The Advancement and Application of the Single-Cell Transcriptome in Biological and Medical Research.

Single-cell RNA sequencing technology (scRNA-seq) has been steadily developing since its inception in 2009. Unlike bulk RNA-seq, scRNA-seq identifies the heterogeneity of tissue cells and reveals gene expression changes in individual cells at the microscopic level. Here, we review the development of scRNA-seq, which has gone through iterations of reverse transcription, in vitro transcription, smart-seq, drop-seq, 10 × Genomics, and spatial single-cell transcriptome technologies. The technology of 10 × Genomics has been widely applied in medicine and biology, producing rich research results. Furthermore, this review presents a summary of the analytical process for single-cell transcriptome data and its integration with other omics analyses, including genomes, epigenomes, proteomes, and metabolomics. The single-cell transcriptome has a wide range of applications in biology and medicine. This review analyzes the applications of scRNA-seq in cancer, stem cell research, developmental biology, microbiology, and other fields. In essence, scRNA-seq provides a means of elucidating gene expression patterns in single cells, thereby offering a valuable tool for scientific research. Nevertheless, the current single-cell transcriptome technology is still imperfect, and this review identifies its shortcomings and anticipates future developments. The objective of this review is to facilitate a deeper comprehension of scRNA-seq technology and its applications in biological and medical research, as well as to identify avenues for its future development in alignment with practical needs.

Age-related compositional and functional changes in the adult and breastfed buffalo rumen microbiome.

Introduction: The buffalo is an important domestic animal globally, providing milk, meat, and labor to more than 2 billion people in 67 countries. The rumen microorganisms of buffaloes play an indispensable role in enabling the healthy functionality and digestive function of buffalo organisms. Currently, there is a lack of clarity regarding the differences in the composition and function of rumen microorganisms among buffaloes at different growth stages. Methods: In this study, metagenomics sequencing technology was applied to examine the compositional and functional differences of rumen microorganisms in adult and breastfed buffaloes. Results: The results revealed that the rumen of adult buffaloes had significantly higher levels of the following dominant genera: Prevotella, UBA1711, RF16, Saccharofermentans, F23-D06, UBA1777, RUG472, and Methanobrevibacter_A. Interestingly, the dominant genera specific to the rumen of adult buffaloes showed a significant positive correlation (correlation>0.5, p-value<0.05) with both lignocellulose degradation-related carbohydrate-active enzymes (CAZymes) and immune signaling pathways activated by antigenic stimulation. The rumen of breastfed buffaloes had significantly higher levels of the following dominant genera: UBA629, CAG- 791, Selenomonas_C, Treponema_D, Succinivibrio, and RC9. Simultaneously, the rumen-dominant genera specific to breastfed buffaloes were significantly positively correlated (correlation>0.5, p-value<0.05) with CAZymes associated with lactose degradation, amino acid synthesis pathways, and antibiotic-producing pathways. Discussion: This indicates that rumen microorganisms in adult buffaloes are more engaged in lignocellulose degradation, whereas rumen microorganisms in breastfed buffaloes are more involved in lactose and amino acid degradation, as well as antibiotic production. In conclusion, these findings suggest a close relationship between differences in rumen microbes and the survival needs of buffaloes at different growth stages.

Multi-omics reveals the mechanism of rumen microbiome and its metabolome together with host metabolome participating in the regulation of milk production traits in dairy buffaloes.

