Age‐related variation in the trophic characteristics of a marsupial carnivore,the Tasmanian devil Sarcophilus harrisii

Age‐related changes in diet have implications for competitive interactions and for predator–prey dynamics, affecting individuals and groups at different life stages. To quantify patterns of variation and ontogenetic change in the diets of Tasmanian devils Sarcophilus harrisii, a threatened marsupial carnivore, we analyzed variation in the stable isotope composition of whisker tissue samples taken from 91 individual devils from Wilmot, Tasmania from December 2014 to February 2017. Both δ¹³C and δ¹⁵N decreased with increasing age in weaned Tasmanian devils, indicating that as they age devils rely less on small mammals and birds, and more on large herbivores. Devils <12 months old had broader group isotopic niches, as estimated by Bayesian standard ellipses (SEA_B mode = 1.042) than devils from 12 to 23 months old (mode = 0.541) and devils ≥24 months old (mode = 0.532). Devils <24 months old had broader individual isotopic niches (SEA_B mode range 0.492–1.083) than devils ≥24 months old (mode range 0.092–0.240). A decrease in δ¹⁵N from the older whisker sections to the more recently grown sections in devils <24 months old likely reflects the period of weaning in this species, as this pattern was not observed in devils ≥24 months old. Our data reveal changes in the isotopic composition of devil whiskers with increasing age, accompanied by a reduction in isotopic variation both among population age classes and within individuals, reflecting the effect of weaning in early life, and a likely shift from an initially diverse diet of small mammals, birds, and invertebrates towards increasing consumption of larger herbivores in adulthood.     

生态系统光合与呼吸拆分的同位素理论、方法和应用进展

生态系统光合和呼吸是构成净生态系统CO₂交换量(NEE)的重要组分。涡度相关技术可直接观测生态系统NEE,并通过建立温度回归或光响应曲线等函数将NEE统计拆分为生态系统光合和呼吸,但是存在自相关和高估白天呼吸等问题。稳定同位素红外光谱技术的进步使高时间分辨率大气CO₂及其稳定碳同位素组成(δ¹³C)的连续观测成为可能,与涡度相关技术观测的NEE数据相结合,可实现昼夜和季节尺度生态系统光合和呼吸拆分。本文系统阐述了生态系统光合与呼吸的同位素通量拆分方法的基本理论与假设,阐述了同位素通量观测技术的发展及其应用进展,综述了同位素通量拆分理论解析生态系统光合与呼吸过程的新机制认识,最后总结并展望了同位素通量拆分理论的不确定性以及开展多种拆分方法综合比较的必要性。     

Application of isotopic ratio mass spectrometry for the in vitro determination of demethylation activity in human liver microsomes using N-methyl-13C-labeled substrates

The reaction of demethylation mediated by cytochrome P450 (CYP) leads to the equimolar production of demethylated metabolite and formaldehyde. From a 13C-substrate labeled on a carbon of the methyl moiety, [13C]formaldehyde (H13CHO) is liberated. A highly sensitive and specific assay involving the oxidation of H13CHO to 13CO(2) by a double-enzymatic-step reaction is reported. The 13CO(2) was quantified by the method of reverse isotopic dilution based on gas chromatography-isotope ratio mass spectrometry analysis. The method first involves the limiting step of the CYP-dependent reaction, which is stopped with a mixture of zinc sulfate 5 mM and trichloroacetic acid 100 mM. Then, the transformation of H13CHO to 13CO(2) is performed with the formaldehyde (0.2 unit) and the formate (0.2 unit) dehydrogenase NAD-dependent enzymes. The recovery of 13CO(2) from the incubation mixture was equal to 91.4 +/- 3.0%. The accuracy and the precision of the present method were within 12 and 10%, respectively. The limit of quantification was set to 25 pmol. The performance of the assay was validated on human liver microsomes with five probes: [13C]erythromycin, [1-13C]caffeine, [3-13C]caffeine, [7-13C]caffeine, and [13C(2)]aminopyrine. This method is useful for the rapid determination of N-demethylase activity of human liver microsomes from methyl-13C-substrates.     

Geochemical Peculiarities of the Carbonate Constructions Formed during Microbial Oxidation of Methane under Anaerobic Conditions

The aragonite constructions of the Black Sea are formed in a stable anaerobic zone and are a perfect object to study the natural mechanism of anaerobic methane oxidation. The most probable pathway of methane oxidation is its methanogen-mediated reaction with bicarbonates, dissolved in seawater, with the formation of water and acetate, which is then consumed by other components of the anaerobic community. Comparison of the ¹³C values of carbonate minerals and organic matter once more demonstrated that the formation of the organic matter of biomass is accompanied by intense fractionation of carbon isotopes, as a result of which the total organic matter of biomass acquires an extremely light isotopic composition, characterized by ¹³C values as low as –83.8.     

