Fish Uses CTLA-4 Immune Checkpoint to Suppress mTORC1-Controlled T-Cell Glycolysis and Immunity.

As an immune checkpoint, cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) suppresses the activation, proliferation, and effector function of T cells, thus preventing an overexuberant response and maintaining immune homeostasis. However, whether and how this immune checkpoint functions in early vertebrates remains unknown. In the current study, using a Nile tilapia (Oreochromis niloticus) model, we investigated the suppression of T cell response by CTLA-4 in bony fish. Tilapia CTLA-4 is constitutively expressed in lymphoid tissues, and its mRNA and protein expression in lymphocytes are upregulated following PHA stimulation or Edwardsiella piscicida infection. Blockade of CTLA-4 signaling enhanced T cell activation and proliferation but inhibited activation-induced T cell apoptosis, indicating that CTLA-4 negatively regulated T cell activation. In addition, blocking CTLA-4 signaling in vivo increased the differentiation potential and cytotoxicity of T cells, resulting in an enhanced T cell response during E. piscicida infection. Tilapia CTLA-4 competitively bound the B7.2/CD86 molecule with CD28, thus antagonizing the CD28-mediated costimulatory signal of T cell activation. Furthermore, inhibition of mammalian/mechanistic target of rapamycin complex 1 (mTORC1) signaling, c-Myc, or glycolysis markedly impaired the CTLA-4 blockade-enhanced T cell response, suggesting that CTLA-4 suppressed the T cell response of tilapia by inhibiting mTORC1/c-Myc axis-controlled glycolysis. Overall, the findings indicate a detailed mechanism by which CTLA-4 suppresses T cell immunity in tilapia; therefore, we propose that early vertebrates have evolved sophisticated mechanisms coupling immune checkpoints and metabolic reprogramming to avoid an overexuberant T cell response.

IL-10 Negatively Controls the Primary T Cell Response of Tilapia by Triggering the JAK1/STAT3/SOCS3 Axis That Suppresses NF-κB and MAPK/ERK Signaling.

The braking mechanisms to protect the host from tissue damage and inflammatory disease caused by an overexuberant immune response are common in many T cell subsets. However, the negative regulation of T cell responses and detailed mechanisms are not well understood in early vertebrates. In the current study, using a Nile tilapia (Oreochromis niloticus) model, we investigated the suppression of T cell immunity by IL-10. Tilapia encodes an evolutionarily conserved IL-10, whose expression in lymphocytes is markedly induced during the primary adaptive immune response against Aeromonas hydrophila infection. Activated T cells of tilapia produce IL-10, which in turn inhibits proinflammatory cytokine expression and suppresses PHA-induced T cell activation. Moreover, administration of IL-10 impairs the proliferation of tilapia T cells, reduces their potential to differentiate into Th subsets, and cripples the cytotoxic function, rendering the animals more vulnerable to pathogen attack. After binding to its receptor IL-10Ra, IL-10 activates the JAK1/STAT3 axis by phosphorylation and enhances the expression of the suppressor of cytokine signaling 3 (SOCS3), which in turn attenuates the activation of the NF-κB and MAPK/ERK signaling pathways, thus suppressing the T cell response of tilapia. Our findings elucidate a negative regulatory mechanism of T cell immunity in a fish species and support the notion that the braking mechanism of T cells executed through IL-10 existed prior to the divergence of the tetrapod lineage from teleosts. Therefore, this study, to our knowledge, provides a novel perspective on the evolution of the adaptive immune system.

TGF-ß1 suppresses the T-cell response in teleost fish by initiating Smad3- and Foxp3-mediated transcriptional networks.