Recently, it has been discovered that certain dairy buffaloes can produce higher milk yield and milk fat yield under the same feeding management conditions, which is a potential new trait. It is unknown to what extent, the rumen microbiome and its metabolites, as well as the host metabolism, contribute to milk yield and milk fat yield. Therefore, we will analyze the rumen microbiome and host-level potential regulatory mechanisms on milk yield and milk fat yield through rumen metagenomics, rumen metabolomics, and serum metabolomics experiments. Microbial metagenomics analysis revealed a significantly higher abundance of several species in the rumen of high-yield dairy buffaloes, which mainly belonged to genera, such as Prevotella, Butyrivibrio, Barnesiella, Lachnospiraceae, Ruminococcus, and Bacteroides. These species contribute to the degradation of diets and improve functions related to fatty acid biosynthesis and lipid metabolism. Furthermore, the rumen of high-yield dairy buffaloes exhibited a lower abundance of methanogenic bacteria and functions, which may produce less methane. Rumen metabolome analysis showed that high-yield dairy buffaloes had significantly higher concentrations of metabolites, including lipids, carbohydrates, and organic acids, as well as volatile fatty acids (VFAs), such as acetic acid and butyric acid. Meanwhile, several Prevotella, Butyrivibrio, Barnesiella, and Bacteroides species were significantly positively correlated with these metabolites. Serum metabolome analysis showed that high-yield dairy buffaloes had significantly higher concentrations of metabolites, mainly lipids and organic acids. Meanwhile, several Prevotella, Bacteroides, Barnesiella, Ruminococcus, and Butyrivibrio species were significantly positively correlated with these metabolites. The combined analysis showed that several species were present, including Prevotella.sp.CAG1031, Prevotella.sp.HUN102, Prevotella.sp.KHD1, Prevotella.phocaeensis, Butyrivibrio.sp.AE3009, Barnesiella.sp.An22, Bacteroides.sp.CAG927, and Bacteroidales.bacterium.52-46, which may play a crucial role in rumen and host lipid metabolism, contributing to milk yield and milk fat yield. The "omics-explainability" analysis revealed that the rumen microbial composition, functions, metabolites, and serum metabolites contributed 34.04, 47.13, 39.09, and 50.14%, respectively, to milk yield and milk fat yield. These findings demonstrate how the rumen microbiota and host jointly affect milk production traits in dairy buffaloes. This information is essential for developing targeted feeding management strategies to improve the quality and yield of buffalo milk.

Multi-omics reveal the effects and regulatory mechanism of dietary neutral detergent fiber supplementation on carcass characteristics, amino acid profiles, and meat quality of finishing pigs.

This study aimed to reveal the effects and regulatory mechanism of dietary NDF on the performance of pigs by multi-omics analysis. Results showed that 16 % dietary NDF significantly improved meat quality, increased flavor amino acid content, and reduced backfat thickness and the feed-to-gain ratio. 16S rDNA sequencing showed that 16 % NDF significantly increased the abundance of Akkermansia, Lachnoclostridium, and Ruminococcus. Transcript analysis showed that genes related to muscle development and lipid metabolism were significantly modified. Metabonomic analysis showed that 16 % NDF significantly increased amino and fatty acid related metabolites. Correlation analysis suggested that 16 % NDF treatment may alter the gut microbiota and metabolites, regulate the expression of genes related to lipid and amino metabolism, and ultimately affect the flavor and performance of pigs. This study provides a novel understanding about the effect and regulatory mechanism of NDF supplements on the finishing pigs and a relevant reference for the improvement of diet formulation.

The multi-kingdom microbiome of the goat gastrointestinal tract.

BACKGROUND: Goat is an important livestock worldwide, which plays an indispensable role in human life by providing meat, milk, fiber, and pelts. Despite recent significant advances in microbiome studies, a comprehensive survey on the goat microbiomes covering gastrointestinal tract (GIT) sites, developmental stages, feeding styles, and geographical factors is still unavailable. Here, we surveyed its multi-kingdom microbial communities using 497 samples from ten sites along the goat GIT. RESULTS: We reconstructed a goat multi-kingdom microbiome catalog (GMMC) including 4004 bacterial, 71 archaeal, and 7204 viral genomes and annotated over 4,817,256 non-redundant protein-coding genes. We revealed patterns of feeding-driven microbial community dynamics along the goat GIT sites which were likely associated with gastrointestinal food digestion and absorption capabilities and disease risks, and identified an abundance of large intestine-enriched genera involved in plant fiber digestion. We quantified the effects of various factors affecting the distribution and abundance of methane-producing microbes including the GIT site, age, feeding style, and geography, and identified 68 virulent viruses targeting the methane producers via a comprehensive virus-bacterium/archaea interaction network. CONCLUSIONS: Together, our GMMC catalog provides functional insights of the goat GIT microbiota through microbiome-host interactions and paves the way to microbial interventions for better goat and eco-environmental qualities. Video Abstract.

