Ammonia (NH₃) emissions should be mitigated to improve environmental quality.  Croplands are one of the largest NH₃ sources, they must be managed properly to reduce their emissions while achieving the target yields.  Herein, we report the NH₃ emissions, crop yield and changes in soil fertility in a long-term trial with various fertilization regimes, to explore whether NH₃ emissions can be significantly reduced using the 4R nutrient stewardship (4Rs), and its interaction with the organic amendments (i.e., manure and straw) in a wheat–maize rotation.  Implementing the 4Rs significantly reduced NH₃ emissions to 6 kg N ha^–1 yr^–1 and the emission factor to 1.72%, without compromising grain yield (12.37 Mg ha^–1 yr^–1) and soil fertility (soil organic carbon of 7.58 g kg^–1) compared to the conventional chemical N management.  When using the 4R plus manure, NH₃ emissions (7 kg N ha^–1 yr^–1) and the emission factor (1.74%) were as low as 4Rs, and grain yield and soil organic carbon increased to 14.79 Mg ha^–1 yr^–1 and 10.09 g kg^–1, respectively.  Partial manure substitution not only significantly reduced NH₃ emissions but also increased crop yields and improved soil fertility, compared to conventional chemical N management.  Straw return exerted a minor effect on NH₃ emissions.  These results highlight that 4R plus manure, which couples nitrogen and carbon management can help achieve both high yields and low environmental costs.

    The vacuolar proton pump ATPase (V-H⁺-ATPase), which is a multi-subunit membrane protein complex, plays a major role in the activation of ion and nutrient transport and has been suggested to be involved in several physiological processes, such as cell expansion and salt tolerance. In this study, three genes encoding V-H⁺-ATPase subunits B (ScVHA-B, GenBank: JF826506), C (ScVHA-C, GenBank: JF826507) and H (ScVHA-H, GenBank: JF826508) were isolated from the halophyte Suaeda corniculata. The transcript levels of ScVHA-B, ScVHA-C and ScVHA-H were increased by salt, drought and saline-alkali treatments. V-H⁺-ATPase activity was also examined under salt, drought and saline-alkali stresses. The results showed that V-H⁺-ATPase activity was correlated with salt, drought and saline-alkali stress. Furthermore, V-H⁺-ATPase subunits B, C and H (ScVHA-B, ScVHA-C and ScVHA-H) from S. corniculata were introduced separately into the alfalfa genome. The transgenic alfalfa was verified by Southern and Northern blot analysis. During salt and saline-alkali stresses, transgenic linevacuolar proton pump, salt tolerance, saline-alkali tolerance, alfalfa
s carrying the B, C and H subunits had higher germination rates than the wild type (WT). More free proline, higher superoxide dismutase (SOD) activity and lower malondialdehyde (MDA) levels were detected in the transgenic plants under salt and saline-alkali treatments. Moreover, the ScVHA-B transgenic lines showed greater tolerance to salt and saline-alkali stresses than the WT. These results suggest that overexpression of ScVHA-B, ScVHA-C and ScVHA-H improves tolerance to salt and saline-alkali stresses in transgenic alfalfa.

    Carboxylesterase is a multifunctional superfamily and can be found in almost all living organisms. As the metabolic enzymes, carboxylesterases are involved in insecticides resistance in insects for long time. In our previous studies, the enhanced carboxylesterase activities were found in the chlorantraniliprole resistance strain of diamondback moth (DBM). However, the related enzyme gene of chlorantraniliprole resistance has not been clear in this strain. Here, a full-length cDNA of carboxylesterase pxCCE016b was cloned and exogenously expressed in Escherichia coli at the first time, which contained a 1 693 bp open reading frame (ORF) and encoded a protein of 542 amino acids. Sequence analysis showed that this cDNA has a predicted mass of 61.56 kDa and a theoretical isoelectric point value of 5.78. The sequence of deduced amino acid possessed the classical structural features: a type-B carboxylesterase signature 2 (EDCLYLNVYTK), a type-B carboxylesterase serine active site (FGGDPENITIFGESAG) and the catalytic triad (Ser186, Glu316, and His444). The real-time quantitative PCR (qPCR) analysis showed that the expression level of the pxCCE016b was significantly higher in the chlorantraniliprole resistant strain than in the susceptible strain. Furthermore, pxCCE016b was highly expressed in the midgut and epidermis of the DBM larvae. When the 3rd-instar larvae of resistant DBM were exposed to abamectin, alpha-cypermethrin, chlorantraniliprole, spinosad, chlorfenapyr and indoxacarb insecticides, the up-regulated expression of pxCCE016b was observed only in the group treated by chlorantraniliprole. In addition, recombinant vector pET-pxCCE016b was constructed with the most coding region (1 293 bp) and large number of soluble recombinant proteins (less than 48 kDa) were expressed successfully with prokaryotic cell. Western blot analysis showed that it was coded by pxCCE016b. All the above findings provide important information for further functional study, although we are uncertainty whether the pxCCE016b gene is actually involved in chlorantraniliprole resistance.

The insecticide chlorantraniliprole exhibits good efficacy and plays an important role in controlling the diamondback moth, Plutella xylostella Linnaeus. However, resistance to chlorantraniliprole has been observed recently in some field populations. At present study, diamondback moths with resistance to chlorantraniliprole (resistant ratio (RR) was 82.18) for biochemical assays were selected. The assays were performed to determine potential resistance mechanisms. The results showed that the selected resistant moths (GDLZ-R) and susceptible moth could be synergized by known metabolic inhibitors such as piperonyl butoxide (PBO), triphenyl phosphate (TPP) and diethyl-maleate (DEM) at different levels (1.68-5.50-fold and 2.20-2.89-fold, respectively), and DEM showed the maximum synergism in both strains. In enzymes assays, a high level of glutathione-S-transferase (GST) was observed in the resistant moth, in contrast, moths that are susceptible to the insecticide had only 1/3 the GST activity of the resistant moths. The analysis of short-term exposure of chlorantraniliprole on biochemical response in the resistant strain also showed that GST activity was significantly elevated after exposure to a sub-lethal concentration of chlorantraniliprole (about 1/3 LC50, 12 mg L-1) 12 and 24 h, respectively. The results show that there is a strong correlation between the enzyme activity and resistance, and GST is likely the main detoxification mechanism responsible for resistance to chlorantraniliprole in P. xylostella L., cytochrome P450 monooxygenase (P450) and carboxy-lesterase (CarE) are involved in to some extent.