Transforming growth factor-ß1 (TGF-ß1) can suppress the activation, proliferation, and function of many T-cell subsets, protecting organisms from inflammatory and autoimmune disease caused by an overexuberant immune response. However, whether and how TGF-ß1 regulates T-cell immunity in early vertebrates remain unknown. Here, using a Nile tilapia (Oreochromis niloticus) model, we investigated suppression of the T-cell response by TGF-ß1 in teleost species. Tilapia encodes an evolutionarily conserved TGF-ß1, the expression of which in lymphocytes is significantly induced during the immune response following Edwardsiella piscicida infection. Once activated, tilapia T cells increase TGF-ß1 production, which in turn suppresses proinflammatory cytokine expression and inhibits T-cell activation. Notably, we found administration of TGF-ß1 cripples the proliferation of tilapia T cells, reduces the potential capacity of Th1/2 differentiation, and impairs the cytotoxic function, rendering the fish more vulnerable to bacterial infection. Mechanistically, TGF-ß1 initiates the TGF-ßR/Smad signaling pathway and triggers the phosphorylation and nuclear translocation of Smad2/3. Smad3 subsequently interacts with several transcriptional partners to repress transcription of cytokines IL-2 and IFN-γ but promote transcription of immune checkpoint regulator CTLA4 and transcription factor Foxp3. Furthermore, TGF-ß1/Smad signaling further utilizes Foxp3 to achieve the cascade regulation of these T-cell genes. Taken together, our findings reveal a detailed mechanism by which TGF-ß1 suppresses the T cell-based immunity in Nile tilapia and support the notion that TGF-ß1 had already been employed to inhibit the T-cell response early in vertebrate evolution, thus providing novel insights into the evolution of the adaptive immune system.

Inducible IL-2 production and IL-2+ cell expansion are landmark events for T-cell activation of teleost.

As a multipotent cytokine, interleukin (IL)-2 plays important roles in activation, differentiation and survival of the lymphocytes. Although biological characteristics and function of IL-2 have been clarified in several teleost species, evidence regarding IL-2 production at the cellular and protein levels is still scarce in fish due to the lack of reliable antibody. In this study, we developed a mouse anti-Nile tilapia IL-2 monoclonal antibody (mAb), which could specifically recognize IL-2 protein and identify IL-2-producing lymphocytes of tilapia. Using this mAb, we found that CD3+ T cells, but not CD3- lymphocytes, are the main cellular source of IL-2 in tilapia. Under resting condition, both CD3+CD4-1+ T cells and CD3+CD4-1- T cells of tilapia produce IL-2. Moreover, the IL-2 protein level and the frequency of IL-2+ T cells significantly increased once T cells were activated by phytohemagglutinin (PHA) or CD3 plus CD28 mAbs in vitro. In addition, Edwardsiella piscicida infection also induces the IL-2 production and the expansion of IL-2+ T cells in the spleen lymphocytes. These findings demonstrate that IL-2 takes part in the T-cell activation and anti-bacterial adaptive immune response of tilapia, and can serve as an important marker for T-cell activation of teleost fish. Our study has enriched the knowledge regarding T-cell response in fish species, and also provide novel perspective for understanding the evolution of adaptive immune system.

Transcription factor networks drive perforin activity in the anti-bacterial immune response of tilapia.

Perforin, produced by natural killer (NK) cells and cytotoxic T lymphocytes (CTLs), is one of the effectors of cell-mediated cytotoxicity (CMC) in vertebrates, playing a paramount role in killing target cells. However, whether and how perforin is involved in adaptive immune responses in early vertebrates remains unclear. Using Nile tilapia (Oreochromis niloticus) as a model, we investigated the characteristics of perforin in early vertebrates. Oreochromis niloticus perforin (OnPRF) possesses 2 conserved functional domains, membrane attack complex/perforin (MACPF) and protein kinase C conserved region 2 (C2) domains, although they share low amino acid sequence similarity with other homologs. OnPRF was widely expressed in various immune tissues and could respond to lymphocyte activation and T-cell activation in vitro at both the transcriptional and protein levels, indicating that it may be involved in adaptive immune responses. Furthermore, after infection with Edwardsiella piscicida and Aeromonas hydrophila, the mRNA and protein levels of OnPRF were significantly up-regulated within the adaptive immune response period. Additionally, we revealed that many transcription factors were involved in the transcriptional regulation of OnPRF, including p65, c-Fos, c-Jun, STAT1 and STAT4, and there was a synergy among these transcription factors. Overall, these findings demonstrate the involvement of OnPRF in T-cell activation and adaptive immune response in tilapia, thus providing new evidence for comprehending the evolution of immune response in early vertebrates.