NiS gradient distribution on arrayed porous carbonized grapefruit peel for water splitting.

Metal-based catalysts on biomass carbon substrates can combine their respective advantages of composition and structure to improve the catalytic performance. Herein, NiS supported on grapefruit peel derived array porous carbon (APC) was obtained via a carbonization process without emission of toxic gases. The natural S source from grapefruit peel reacted with nickel salt solution. The gradient distribution of NiS and S on the APC substrates can be altered by the concentration of impregnating salt solution. Theoretical calculations showed that the S gradient distribution on APC could tune the electronic structure and optimize the adsorption energies of the intermediates. NiS was firmly anchored on the porous carbon framework, resulting in enhanced high intrinsic activity, exposure of more active sites, and accelerated mass transfer. The active mass density was proposed to build a relationship between active metal content and electrolyte diffusion capacity for the evaluation of catalytic properties.

ReS2 Cocatalyst Improves the Hydrogen Production Performance of the CdS/ZnS Photocatalyst.

Photocatalysis provides an exciting solution to the current growing energy challenge. However, the activity and stability of photocatalysts are two important issues in photocatalytic applications. In this work, we have successfully developed an efficient and stable photocatalyst by loading ReS2 nanoparticles onto a CdS/ZnS heterojunction. After loading ReS2, there is a strong interaction between the CdS/ZnS heterojunction and ReS2, which accelerates the photogenerated charge migration and effectively inhibits the recombination of photogenerated electrons and holes. Accordingly, CdS/ZnS-ReS2 displays excellent photocatalytic activity and stability with the highest hydrogen production rate of 10 722 µmol g-1 h-1, which is approximately 178 times higher than that of the pure CdS and 5 times better than that of CdS/ZnS. This work not only facilitates solar energy conversion to improve photocatalytic activity and stability but also broadens the application of ReS2 as a cocatalyst.

Regulation Effectiveness and Mechanism of Biotransformation Pathway on the Toxicity of Microcystin-LR Target to Protein Phosphatase 2A.

Biotransformation is recognized as a potential pathway to regulate the environmental risk of microcystins (MCs). To explore the regulation effectiveness and mechanism of the biotransformation pathway, six typical MCLR-biotransformation products (MCLR-BTPs) were prepared, and their inhibition effects on protein phosphatase 2A (PP2A) were evaluated. The inhibition effects of the MCLR-BTPs generally decreased with the increase in biothiol molecular weights and polarity, indicating that biotransformation was an effective pathway through which to regulate MCLR toxicity. To further explore the regulation mechanism, the key interaction processes between the MCLR/MCLR-BTPs and the PP2A were explored by homology modeling and molecular docking. The introduced biothiols blocked the covalent binding of Mdha7 to Cys269 but strengthened the hydrogen bond "Mdha7"→Arg268. The changed "Mdha7" intervened the combination of MCLR-BTPs to PP2A by weakening the hydrogen bonds Arg4←Arg214, Arg4→Pro213, Adda5←His118, and Ala1←Arg268, and the ionic bond Glu6-Mn12+. The weakening combination of the MCLR-BTPs to PP2A further attenuated the interactions between the conserved domain and the Mn2+ ions (including the ionic bonds Asp57-Mn12+ and Asp85-Mn12+ and the metal bonds Asp57-Mn12+ and Asn117-Mn12+) and increased the exposure of the Mn2+ ions. Meanwhile, the weakened hydrogen bond Arg4←Arg214 facilitated the combination of the phosphate group to Arg214 (with increased exposure). In this way, the catalytic activity of the PP2A was restored.

