Purification and characterization of two anionic trypsins from the hepatopancreas of carp

SUMMARY: Two trypsins, designated as trypsin A and trypsin B, have been purified from the hepatopancreas of carp. The purification procedures consisted of ammonium sulfate fractionation, and chromatographies on DEAE-Sephacel, Ultrogel AcA54 and Q-Sepharose. Trypsin A was purified to homogeneity with the molecular mass of approximately 28 kDa, while trypsin B gave two close bands of 28.5 kDa and 28 kDa on sodium dodecylsulfate polyacrylamide gel electrophoresis both under reducing and non-reducing conditions. On native-PAGE, both trypsin A and trypsin B showed a single band. Trypsin A and trypsin B revealed optimum temperature of 40°C and 45°C, respectively, and shared the same optimum pH 9.0 using Boc-Phe-Ser-Arg-MCA as substrate. Both enzymes were effectively inhibited by trypsin inhibitors and their susceptibilities were similar. The NH₂-terminal amino acid sequences of trypsin A and trypsin B were determined to 37th and 40th amino acid residue, respectively. Their sequences were very homologous, but not identical to that of a trypsin-type serine proteinase from carp muscle and these of other trypsins. Immunoblotting test using the antibody raised against trypsin A cross-reacted with trypsin B positively.     

Improvement in recruitment of Japanese sardine with delays of the spring phytoplankton bloom in the Sea of Japan

To evaluate the impact of temporal variation of primary productivity on the recruitment of Japanese sardine (Sardinops melanostictus) in the Sea of Japan, the phenology of sea surface phytoplankton abundance was estimated from 8 day multiple satellite (SeaWiFS, MODIS‐Aqua, MERIS, and VIIRS) derived sea surface chlorophyll (SSChl) a concentrations from January 1998 to December 2015. Because relationships between SSChl a and in situ chlorophyll a concentrations were significantly different among periods based on the satellite combinations used, maximum and minimum SSChl a concentrations of 1 year were relativized as 1 and 0, respectively. Spatio‐temporal variation of relativized SSChl a concentrations was determined by using empirical orthogonal function (EOF) analysis. Scores in the first EOF mode denoted the basin‐scale variations of SSChl a concentrations in the Sea of Japan, and the major peak from the end of February to the end of May displayed the spring bloom. The logarithm of recruitment per spawner (LNRPS) for sardine was positively affected by delays in the start and end dates of the spring phytoplankton bloom. The delay of the date of the lowest sea surface temperature contributed to the delay of the end of the spring bloom during the period 1998–2015 and elevated the LNRPS during the period 1982–2015. Sardine spawns in the southern Sea of Japan from April to May, hence, delays of the spring bloom prolonged its overlap with sardine larval periods, and thus improved the recruitment of Japanese sardine in the Sea of Japan.     

Regression tree analysis of the relationship between the concentrations of antimicrobial components and the microbiota of normal milk from dairy cows

The purpose of this study was to determine the concentrations of antimicrobial components (immunoglobulin A (IgA), lactoferrin (LF), lingual antimicrobial peptide (LAP), and S100A7) in normal milk and their relation to host factors (Age, somatic cell count (SCC), days in milk, richness, and alpha diversity of the milk microbiota) in dairy cows using multivariate regression tree analyses, and to clarify how the milk microbiota is related to the obtained results. Thirty normal milk samples were collected from a commercial dairy farm in June 2020. The thresholds that predicted the concentration of each antimicrobial component in milk were obtained by regression tree analysis, and the beta-diversity of the milk microbiota composition between groups divided according to each threshold was compared by an analysis of similarities test. The IgA and LF concentrations were mainly predicted by the SCC (177,500 and 70,000 cells/ml, respectively), and the LAP and S100A7 concentrations were predicted by Age (29.667 and 40.3 months, respectively). No relationship was observed between the concentration of IgA, LAP, or S100A7 and the milk microbiota composition between the groups divided by the threshold for prediction, but the milk microbiota composition was significantly different between the groups divided by the threshold for predicting the LF concentration. Our results indicated that the LF concentration in normal milk may be associated with the milk microbiota composition.     

