obligatory electron acceptor.

In a previous paper (Krawiec, Z., Bilinski, T., Schüller, C. & Ruis, H., 2000, Acta Biochim. Polon. 47, 201-207) we have shown that catalase T holoenzyme is synthesized in the absence of oxygen after treatment of anaerobic yeast cultures with 0.3 M. NaCl, or during heat shock. This finding suggests that heme moiety of the enzyme can either be formed de novo in the absence of oxygen, or derives from the preexisting heme pool present in cells used as inoculum. The strain bearing hem1 mutation, resulting in inability to form delta-aminolevulinate (ALA), the first committed precursor of heme, was used in order to form heme-depleted cells used as inocula. The cultures were supplemented with ALA at the end of anaerobic growth prior the stress treatment. The appearance of active catalase T in the stressed cells strongly suggests that heme moiety of catalase T is formed in the absence of oxygen. This finding suggests the necessity to reconsider current opinions concerning mechanisms of heme synthesis and the role of heme as an oxygen sensor.

Deficiency in superoxide dismutases shortens life span of yeast cells.

Deficiencies in superoxide dismutases (Cu,ZnSOD or Mn-SOD) strongly shorten the life span of yeast cells. The effects of these deficiencies are additive. In contrast, deficiencies in catalases do not influence life span. Our results confirm that free radical processes may be involved in aging.

Sensitivity of antioxidant-deficient yeast Saccharomyces cerevisiae to peroxynitrite and nitric oxide.

Sensitivity of Saccharomyces cerevisiae strains deficient in superoxide dismutases and catalases and of decreased level of glutathione to peroxynitrite and a nitric oxide donor, S-nitrosoglutathione was compared. Moderate but significant differences observed point to increased sensitivity to both agents of yeast deficient in antioxidant defense, the superoxide dismutase-deficient strain showing the highest sensitivity, The sequence of sensitivity of various strains to peroxynitrite and nitric oxide was the same. The results are compatible with the view that cytotoxic effects of peroxynitrite involve formation of secondary reactive oxygen species.

Luminol luminescence induced by oxidants in antioxidant-deficient yeasts Saccharomyces cerevisiae.

Luminol chemiluminescence induced in the presence of yeast cells and yeast cell homogenates was significantly induced by exogenous oxidants (hydrogen peroxide and menadione). tert-Butyl hydroperoxide did not stimulate chemiluminescence by itself but augmented menadione-induced chemiluminescence. Comparison of yeast strains deficient in catalase, superoxide dismutase or glutathione showed that only glutathione-deficient strains showed elevated chemiluminescence in this system. These results support the idea that more reactive species than hydrogen peroxide and superoxide are critical in the induction of luminol chemiluminescence.

Menadione toxicity in Saccharomyces cerevisiae cells: activation by conjugation with glutathione.

Menadione (2-methyl-1,4-naphthoquinone) has been used extensively as an oxidant stressor at the cellular level. However, the mechanism of cytotoxicity of this compound still remains controversial. This study deals with the role of intracellular glutathione in the resistance of the yeast Saccharomyces cerevisiae to menadione. Incubation with 0.5 mM menadione resulted in a decrease of total glutathione concentration in yeast cells, intracellular formation of menadione S-glutathione conjugate and export of the conjugate from cells. GSH-deficient mutants showed lower stimulation of superoxide and hydrogen peroxide production upon exposure to menadione and were more resistant to menadione than wild-type isogenic strains. These results indicate that in yeast cells the formation of S-glutathione conjugate is a major pathway of menadione metabolism and that this reaction leads to redox activation of menadione but permits its removal from the cells.

Protective role of superoxide dismutase in iron toxicity in yeast.

It has been found that yeast mutants deficient in cytosolic superoxide dismutase CuZnSOD are hypersensitive to ferrous iron. In contrast mutants that are deficient in catalases and cytochrome c peroxidase do not differ from the standard strain in this respect. These findings suggest that iron toxicity may depend on the redox status of the cell. They also shed light on the role of superoxide dismutases in preventing the toxic effects of oxygen.

Increased stress parameter synthesis in the yeast Saccharomyces cerevisiae after treatment with 4-hydroxy-2-nonenal.

The metabolism of glutathione (GSH), a marker of oxidative stress and trehalose, a rather general physiological stress marker, was examined in exponentially growing Saccharomyces cerevisiae cells after treatment with 4-hydroxynonenal (HNE). GSH was entirely depleted within a 2 h incubation with 250 microM HNE. After removal of the aldehyde it was replenished by de novo synthesis leading to an overshooting GSH level, which later decreased to the basal level. In addition, trehalose was elevated 4-fold in HNE-treated yeast cells compared to control cells. We conclude that increased GSH levels upon HNE treatment are a general phenomenon of eukaryotic cells to ensure protection and survival during further harsh conditions. Furthermore, we have discovered a new indication for the stress marker trehalose in S. cerevisiae.

Iron toxicity in yeast.