Mechanism for the Potential Inhibition Effect of Microcystin-LR Disinfectant By-Products on Protein Phosphatase 2A.

The secondary contamination of microcystin disinfection by-products (MC-DBPs) is of concern due to the residual structure similar to their original toxin. Based on identification and preparation, the potential inhibition effect of typical MCLR-DBPs (associated with the oxidation of Adda5) on PP2A was confirmed in the sequence of MCLR > P1 > P4 > P3 ≈ P2 > P7 ≈ P6 ≈ P5 > P8. To elucidate the molecular mechanism underlying the inhibition effect, the interaction models for typical MCLR-DBPs and PP2A were constructed using a modeling-based-on-ligand-similarity approach, and the candidate interaction parameters between typical MCLR-DBPs and PP2A were obtained by molecular docking. By analyzing the correlation between inhibition data and candidate interaction parameters, the key interaction parameters were filtered as hydrogen bonds "Adda5"←Asn117, "Adda5"←His118, MeAsp3←Arg89, Arg4←Arg214, Arg4→Pro213; ionic bonds Glu6-Arg89, Asp85-Mn12+, Asp57-Mn22+; and metal bonds Glu6-Mn12+, Glu6-Mn22+. With the gradual intensification of chlorination, Adda5 was destroyed to varying degrees. The key interactions changed correspondingly, resulting in the discrepant inhibition effects of typical MCLR-DBPs on PP2A.

Rapid surface reconstruction strategies for oxygen evolution reactions: chemical grafting of MXene quantum dots on Ni-Co layered double hydroxides.

The surface reconstruction of transition metal-based catalysts with their specific catalytic mechanism is currently one of the hotspots and difficulties in the electrocatalytic oxygen evolution reaction (OER). Herein, a chemical grafting strategy was proposed to facilitate the surface reconstruction of Ni-Co layered double hydroxide@MXene quantum dot (Ni-Co LDH@MQDs) electrocatalysts to optimize the OER kinetics. The surface reconstruction of Ni-Co LDH@MQDs was predicted and monitored by a combination of ab initio molecular dynamics, density functional theory and experimental verification. Compared with weak electrostatic bonds, the rapid surface evolution of electrocatalysts can be revealed due to the strong chemical grafting between the MQDs and LDHs. The reconstituted Ni-Co LDH@MQD electrocatalysts undergo an unconventional bifunctional mechanism to lower the barriers of the rate-limiting step of the OER. This work provides a research strategy for transition metal catalysts for efficient catalysis by designing surface reconfiguration.

A pipeline to evaluate the discrepant interactions between typical nitrogenous disinfection byproduct haloacetonitriles and human hemoglobin.

To evaluate the interaction between haloacetonitriles (HANs) and human hemoglobin (Hb), a pipeline was established based on fluorescence spectra, mass spectra and molecular docking. Fluorescence spectra analysis showed the fluorescence of Hb was statically quenched by HANs in the sequence of TCAN > DBAN > DCAN > IAN > BAN > CAN. HANs could combine to multiple surface sites of Hb accounting for "hydrogen bonds" and "van der Waals forces". The high-resolution mass spectra analysis for Hb with and without HANs further confirmed the formation of multiple HAN-Hb complexes with different conversion rates. With the assistance of MOE molecule docking, the potential combination sites and related interactions parameters between HANs and Hb were filtrated. By analyzing the correlations between the candidate interactions parameters and fluorescence quenching constants/MS conversion rates, the combination sites of HANs were fixed at Asp126 (α1/α2), Lys127 (α1/α2) in the form of "hydrogen bonds" X â†’ Asp126 (α1/α2), N â†’ Lys127 (α1/α2). In this way, the potential interactions between HANs and Hb were effectively evaluated.

