Slc4-like anion transporters of the larval mosquito alimentary canal

Mosquito larvae exhibit luminal pH extremes along the axial length of their alimentary canal that range from very alkaline (pH>10) in the anterior midgut to slightly acid in the hindgut. The principal buffer in the system is thought to be bicarbonate and/or carbonate, because the lumen is known to contain high levels of bicarbonate/carbonate and is surrounded by various epithelial cell types which express a variety of carbonic anhydrases. However, the precise mechanisms responsible for the transport of bicarbonate/carbonate into and out of the lumen are unclear. In the present study, we test the hypothesis that SLC4-like anion transporters play a role in bicarbonate/carbonate accumulation in the larval mosquito alimentary canal. Molecular, physiological and immnuohistochemical characterizations of Slc4-like transporters in the gut of larval mosquitoes (Aedes aegypti and Anopheles gambiae) demonstrate the presence of both a Na(+)-independent chloride/bicarbonate anion exchanger (AE) as well as a Na(+)-dependent anion exchanger (NDAE). Notably, immunolocalization experiments in Malpighian tubules show that the two proteins can be located in the same tissue, but to different cell types. Immunolabeling experiments in the gastric caecae show that the two proteins can be found in the same cells, but on opposite sides (basal vs. apical). In summary, our results indicate that the alimentary canal of larval mosquitoes exhibits robust expression of two SLC4-like transporters in locations that are consistent with a role in the regulation of luminal pH. The precise physiological contributions of each transporter remain to be determined.     

Bicarbonate enhances the peroxidase activity of Cu,Zn-superoxide dismutase. Role of carbonate anion radical.

We examined the effect of bicarbonate on the peroxidase activity of copper-zinc superoxide dismutase (SOD1), using the nitrite anion as a peroxidase probe. Oxidation of nitrite by the enzyme-bound oxidant results in the formation of the nitrogen dioxide radical, which was measured by monitoring 5-nitro-gamma-tocopherol formation. Results indicate that the presence of bicarbonate is not required for the peroxidase activity of SOD1, as monitored by the SOD1/H(2)O(2)-mediated nitration of gamma-tocopherol in the presence of nitrite. However, bicarbonate enhanced SOD1/H(2)O(2)-dependent oxidation of tocopherols in the presence and absence of nitrite and dramatically enhanced SOD1/H(2)O(2)-mediated oxidation of unsaturated lipid in the presence of nitrite. These results, coupled with the finding that bicarbonate protects against inactivation of SOD1 by H(2)O(2), suggest that SOD1/H(2)O(2) oxidizes the bicarbonate anion to the carbonate radical anion. Thus, the amplification of peroxidase activity of SOD1/H(2)O(2) by bicarbonate is attributed to the intermediary role of the diffusible oxidant, the carbonate radical anion. We conclude that, contrary to a previous report (Sankarapandi, S., and Zweier, J. L. (1999) J. Biol. Chem. 274, 1226-1232), bicarbonate is not required for peroxidase activity mediated by SOD1 and H(2)O(2). However, bicarbonate enhanced the peroxidase activity of SOD1 via formation of a putative carbonate radical anion. Biological implications of the carbonate radical anion in free radical biology are discussed.     

Effect of various salts on the growth and development of Geotrichum candidum,the causal agent of carrot sour rot