It has been found that yeast cells are sensitive to iron overload only when grown on glucose as a carbon source. Effective concentration of ferrous iron is much higher than that found in natural environments. Effects of ferrous iron are strictly oxygen dependent, what suggest that the formation of hydroxyl radicals in the Fenton reaction is a cause of the toxicity. Respiratory deficiency and pretreatment of cells with antimycin A prevent toxic effects in the late exponential phase of growth, whereas uncouplers and 2mM magnesium salts completely protect even the most vulnerable exponential cells. Generally, toxic effects correlate with the ability of cells to take up this metal. The results presented suggest that during ferrous iron overload iron is transported through the unspecific divalent cation uptake system which is known in fungi. The data suggest that recently described high and low affinity systems of iron uptake in yeast are the only source of iron in natural environments.

Lack of evidence of oxidative damage in antioxidant-deficient strains of Saccharomyces cerevisiae.

The content of reactive carbonyls and of glutathione-protein mixed disulfides, two indices of oxidative stress, were compared in wild-type Saccharomyces cerevisiae and in strains deficient in superoxide dismutase and catalase, and of decreased glutathione level. Both indices were higher in stationary than in logarithmic cultures and were not increased in antioxidant-deficient strains. Oxidation of dichlorofluorescin, an estimate of peroxide production, measured in the presence of exogenous peroxidase, was higher in antioxidant-deficient strains. These results corroborate our previous results on compensatory antioxidant mechanisms in the mutant yeast strains.

Glutathione depletion in the yeast Saccharomyces cerevisiae.

The effect of chosen compounds on the total glutathione (GSH) level in stationary cultures of S. cerevisiae was compared. 1-Chloro-2,4-dinitrobenzene, 1-fluoro-2,4-dinitrobenzene, maleimide, iodacetamide and allyl alcohol (1 mM), and menadione (0.5 mM) caused an almost complete GSH depletion during several minutes. Bromobenzoic acid and chloramine T (I mM), and daunomycin (60 mu M) induced a slower GSH decrease, down to 30-70% after 60 min. Paraquat (1 mM), CuSO(4) (0.5 mM) and cadmium acetate (1 mM) decreased glutathione level down to ca 70%. Diamide (0.5 mM), phenazine methosulphate, phenylhydrazine, acetylphenylhydrazine and H(2)O(2) (1 mM), and t-butyl hydroperoxide (2 mM) did not affect total GSH during 60-min exposure. There was no clear-cut dependence between the ability of various chemicals to deplete cellular GSH and their increased toxicity to a glutathione-poor mutant.

What determines the antioxidant potential of yeast cells?

Reactivity which organic radicals was compared in yeast (Saccharomyces cerevisiae) strains defective in catalase and superoxide dismutase, and with a decreased level of glutathione. Yeast cell homogenates did not show considerable strain-related differences in the ability to scavenge nitroxide (TEMPO) stable free radicals and alkoxyl free radicals generated by decomposition of the free radical initiator AAPH. The "total antioxidant status" based on scavenging of ABTS free radicals showed a good correlation with the radiation resistance of the yeasts. These results point to the importance of other factors, apart from antioxidative enzymes and glutathione, in the determination of cellular resistance to ionizing radiation and other types of free-radical stress.

Resistance to ionizing radiation and antioxidative defence in yeasts. Are antioxidant-deficient cells permanently stressed?

Radiation sensitivity of the yeast (Saccharomyces cerevisiae) strains defective in catalase and superoxide dismutase, and of decreased level of glutathione was compared. Small differences in the radiation sensitivity pointed to the highest sensitivity of the wild-type strain as compared with the antioxidant-defective mutants. Heat shock increases the radiation resistance to a different extent, the response being highest in the wild-type strain and lowest in the glutathione-deficient strain. Catalase induction by heat shock follows the same pattern, significant activity of this enzyme being detectable in the glutathione-deficient mutant under normal growth conditions. These results suggest that compensatory induction of elements of the antioxidant defence system may occur in yeast mutants deficient in antioxidants under normal growth conditions.

Assessment of growth inhibition by aldehydic lipid peroxidation products and related aldehydes by Saccharomyces cerevisiae.

The effect of pretreatment with aldehydes on the subsequent colony forming efficiency (CFE) of Saccharomyces cerevisiae was investigated. All 21 aldehydes tested inhibited CFE in a dose-dependent manner. The effective doses, however, differed markedly from 300 mM to 0.07 mM depending on the functional groups and chain length of the aldehydes. Amongst the nine representatives of n-alkanals, formaldehyde was the most potent inhibitor, reducing CFE to 50 per cent at a dose of 0.3 mM (IC50). In the series of 2-trans-alkenals, acrolein was most effective with an IC50 of 0.08 mM and amongst the 4-hydroxy 2-trans-alkenals, 4-hydroxynonenal was most effective with IC50 of 0.07 mM. In general, effectiveness decreased in the order: 4-hydroxyalkenals > 2-alkenals >> n-alkenals. It is proposed that S. cerevisiae is a promising target cell to elucidate further the molecular mechanisms by which aldehydes, particularly the lipid peroxidation product 4-hydroxynonenal, inhibits cell proliferation.