Insight into the Molecular Mechanism for the Discrepant Inhibition of Microcystins (MCLR, LA, LF, LW, LY) on Protein Phosphatase 2A.

Microcystins (MCs) exhibit diversified inhibition effects on protein phosphatases (PPs) due to their structural differences. To fully evaluate the potential mechanism for the discrepant inhibition effects, the five most frequent MCs with varying residues at position Z4 were selected as the tested toxins. Their inhibition sequence on PP2A was detected as follows: MCLR > MCLW > MCLA > MCLF > MCLY. Combined with homology modeling and molecular docking technology, the major interaction parameters between the MCs and PP2A were obtained. The correlation analysis for the major interaction parameters and inhibition effects showed that the hydrophobicity of Z4 had an important influence on the interaction of the MCs to PP2A. The introduction of hydrophobic Z4 directly weakened hydrogen bonds Z4→Pro213 and Z4←Arg214, indirectly weakened hydrogen bonds Adda5←Asn117, Glu6←Arg89, and MeAsp3←Arg89, but indirectly enhanced ionic bonds Glu6←Arg89, Glu6-Mn12+, and Glu6-Mn22+. In this way, the combination of the MCs with PP2A was blocked, and thus, the interactions between PP2A and the Mn2+ ions (in the catalytic center) were further affected; metal bonds Asp85-Mn12+ and Asp85-Mn22+ were weakened, while metal bond His241-Mn12+ was enhanced. As a result, the interactions in the catalytic center were inhibited to varying degrees, resulting in the reduced toxicity of MCs.

pH Oscillator-Driven Jellyfish-like Hydrogel Actuator with Dissipative Synergy between Deformation and Fluorescence Color Change.

Development of soft actuators with complex practical functions is significant for imitating the behaviors of living organisms. However, it is still a challenge to fabricate artificial soft actuators with jellyfish-like synergistic deformation and fluorescence color change (SDFC) and autonomous dynamic behavior, but such a system could obviously endow the classic soft actuators with more functions. Herein, we proposed to utilize tetra(4-pyridylphenyl)ethylene (TPE-4N) luminogen with pH-responsive aggregation-induced emission (AIE) to fabricate the AIE active hydrogel, which could be further employed to obtain an anisotropic bilayer soft actuator based on strong interfacial adhesion with acrylic acid (AA) gels. Furthermore, artificial flower-shape actuators showing SDFC behaviors were demonstrated. On the basis of these findings, jellyfish-inspired autonomous gel actuators driven by a pH oscillator have been fabricated, in which periodical SDFC behaviors completely regulated by the system itself without repetitive on/off switches of external stimuli were well synchronized with the pH oscillator. The described combination of nonlinear chemistry and responsive hydrogels actuator opens pathways toward out-of-equilibrium SDFC devices with autonomous behavior useful for biomimetic fields.

The microbiome of the buffalo digestive tract.

Buffalo is an important livestock species. Here, we present a comprehensive metagenomic survey of the microbial communities along the buffalo digestive tract. We analysed 695 samples covering eight different sites in three compartments (four-chambered stomach, intestine, and rectum). We mapped ~85% of the raw sequence reads to 4,960 strain-level metagenome-assembled genomes (MAGs) and 3,255 species-level MAGs, 90% of which appear to correspond to new species. In addition, we annotated over 5.8 million nonredundant proteins from the MAGs. In comparison with the rumen microbiome of cattle, the buffalo microbiota seems to present greater potential for fibre degradation and less potential for methane production. Our catalogue of microbial genomes and the encoded proteins provides insights into microbial functions and interactions at distinct sites along the buffalo digestive tract.

Molecularly Engineered Covalent Organic Frameworks for Hydrogen Peroxide Photosynthesis.