Nutrient limitations to bacterial and fungal growth during cellulose decomposition in tropical forest soils

Nutrients constrain the soil carbon cycle in tropical forests, but we lack knowledge on how these constraints vary within the soil microbial community. Here, we used in situ fertilization in a montane tropical forest and in two lowland tropical forests on contrasting soil types to test the principal hypothesis that there are different nutrient constraints to different groups of microorganisms during the decomposition of cellulose. We also tested the hypotheses that decomposers shift from nitrogen to phosphorus constraints from montane to lowland forests, respectively, and are further constrained by potassium and sodium deficiency in the western Amazon. Cellulose and nutrients (nitrogen, phosphorus, potassium, sodium, and combined) were added to soils in situ, and microbial growth on cellulose (phospholipid fatty acids and ergosterol) and respiration were measured. Microbial growth on cellulose after single nutrient additions was highest following nitrogen addition for fungi, suggesting nitrogen as the primary limiting nutrient for cellulose decomposition. This was observed at all sites, with no clear shift in nutrient constraints to decomposition between lowland and montane sites. We also observed positive respiration and fungal growth responses to sodium and potassium addition at one of the lowland sites. However, when phosphorus was added, and especially when added in combination with other nutrients, bacterial growth was highest, suggesting that bacteria out-compete fungi for nitrogen where phosphorus is abundant. In summary, nitrogen constrains fungal growth and cellulose decomposition in both lowland and montane tropical forest soils, but additional nutrients may also be of critical importance in determining the balance between fungal and bacterial decomposition of cellulose.     

Biochar improves sediment microbial fuel cell performance in low conductivity freshwater sediment

Purpose

The low conductivity of sediments for mass and electron transport is the most severe limiting factor in sediment microbial fuel cells (SMFCs), so that sediment ameliorations yielded more remarkable effects than electrode improvements. The objective of this research was to enhance the electricity generation of SMFCs with amendments of biochar to freshwater sediments for conductivity enhancement.

Materials and methods

Laboratory-scale SMFCs were constructed and biochars were produced from coconut shells at different temperatures. Variations in the power output, electrode potential, internal resistance, total organic carbon (TOC) content, and microbial communities were measured.

Results and discussion

Amending with biochar reduced the charge transfer resistances of SMFCs and enriched the Firmicutes (mainly Fusibacter sp.) in the sediment, which improved the SMFC power generation by two- to tenfold and enhanced the TOC removal rate by 1.7- to fourfold relative to those without the amendment.

Conclusions

The results suggested that biochar amendment is a promising strategy to enhance SMFC power production, and the electrical conductivity of biochar should be considered important when interpreting the impact biochar has on the electrical performance of soil or freshwater sediment MFCs.

     

Influences of indigenous phototrophs on methane emissions from a straw-amended paddy soil

A pot experiment was conducted to investigate the influences of indigenous phototrophs on methane (CH₄) emissions from a paddy soil where rice straw was incorporated or was surface-applied. During the cultivation, half of the pots were covered with aluminum foil, except for the minimum space for rice plants, to prevent ambient light reaching the floodwater or the soil surface. Growth of oxygen-producing phototrophs was hardly observed in the unilluminated plots, whereas intensive growth of algae, duckweed and hydrophytes was found in the illuminated ones. Plant growth was not affected by the different treatments. Seasonal changes in CH₄ emission determined by a closed chamber method indicated that illumination had no or only minor effects on CH₄ emissions when rice straw was incorporated or was not applied, but significantly reduced CH₄ emissions when rice straw was surface-applied. Methanogenesis occurring in the soil-floodwater interface was further investigated in two lab-scale model experiments measuring methanogenic activity. As a result, more activated methanogenesis was found in the surface-applied rice straw and the soil around the straw compared with the soil incubated without rice straw. The magnitude of the methanogenic activity in the rice straw incubated under illuminated conditions was significantly lower than that incubated in the dark. Consequently, this study demonstrates that methanogenesis in paddy soil occurs even in the soil-floodwater interface if plant residues like rice straw exist, and such methanogenesis is likely to be suppressed by growth of indigenous phototrophs under illumination.     