This study evaluated the efficacy of ammonium, calcium, potassium and sodium salts as possible alternatives to synthetic fungicides in the control of Geotrichum candidum, the causal agent of sour rot on carrots. In vitro mycelial growth of G. candidum was completely halted by ammonium bicarbonate and carbonate; calcium oxide; potassium benzoate, carbonate and sorbate; sodium benzoate, carbonate and fluoride (2% w/v). Potassium and sodium bicarbonate also reduced mycelial growth by 77.78% and 90.60%, respectively, and the difference between the effects of sodium bicarbonate and the first group of salts was not statistically significant (p < 0.05). With the exception of potassium and sodium bicarbonate, the above‐mentioned salts also halted or strongly reduced arthrospore germination. Potassium bicarbonate, and sodium bicarbonate, acetate and propionate significantly increased conidiation (p < 0.05). Of all the salts tested in vitro, only ammonium bicarbonate and carbonate, calcium oxide and sodium fluoride were toxic to G. candidum. In in vivo studies, all the calcium salts tested (acetate, chloride, citrate, formate, lactate, oxide, propionate and silicate), several of the sodium salts (acetate, bicarbonate, chloride and fluoride) and potassium bicarbonate exhibited both protective and curative activity against G. candidum, significantly reducing the severity of sour rot in comparison to pathogen‐inoculated controls (p < 0.05). Although no curative was observed with ammonium bicarbonate, ammonium carbonate, potassium carbonate, potassium chloride, sodium carbonate or sodium citrate, these salts also demonstrated significant protective activity against sour rot when compared to controls (p < 0.05). In sum, the study findings show that all of the selected salts may be used to control carrot sour rot, except for sodium fluoride, which exhibited phytotoxicity to carrots.     

Evaluation of antifungal activity of carbonate and bicarbonate salts alone or in combination with biocontrol agents in control of citrus green mold

The aim of this research was to determine if the attacks of green mold on orange could be reduced by edible salts alone or in combination with biocontrol agent. For this purpose toxicity to Pantoea digitatum and practical use of sodium carbonate (SC), sodium bicarbonate (SBC) and potassium carbonate, and potassium bicarbonate alone or in combination with antagonistic bacteria (Pseudomonas fluorescens isolate PN, Bacillus subtilis isolate VHN, Pantoea agglomerans isolate CA) to control green mold were determined. All were fungistatic. SC and SBC were equal and superior to the other salts for control of green mold on oranges inoculated 6h before treatment and were chosen for subsequent trails under cold storage conditions. The biocontrol agents were found completely tolerant to 3% sodium bicarbonate and sodium carbonate at room temperature; although their culturability was reduced by > 1000-fold after 60 min in 1% other salt solutions. Satisfactory results were also obtained with the combined treatment for control of green mold. A significant increase in biocontrol activity of all isolate was observed when combined with sodium carbonate and sodium bicarbonate. The treatments comprising CA combined with SB was as effective as fungicide treatment. Thus, use of sodium bicarbonate treatment at 3% followed by the antagonist P. agglomerans CA could be an alternative to chemical fungicides for control of green mold on oranges.     

Carbonate ion-selective electrode with reduced interference from salicylate

A carbonate ion-selective electrode for determination of total carbon dioxide species such as carbon dioxide, bicarbonate and carbonate with reduced interference from salicylate is described. Derivatives of trifluoroacetophenone were used as neutral carriers for carbonate. A polymer-free liquid carbonate-selective membrane with a cellophane outer membrane was found to give a carbonate-selective electrode with a negligible response to salicylate. The electrical contact was obtained by insertion of a silver/silver chloride electrode directly into the liquid membrane. The electrode does not require any aqueous filling solution and is therefore maintenance-free. The response to carbon dioxide species was found to be highly reproducible with a response time of 1-2 min at total carbon dioxide concentrations in the range from 5 to 50 mM. The lifetime of the electrode was at least 3 months. The electrode is regarded as very promising for clinical analysis of carbon dioxide species in body fluids such as plasma and serum.     

Reef corals, zooxanthellae and free-living algae: a microcosm study that demonstrates synergy between calcification and primary production