Synthesizing H2 O2 from water and air via a photocatalytic approach is ideal for efficient production of this chemical at small-scale. However, the poor activity and selectivity of the 2 e- water oxidation reaction (WOR) greatly restricts the efficiency of photocatalytic H2 O2 production. Herein we prepare a bipyridine-based covalent organic framework photocatalyst (denoted as COF-TfpBpy) for H2 O2 production from water and air. The solar-to-chemical conversion (SCC) efficiency at 298 K and 333 K is 0.57 % and 1.08 %, respectively, which are higher than the current reported highest value. The resulting H2 O2 solution is capable of degrading pollutants. A mechanistic study revealed that the excellent photocatalytic activity of COF-TfpBpy is due to the protonation of bipyridine monomer, which promotes the rate-determining reaction (2 e- WOR) and then enhances Yeager-type oxygen adsorption to accelerate 2 e- one-step oxygen reduction. This work demonstrates, for the first time, the COF-catalyzed photosynthesis of H2 O2 from water and air; and paves the way for wastewater treatment using photocatalytic H2 O2 solution.

Cloning and expression of three heat shock protein genes in the gills of Cherax quadricarinatus responding to bacterial challenge.

Cherax quadricarinatus is seriously affected by multiple types of pathogens, including bacteria and viruses, and has been widely transplanted around the world. Heat shock proteins (Hsps) are a group of molecular chaperones that play important roles in promoting the proper refolding and blocking the aggregation of denatured proteins. In this study, CqHsp60, CqHsp70 and CqHsp90 from C. quadricarinatus were cloned, and their expression patterns were analysed. The CDS (coding sequence) lengths of the CqHsp60, CqHsp70 and CqHsp90 genes were 1731 bp, 1932 bp and 2199 bp, encoding 576, 643 and 732 amino acids, respectively. CqHsp60 was 99.13%, 98.78% and 88.63% identical to the corresponding sequences of Cherax cainii, Cherax destructor and Eriocheir sinensis, respectively. CqHsp70 showed 99.84%, 92.73% and 91.58% identity to the corresponding sequences of C. cainii, C. destructor and E. sinensis, while CqHsp90 was 98.25%, 98.51% and 91.41% identical with those of C. cainii, C. destructor and E. sinensis, respectively. The expression patterns of the three CqHsps were different between males and females. CqHsp60 and CqHsp70 exhibited the highest expression in the hepatopancreas of males and the gonads of females, and CqHsp90 presented the highest expression in the gonads of males and hepatopancreas of females. After pathogenic inoculation, the death trend of C. quadricarinatus at different time points was the same in association with different pathogens, with most deaths occurring within 6 h post-inoculation. The trend of CqHsp transcription at different time points was the same among the groups treated with Vibrio alginolyticus, Vibrio parahemolyticus and Aeromonas hydrophila, exhibiting upregulation first and then downregulation. The expression of CqHsp60 and CqHsp70 in the gills of living C. quadricarinatus was less than 3.5 times that in the PBS group, but in the gills of dead C. quadricarinatus under A. hydrophila inoculation, its expression was more than 5-9 times that in the PBS group. CqHsp90 expression changed dramatically in the V. alginolyticus, V. parahemolyticus and A. hydrophila groups, in which it exceeded 50 times the level in the PBS group. These results indicated that CqHsps could induce the activation of the immune system within a short time and that CqHsp90 could be used as a more effective molecular biomarker than CqHsp70 and CqHsp60 in a pathogenic bacterium-polluted environment.

Heavy metal accumulation, geochemical fractions, and loadings in two agricultural watersheds with distinct climate conditions.