A bayesian model for the analysis of lactation curves of dairy goats

This article aims to determine the effect of certain covariates, such as season of kidding, parity, and time of kidding on the characteristics of the lactation curve of Saanen dairy goats. Characteristics investigated are peak milk yield, time of peak milk yield, total milk production, persistency, and the relationship between fat and protein in milk composition, as well as between lactation curves of the same animal in successive years. The analysis is carried out using a hierarchical Bayesian approach, together with Wood’s model, to model lactation. Posterior distributions of quantities of interest are obtained by means of the Markov chain Monte Carlo (MCMC) methods. These clearly illustrate the significant effect of especially parity, but also season and time of kidding on the characteristics of the lactation curve. Total and peak milk yield increase with increasing parity up to about the third or fourth parity, while peak yield is later for first than for later parities. The analysis also enables estimation of lactation characteristics of untested animals, prediction of future characteristics and identification of exceptional animals.     

Effect of bensulfuron-methyl (a sulfonylurea herbicide) on the soil bacterial community of a paddy soil microcosm

The effect of bensulfuron-methyl (BSM) on a soil microbial community in a model paddy microcosm was studied. Total bacterial numbers in the overlying water and surface soil were monitored for 2 months after the application of BSM at the field rate and a ten-fold field rate. Pentachlorophenol (PCP) was used for comparison. Neither chemical affected the total bacterial numbers remarkably, either in the overlying water or in the surface soil. In contrast, the nitrification potential was significantly suppressed by the BSM application. The bacterial community structure, as evaluated by the denaturing gradient gel electrophoresis (DGGE) of PCR amplification products from bacterial 16S rDNA, was unaffected by the BSM treatments over 8 weeks in the surface soil, compared with the control (no pesticide). In contrast, the surface soil exposed to PCP at a ten-fold field rate showed different patterns from the controls at 4 weeks and 8 weeks after application. The DGGE patterns of the overlying water were much more variable than those of the surface soil in any treatments. Cluster analysis showed that the BSM plots were classified within the same group as the control at 1 week after application and that the BSM and PCP plots from 2 weeks onward after application were grouped differently from the control. Of 22 clones excised from the DGGE gels, 20 clones belonged to the Proteobacteria and two belonged to the Verrucomicrobia. It was considered that the impact of BSM on the overall microbial community (total numbers, community structure of soil) was negligible, although BSM had an impact on some specific functions of the soil microbial community (nitrification) and a part of the community (overlying water).     

Recovery of cover-crop-N in the soil-plant system in the Guinea savannah zone of Ghana

In order to understand the efficiency of residue-N use and to estimate the minimum input required to obtain a reasonable level of crop response, it is important to quantify the fate of the applied organic-N. The recovery of N from ¹⁵N-labelled Crotalaria juncea was followed in the soil and the succeeding maize crop. Apparent N recovery (ANR) by maize from unlabelled Crotalaria juncea, Crotalaria retusa, Calopogonium mucunoides, Mucuna pruriens and mineral fertilizer at three locations were also evaluated. The maize crop recovered 4.7% and 7.3% of the ¹⁵N-labelled C. juncea-N at 42 days after sowing (DAS) and at final harvest, respectively. The corresponding ¹⁵N recovery from the soil was 92.4% and 58.5%. The highest mean ANR of 57.4% was with mineral fertilizer, whereas the mean ANR of 14.3% from C. retusa was the lowest. A large pool substitution and added-N interaction effect was observed when comparing N recovery from the labelled and unlabelled C. juncea. The amount of residue-N accounted for by the isotope dilution method at 42 DAS was 97.1% and at final harvest 65.8%. The large residue-N recovery in the soil organic-N pool explains the residual effect usually observed with organic residue application.