A mature, high-biodiversity coral reef microcosm and its chambered subsets were used to examine the relationship between calcification and photosynthesis and its most critical biotic components. Whole ecosystem calcification at 4.0±0.2 kg (40±2 mol) CaCO₃ m⁻² year⁻¹ is related to its primary components (stony coral 17.6%, Halimeda 7.4%, Tridacna 9.0%, algal turf, coralline and foraminifera 29.4%, and miscellaneous invertebrates 36%). Through analysis of the microcosm's daily carbonate system, it is demonstrated that bicarbonate ion, not carbonate ion, is the principal component of total alkalinity reduction in the water column (thus, bicarbonate ion is the principal measured component of calcification as normally measured on reef transects). While chamber-isolated free-living algae remove carbon dioxide, and raise pH and carbonate ion equivalent to that in the microcosm as a whole, no total alkalinity reduction (calcification) occurs. On the other hand, chamber isolated stony corals remove considerable bicarbonate, with very little pH or carbonate ion elevation. Combining the non-calcifying free-living macroalgae Chondria with stony corals in chamber subsets, it is possible to remove more carbon dioxide (elevating pH) and thereby increase coral calcification rates by 60 and 120% above zooxanthellae-mediated rates to 20.6 kg (206 mol) and 18.5 kg (185 mol) CaCO₃ m⁻² year⁻¹ for Acropora and Montipora, respectively. These findings, which support the McConnaughey and Whelan hypothesis of bicarbonate ion neutralization in coral calcification, are easily demonstrated in the controlled microcosm environment.     

Effects of variations in carbonate chemistry on the calcification rates of Madracis auretenra (= Madracis mirabilis sensu Wells, 1973): bicarbonate concentrations best predict calcification rates

Physiological data and models of coral calcification indicate that corals utilize a combination of seawater bicarbonate and (mainly) respiratory CO₂ for calcification, not seawater carbonate. However, a number of investigators are attributing observed negative effects of experimental seawater acidification by CO₂ or hydrochloric acid additions to a reduction in seawater carbonate ion concentration and thus aragonite saturation state. Thus, there is a discrepancy between the physiological and geochemical views of coral biomineralization. Furthermore, not all calcifying organisms respond negatively to decreased pH or saturation state. Together, these discrepancies suggest that other physiological mechanisms, such as a direct effect of reduced pH on calcium or bicarbonate ion transport and/or variable ability to regulate internal pH, are responsible for the variability in reported experimental effects of acidification on calcification. To distinguish the effects of pH, carbonate concentration and bicarbonate concentration on coral calcification, incubations were performed with the coral Madracis auretenra (= Madracis mirabilis sensu Wells, 1973) in modified seawater chemistries. Carbonate parameters were manipulated to isolate the effects of each parameter more effectively than in previous studies, with a total of six different chemistries. Among treatment differences were highly significant. The corals responded strongly to variation in bicarbonate concentration, but not consistently to carbonate concentration, aragonite saturation state or pH. Corals calcified at normal or elevated rates under low pH (7.6–7.8) when the seawater bicarbonate concentrations were above 1800 μm . Conversely, corals incubated at normal pH had low calcification rates if the bicarbonate concentration was lowered. These results demonstrate that coral responses to ocean acidification are more diverse than currently thought, and question the reliability of using carbonate concentration or aragonite saturation state as the sole predictor of the effects of ocean acidification on coral calcification.     

Evaluation of Food Additives and Low-toxicity Compounds for the Control of Bean Rust and Wheat Leaf Rust

The efficacy of low‐toxicity chemicals as possible alternatives to synthetic fungicides for the control of Uromyces appendiculatus and Puccinia triticina was evaluated. A preliminary selection of food additives was performed through in vitro and in vivo preliminary screenings. The ED₅₀ and minimum inhibition concentration (MIC) values showed that most of the food additives used in this study were more toxic to U. appendiculatus than to P. triticina. Acetic acid, potassium carbonate, sodium carbonate and sodium molybdate were the food additives that were more toxic to the urediniospores of P. triticina. Selected compounds and concentrations were tested on bean and wheat plants grown in pots under controlled conditions. Acetic acid, ammonium bicarbonate, potassium acetate, potassium benzoate, potassium bicarbonate, potassium carbonate, sodium acetate and sodium citrate at 0.03, 0.09, 0.03, 0.006, 0.012, 0.012, 0.03 and 0.03 m , respectively, significantly reduced the disease severity of U. appendiculatus without causing any injury to bean leaves. Ammonium bicarbonate, potassium bicarbonate, sodium bicarbonate and sodium citrate at 0.12, 0.03, 0.12 and 0.03 m , respectively, were the most effective in reducing the disease severity caused by P. triticina without causing any injury to wheat leaves.     