The main aim of this study was to explore the effects of climate conditions on the transport and transformation of heavy metals. Sedimentary geochemical analysis and watershed modeling were used to investigate the distinctions between heavy metal pollution under different climate conditions. The results showed that the average concentrations of Cu, Cd, and Pb in sediments of the subtropical watershed (36.64, 0.60, and 133.69 mg/kg, respectively) were higher than those of the temperate watershed (26.58, 0.19, and 23.17 mg/kg, respectively) because of surface runoff-induced heavy metal loadings under higher precipitation. Also, the labile fractions, which mainly originated from anthropogenic sources, showed higher percentages in the subtropical watershed (67.84-91.33%), thereby indicating that the transport of heavy metals was promoted by surface runoff. Moreover, higher percentages of acid-soluble fractions of Cu and Pb (23.55-33.60%) in the subtropical watershed suggested that higher temperatures accelerated the transformation of heavy metal fractions, thus contributing to the transportation of heavy metals. Overall, climate conditions were the dominant factors for the differences between the subtropical and temperate watersheds. The results of this study suggest that the effects of climate conditions on the transport, enrichment, and bioavailability of heavy metals are of great significance. Such effects should therefore be the focus of future studies.

Watershed diffuse pollution dynamics and response to land development assessment with riverine sediments.

Sediment cores can reflect diffuse pollution history due to the accumulation of pollutants over time, therefore, the quantitative relationship between the sedimentation flux of pollutants and diffuse loads can identify the historical change. Sediment cores were collected from two river reaches in a small agricultural watershed (143 km2), and the total nitrogen (TN) and total phosphorus (TP) concentrations were determined. The sediments were dated using 210Pb isotope radioactivity and the TN and TP sedimentation flux was calculated with Constant Rate of Supply (CRS) and Constant Initial Concentration (CIC) models. Watershed loss loads were simulated using the Soil and Water Assessment Tool (SWAT) in the same temporal period. As the similar natural condition in the post-depositional period of sediments, a linear regression model was used to analyze the relationship between TN and TP sedimentation flux and the hindcast model data. The TP sedimentation flux showed a clear positive relationship with its simulated load (R2 = 0.600 and 0.664) using the CRS model, and better reflected long-term diffuse pollution loss dynamics than nitrogen. The impacts of land use change on diffuse pollution loading were identified with the combination of sedimentation flux from different reaches and watershed modeling. During the expansion of paddy land in the lower reach, the difference of TP sedimentation flux between upper and lower reaches narrowed, while gap of TN sedimentation flux increased. Base on the lateral correlations of two sections, the sediment concentration of TP was more reliable for the long term diffuse pollution assessment under land development.

Using river sediments to analyze the driving force difference for non-point source pollution dynamics between two scales of watersheds.

The formation and transportation processes of non-point source (NPS) pollution varied among the studied watersheds in the Northeastern China, so we hypothesized that the driving force behind NPS pollution followed the spatial scale effect. With a watershed outlet sedimentary flux analysis and a distributed NPS pollution loading model, we investigated the temporal dynamics of NPS and the differences in driving forces. Sediment core samples were collected from two adjacent watersheds, the smaller Abujiao watershed and the larger Naoli watershed. The natural climatic conditions, long-term variations in the distribution of land use, soil properties and tillage practices were the same in the two watersheds. The vertical distributions of total nitrogen, total phosphorus, Zn and As at 1-cm intervals in the section showed clear differences between the watersheds. There were higher concentrations of total nitrogen and total phosphorus in the larger watershed, but the heavy metals were more concentrated in the smaller watershed. Lead-210 (210Pb) analyses and the constant rate of supply model provided a dated sedimentary flux, which was correlated with the corresponding yearly loading of NPS total nitrogen and total phosphorus in the two watersheds. The total phosphorus showed a stable relationship in both watersheds with an R2 value that ranged from 0.503 to 0.682. A rose figure comparison also demonstrated that the pollutant flux in the sediment was very different in the two watersheds, which had similar territorial conditions and different hydrological patterns. Redundancy analysis further indicated that expanding paddy areas had a large impact on the sedimentary flux of nitrogen and phosphorus in the smaller watershed, but precipitation had a direct impact on NPS loading in the larger watershed. We concluded that the spatial scale effect affected the NPS pollution via the transport processes in the waterway, which was mainly influenced by branch length and drainage density.