Regulation of AE2-mediated Cl- transport by intracellular or by extracellular pH requires highly conserved amino acid residues of the AE2 NH2-terminal cytoplasmic domain

We reported recently that regulation by intracellular pH (pH(i)) of the murine Cl-/HCO(3)(-) exchanger AE2 requires amino acid residues 310-347 of the polypeptide's NH(2)-terminal cytoplasmic domain. We have now identified individual amino acid residues within this region whose integrity is required for regulation of AE2 by pH. 36Cl- efflux from AE2-expressing Xenopus oocytes was monitored during variation of extracellular pH (pH(o)) with unclamped or clamped pH(i), or during variation of pH(i) at constant pH(o). Wild-type AE2-mediated 36Cl- efflux was profoundly inhibited by acid pH(o), with a value of pH(o50) = 6.87 +/- 0.05, and was stimulated up to 10-fold by the intracellular alkalinization produced by bath removal of the preequilibrated weak acid, butyrate. Systematic hexa-alanine [(A)6]bloc substitutions between aa 312-347 identified the greatest acid shift in pH(o(50)) value, approximately 0.8 pH units in the mutant (A)6 342-347, but only a modest acid-shift in the mutant (A)6 336-341. Two of the six (A)6 mutants retained normal pH(i) sensitivity of 36Cl- efflux, whereas the (A)6 mutants 318-323, 336-341, and 342-347 were not stimulated by intracellular alkalinization. We further evaluated the highly conserved region between aa 336-347 by alanine scan and other mutagenesis of single residues. Significant changes in AE2 sensitivity to pH(o) and to pH(i) were found independently and in concert. The E346A mutation acid-shifted the pH(o(0) value to the same extent whether pH(i) was unclamped or held constant during variation of pH(o). Alanine substitution of the corresponding glutamate residues in the cytoplasmic domains of related AE anion exchanger polypeptides confirmed the general importance of these residues in regulation of anion exchange by pH. Conserved, individual amino acid residues of the AE2 cytoplasmic domain contribute to independent regulation of anion exchange activity by pH(o) as well as pH(i).     

Tracing mangrove carbon in suspended matter and aquatic fauna of the Gautami–Godavari Delta,Bay of Bengal (India)

Stable carbon isotopic composition and C/N ratio were used to trace the input of carbon associated with mangrove litter into the estuary of the Godavari–Gautami delta system and Kakinada bay (Andhra Pradesh, India). Suspended organic matter in the mangrove channels was more depleted in ¹³C (average ¹³C = –24.5) than in Kakinada bay which showed ¹³C values for suspended matter (average ¹³C = –22.7) closer to those expected for marine phytoplankton. Suspended organic matter from mangrove channels was enriched in nitrogen (average C/N atom ratio 12.7) and ¹³C (average ¹³C = –24.5) relative to mangrove leaf litter, which had a C/N ratio of 75 and a ¹³C value of –28. Lowest C/N ratios for suspended matter were observed during southwest monsoon when rainfall was highest. Although in general, mangrove litter fall was also lower during this period, no clear correlation was observed between litter fall and C/N ratio of suspended matter. In general, the composition of suspended matter pointed towards phytoplankton as a major component. Isotopic composition of zooplankton suggested selective feeding on ¹³C-enriched, marine phytoplankton in open Kakinada bay and on ¹³C-depleted organic matter, such as estuarine phytoplankton and mangrove litter, in the mangrove channels. From the ¹³C signature, it appeared that mangrove carbon was present to some extent in zooplankton and macrofauna from the mangrove mudflats and channels, but the signal rapidly decreased in Kakinada bay. Nitrogen isotopic composition of zooplankton and macrofauna indicated a progressive enrichment of ¹⁵N away from the mangrove forest towards the northern part of Kakinada bay, in approach of Kakinada city. This is thought to reflect input of anthropogenic nitrogen enriched in ¹⁵N and subsequent uptake of this enriched nitrogen into the aquatic food chain.     