Zinc deficiency in wetland rice along a toposequence of hydromorphic soils in the Philippines

Summary Zinc deficiency in rice induced by prolonged water saturation of the soil is widespread in the Philippines. The poor drainage associated with the disorder is mostly due to upwelling of shallow artesian water in footslope areas and to ponding of water from continually flowing artesian wells common in such areas. One toposequence where no Zn deficiency occurred in the highest field and progressively severe Zn deficiency in successive lower fields, was studied in detail. With lower elevation soils showed an increase in organic matter and free carbonate, associated with increased upwelling and ponding of artesian water very high in dissolved Mg and Ca bicarbonate.     

An Automated System for Monitoring and Regulating the pH of Bicarbonate Buffers

The bicarbonate buffer is considered as the most biorelevant buffer system for the simulation of intestinal conditions. However, its use in dissolution testing of solid oral dosage forms is very limited. The reason for this is the thermodynamic instability of the solution containing hydrogen carbonate ions and carbonic acid. The spontaneous loss of carbon dioxide (CO₂) from the solution results in an uncontrolled increase of the pH. In order to maintain the pH on the desired level, either a CO₂ loss must be completely avoided or the escaped CO₂ has to be replaced by quantitative substitution, i.e. feeding the solution with the respective amount of gas, which re-acidifies the buffer after dissociation. The present work aimed at the development of a device enabling an automatic pH monitoring and regulation of hydrogen carbonate buffers during dissolution tests.     

Abiotic influences on bicarbonate use in the giant kelp,Macrocystis pyrifera,in the Monterey Bay

In the Monterey Bay region of central California, the giant kelp Macrocystis pyrifera experiences broad fluctuations in wave forces, temperature, light availability, nutrient availability, and seawater carbonate chemistry, all of which may impact their productivity. In particular, current velocities and light intensity may strongly regulate the supply and demand of inorganic carbon (Ci) as substrates for photosynthesis. Macrocystis pyrifera can acquire and utilize both CO₂ and bicarbonate (HCO₃^?) as Ci substrates for photosynthesis and growth. Given the variability in carbon delivery (due to current velocities and varying [DIC]) and demand (in the form of saturating irradiance), we hypothesized that the proportion of CO₂ and bicarbonate utilized is not constant for M. pyrifera, but a variable function of their fluctuating environment. We further hypothesized that populations acclimated to different wave exposure and irradiance habitats would display different patterns of bicarbonate uptake. To test these hypotheses, we carried out oxygen evolution trials in the laboratory to measure the proportion of bicarbonate utilized by M. pyrifera via external CA under an orthogonal cross of velocity, irradiance, and acclimation treatments. Our Monterey Bay populations of M. pyrifera exhibited proportionally higher external bicarbonate utilization in high irradiance and high flow velocity conditions than in sub‐saturating irradiance or low flow velocity conditions. However, there was no significant difference in proportional bicarbonate use between deep blades and canopy blades, nor between individuals from wave‐exposed versus wave‐protected sites. This study contributes a new field‐oriented perspective on the abiotic controls of carbon utilization physiology in macroalgae.     

Bicarbonate produced from carbon capture for algae culture

Using captured CO(2) to grow microalgae is limited by the high cost of CO(2) capture and transportation, as well as significant CO(2) loss during algae culture. Moreover, algae grow poorly at night, but CO(2) cannot be temporarily stored until sunrise. To address these challenges, we discuss a process where CO(2) is captured as bicarbonate and used as feedstock for algae culture, and the carbonate regenerated by the culture process is used as an absorbent to capture more CO(2). This process would significantly reduce carbon capture costs because it does not require additional energy for carbonate regeneration. Furthermore, not only would transport of the aqueous bicarbonate solution cost less than for that of compressed CO(2), but using bicarbonate would also provide a superior alternative for CO(2) delivery to an algae culture system.     

Potential role for microbial ureolysis in the rapid formation of carbonate tufa mounds

Modern carbonate tufa towers in the alkaline (~pH 9.5) Big Soda Lake (BSL), Nevada, exhibit rapid precipitation rates (exceeding 3 cm/year) and host diverse microbial communities. Geochemical indicators reveal that carbonate precipitation is, in part, promoted by the mixing of calcium-rich groundwater and carbonate-rich lake water, such that a microbial role for carbonate precipitation is unknown. Here, we characterize the BSL microbial communities and evaluate their potential effects on carbonate precipitation that may influence fast carbonate precipitation rates of the active tufa mounds of BSL. Small subunit rRNA gene surveys indicate a diverse microbial community living endolithically, in interior voids, and on tufa surfaces. Metagenomic DNA sequencing shows that genes associated with metabolisms that are capable of increasing carbonate saturation (e.g., photosynthesis, ureolysis, and bicarbonate transport) are abundant. Enzyme activity assays revealed that urease and carbonic anhydrase, two microbial enzymes that promote carbonate precipitation, are active in situ in BSL tufa biofilms, and urease also increased calcium carbonate precipitation rates in laboratory incubation analyses. We propose that, although BSL tufas form partially as a result of water mixing, tufa-inhabiting microbiota promote rapid carbonate authigenesis via ureolysis, and potentially via bicarbonate dehydration and CO₂ outgassing by carbonic anhydrase. Microbially induced calcium carbonate precipitation in BSL tufas may generate signatures preserved in the carbonate microfabric, such as stromatolitic layers, which could serve as models for developing potential biosignatures on Earth and elsewhere.     

Direct EPR detection of the carbonate radical anion produced from peroxynitrite and carbon dioxide

The biological effects of peroxynitrite have been recently considered to be largely dependent on its reaction with carbon dioxide, which is present in high concentrations in intra- and extracellular compartments. Peroxynitrite anion (ONOO-) reacts rapidly with carbon dioxide, forming an adduct, nitrosoperoxocarboxylate (ONOOCO2-), whose decomposition has been proposed to produce reactive intermediates such as the carbonate radical (CO-3). Here, by the use of rapid mixing continuous flow electron paramagnetic resonance (EPR), we directly detected the carbonate radical in flow mixtures of peroxynitrite with bicarbonate-carbon dioxide over the pH range of 6-9. The radical was unambiguously identified by its EPR parameters (g = 2.0113; line width = 5.5 G) and by experiments with bicarbonate labeled with 13C. In this case, the singlet EPR signal obtained with 12C bicarbonate splits into the expected doublet because of 13C (a(13C)= 11.7 G). The singlet spectrum of the unlabeled radical was invariant between pH 6 and 9, confirming that in this pH range the detected radical is the carbonate radical anion (CO-3). Importantly, in addition to contributing to the understanding of nitrosoperoxocarboxylate decomposition pathways, this is the first report unambiguously demonstrating the formation of the carbonate radical anion at physiological pHs by direct EPR spectroscopy.     

A new method for the concentration of Cryptosporidium oocysts from water

A novel method for the concentration of Cryptosporidium oocysts from water has been developed, based upon the precipitation of calcium carbonate. A 10 1 water sample is treated by adding solutions of calcium chloride and sodium bicarbonate and raising the pH value to 10 with sodium hydroxide. Crystals of calcium carbonate form and enmesh particles in the Cryptosporidium oocyst size range. The crystals are allowed to settle, the supernatant fluid is discarded and the calcium carbonate precipitate dissolved in sulphamic acid. The sample can be concentrated further by centrifugation. Recoveries of oocysts from seeded samples of deionized, tap and river water were in excess of 68%.     