Variations in the natural abundance of 15N in ryegrass/white clover shoot material as influenced by cattle grazing

Biological N2 fixation in clover is an important source of N in low external-N input farming systems. Using the natural 15N-abundance method, variations in N2 fixation were investigated in grazed and mowed plots of a ryegrass/white clover field. Ryegrass 15N varied considerably, from 0.2 to 5.6 under mowed conditions and from –3.3 to 11.6 under grazed conditions. Variations in 15N white clover were lower than in ryegrass, especially in the mowed plots (SE = 0.05, n = 20). The variations in the percentage of nitrogen derived from the atmosphere (%Ndfa) in white clover were highest in the grazed plots where it ranged from 12 to 96% (mean = 64%) compared with the mowed plots where it ranged from 64 to 92% (mean = 79%). Thus, the N2 fixation per unit white clover DM in the grazed ley was lower and more variable than under mowing conditions.Urine from dairy cows equivalent to 0, 200, 400 and 800 kg N ha-1 was applied to a ryegrass/white clover plot 6, 4 or 2 weeks before harvest. Without urine application 15N of ryegrass was positive. By increasing urine application (15N = –1) two weeks before sampling, the 15N of ryegrass decreased strongly to about –7 (P < 0.001). However, this effect was only observed when urine was applied two weeks before sampling. When applying 800 kg N four and six weeks before sampling, 15N in ryegrass was not significantly different from the treatment without urine application. White clover 15N was unaffected by whatever changes occurred in 15N of the plant-available soil N pool (reflected in 15N of ryegrass). This indicates that within the time span of this experiment, N2 fixation per unit DM was not affected by urine. Therefore, newly deposited urine may not be the main contributing factor to the variation in %Ndfa found in the grazed fields. This experiment suggested that the natural abundance method can be applied for estimating %Ndfa without disturbance in natural animal-grazed systems.     

Nitrogen partitioning and isotopic discrimination are affected by age and dietary protein content in growing lambs

It is difficult to separate an age-dependent fall in nitrogen use efficiency (NUE; N balance/N intake) in growing ruminants from a progressively decrease in animal protein requirements over time. This study examined the effect of dietary protein content on N partitioning, digestibility and N isotopic discrimination between the animal and its diet (Δ¹⁵N_animal-diet) evaluated at two different fattening periods (early v. late). Twenty-four male Romane lambs (age: 19 ± 4.0 days; BW: 8.3 ± 1.39 kg) were equally allocated to three dietary CP treatments (15%, 17% and 20% CP on a DM basis). Lambs were reared with their mothers until weaning, thereafter housed in individual pens until slaughter (45 kg BW). During the post-weaning period, lambs were allocated twice (early fattening (30 days post-weaning) and late fattening (60 days post-weaning)) to metabolic cages for digestibility and N balance study. When diet CP content increased, the average daily gain of lambs increased (P < 0.05) while the age at slaughter decreased (P = 0.01), but no effect was observed on feed efficiency (P > 0.10). Diet CP content had limited effect on lamb carcass traits. Higher fibre digestibility was observed at the early v. late fattening period (P < 0.001). The N intake and the urinary N excretion increased when diet CP content increased (P < 0.001) and when shifting from early to late fattening period (P < 0.001). Faecal N excretion (P = 0.14) and N balance (P > 0.10) were not affected by diet CP content. Nitrogen digestibility increased (P < 0.001) as the diet CP content increased and on average it was greater at late v. early fattening period (P = 0.02). The NUE decreased (P = 0.001) as the diet CP content increased and as the lamb became older (P < 0.001). However, the age-dependent fall in NUE observed was lower at high v. low dietary CP content (CP × age interaction; P = 0.04). The Δ¹⁵N_animal-diet was positively correlated (P < 0.05) with N intake (r = 0.59), excretion of faecal N (r = 0.41), urinary N (r = 0.69) and total manure N (r = 0.64), while negatively correlated with NUE (r = −0.57). Overall, the experiment showed NUE was lower in older lambs and when lambs were fed high diet CP content, and that Δ¹⁵N_animal-diet was a useful indicator not only for NUE but also for urinary N excretion, which is a major environmental pollution factor on farm.     

Isotopic Fractionation of Hydrogen in Plants

The naturally-occurring stable isotopes deuterium and hydrogen are fractionated by a number of physical and biological processes. Deuterium has a tendency to precipitate out first from a moist air mass. Thus ground water will become isotopically lighter with an increase in latitude, altitude, or distance inland. Water taken up by the plant from the soil undergoes little change until evapotranspiration results in leaf water becoming isotopically heavier. Thus hydrogen isotopes in plants can reveal something of geography (groundwater) and climate. Hydrogen isotopes undergo little fractionation by passage through the food chain, although plant parasites tend to be enriched in D as compared to their hosts, possibly due to higher rates of transpiration in the parasitic plants. The splitting of water in photosynthesis results in the lighter isotope being incorporated into organic matter. An even larger isotopic fractionation results during lipid synthesis and other processes involving the pyruvate dehydrogenase complex. Differences in metabolic pathway between species can be detected by D/H ratios. Hydrogen isotopic differences can be detected between CAM, C₄, and C₃ species. Within C₄ plants, the NADP-ME plants are isotopically distinguishable from NAD-ME and PEP-CK plants.