Characterization of crystalline structures in Opuntia ficus-indica

This research studies the crystalline compounds present in nopal (Opuntia ficus-indica) cladodes. The identification of the crystalline structures was performed using X-ray diffraction, scanning electron microscopy, mass spectrometry, and Fourier transform infrared spectroscopy. The crystalline structures identified were calcium carbonate (calcite) [CaCO₃], calcium-magnesium bicarbonate [CaMg(CO₃)₂], magnesium oxide [MgO], calcium oxalate monohydrate [Ca(C₂O₄)•(H₂O)], potassium peroxydiphosphate [K₄P₂O₈] and potassium chloride [KCl]. The SEM images indicate that calcite crystals grow to dipyramidal, octahedral-like, prismatic, and flower-like structures; meanwhile, calcium-magnesium bicarbonate structures show rhombohedral exfoliation and calcium oxalate monohydrate is present in a drusenoid morphology. These calcium carbonate compounds have a great importance for humans because their bioavailability. This is the first report about the identification and structural analysis of calcium carbonate and calcium-magnesium bicarbonate in nopal cladodes, as well as the presence of magnesium oxide, potassium peroxydiphosphate and potassium chloride in these plants. The significance of the study of the inorganic components of these cactus plants is related with the increasing interest in the potential use of Opuntia as a raw material of products for the food, pharmaceutical, and cosmetic industries.     

Carbonic anhydrase inhibitors: Inhibition studies of a coral secretory isoform with inorganic anions

The inhibition of a coral carbonic anhydrase (CA, EC 4.2.1.1) has been investigated with a series of inorganic anions such as halogenides, pseudohalogenides, bicarbonate, carbonate, nitrate, nitrite, hydrogen sulfide, bisulfite, perchlorate, sulfate. The full-length scleractinian coral Stylophora pistillata CA, STPCA, has a significant catalytic activity for the physiological reaction of CO₂ hydration to bicarbonate, similarly to the ubiquitous human isoforms hCA I (cytosolic) and hCA VI (secreted). The best STPCA anion inhibitors were bromide, iodide, carbonate, and sulfamate, with inhibition constants of 9.0–10.0 μM.     

Carbonic anhydrase inhibitors. Inhibition of the newly isolated murine isozyme XIII with anions

The inhibition of the newly discovered cytosolic carbonic anhydrase (CA, EC 4.2.1.1) isozyme XIII of murine origin (mCA XIII) has been investigated with a series of anions, such as the physiological ones (bicarbonate, chloride), or the metal complexing anions (cyanate, cyanide, azide, hydrogen sulfide, etc), nitrate, nitrite, sulfate, sulfamate, sulfamide as well as with phenylboronic and phenylarsonic acids. The best mCA XIII inhibitors were cyanate, thiocyanate, cyanide and sulfamide, with K(I)-s in the range of 0.25microM-0.74 mM, whereas fluoride, iodide, azide, carbonate and hydrogen sulfide were less effective (K(I)-s in the range of 3.0-5.5mM). The least effective inhibitors were sulfate, chloride and bicarbonate (K(I)-s in the range of 138-267 mM). The affinity of mCA XIII for anions is very different from that of the other cytosolic isozymes (hCA I and II) or the mitochondrial isozyme hCA V. This resistance to inhibition by the physiological anions bicarbonate and chloride suggests an evolutionary adaptation of CA XIII to the presence of high concentrations of such anions (e.g., in the reproductive tract of both female and male), and the possible participation of this isozyme (similarly to CA II, CA IV and CA V) in metabolons with proteins involved in the anion exchange and transport, such as the anion exchangers (AE1-3) or the sodium bicarbonate co-transporter (NBC1 and NBC3) proteins, which remain to be identified.     

Increase of atmospheric CO2 promotes phytoplankton productivity

It is usually thought that unlike terrestrial plants, phytoplankton will not show a significant response to an increase of atmospheric CO₂. Here we suggest that this view may be biased by a neglect of the effects of carbon (C) assimilation on the pH and the dissociation of the C species. We show that under eutrophic conditions, productivity may double as a result of doubling of the atmospheric CO₂ concentration. Although in practice productivity increase will usually be less, we still predict a productivity increase of up to 40% in marine species with a low affinity for bicarbonate. In eutrophic freshwater systems doubling of atmospheric CO₂ may result in an increase of the productivity of more than 50%. Freshwaters with low alkalinity appeared to be very sensitive to atmospheric CO₂ elevation. Our results suggest that the aquatic C sink may increase more than expected, and that nuisance phytoplankton blooms may be aggravated at elevated atmospheric CO₂ concentrations.