Comparative analysis of T-cell immunity between Streptococcus agalactiae susceptible and resistant tilapia (Oreochromis niloticus).

Nile tilapia (Oreochromis niloticus) is one of the important economic fish species cultured worldwide. However, Streptococcus agalactiae has emerged as a significant bacterial threat, severely impacting the economy of tilapia industry. The immune response underlying the resistance of tilapia to S. agalactiae are not well understood, hindering the reasonable evaluation of breeding and the formulation of effective strategies. In this study, we investigated the differences in T-cell immunity between S. agalactiae-resistant and -susceptible tilapia. Compared with susceptible tilapia, resistant tilapia exhibited a higher percentage of T cells and BrdU+ T cells during infection, indicating a superior proliferative capacity. Whether infected or not, T cells from resistant fish demonstrated a greater ability to resist apoptosis. Additionally, T cell effector genes, including interleukin (IL)-2, interferon (IFN)-γ, perforin A, and granzyme B were expressed at higher levels in resistant tilapia after infection. Along with these T-cell immune responses, resistant fish showed more effective clearance of infection. Our study elucidates the T-cell immune responses in resistant tilapia, which may contribute to the high resistance of tilapia to S. agalactiae, and provide valuable theoretical references for the selection and evaluation of disease-resistant fish strains in the future.

The TAK1/JNK axis participates in adaptive immunity by promoting lymphocyte activation in Nile tilapia.

The transforming growth factor beta-activated kinase 1 (TAK1)/c-Jun N-terminal kinase (JNK) axis is an essential MAPK upstream mediator and regulates immune signaling pathways. However, whether the TAK1/JNK axis harnesses the strength in regulation of signal transduction in early vertebrate adaptive immunity is unclear. In this study, by modeling on Nile tilapia (Oreochromis niloticus), we investigated the potential regulatory function of TAK1/JNK axis on lymphocyte-mediated adaptive immune response. Both OnTAK1 and OnJNK exhibited highly conserved sequences and structures relative to their counterparts in other vertebrates. Their mRNA was widely expressed in the immune-associated tissues, while phosphorylation levels in splenic lymphocytes were significantly enhanced on the 4th day post-infection by Edwardsiella piscicida. In addition, OnTAK1 and OnJNK were significantly up-regulated in transcriptional level after activation of lymphocytes in vitro by phorbol 12-myristate 13-acetate plus ionomycin (P + I) or PHA, accompanied by a predominant increase in phosphorylation level. More importantly, inhibition of OnTAK1 activity by specific inhibitor NG25 led to a significant decrease in the phosphorylation level of OnJNK. Furthermore, blocking the activity of OnJNK with specific inhibitor SP600125 resulted in a marked reduction in the expression of T-cell activation markers including IFN-γ, CD122, IL-2, and CD44 during PHA-induced T-cell activation. In summary, these findings indicated that the conserved TAK1/JNK axis in Nile tilapia was involved in adaptive immune responses by regulating the activation of lymphocytes. This study enriched the current knowledge of adaptive immunity in teleost and provided a new perspective for understanding the regulatory mechanism of fish immunity.

CD122 is an activation marker ensuring proper proliferation of T cells in teleost.

As one of subunits for interleukin-2 receptor (IL-2R), CD122 can bind to IL-2 and then activate downstream signal transduction to participate in adaptive immune response. Although CD122 has been identified and investigated from several teleost species, studies on its function at T-cell level are still scarce for lack of specific antibodies. In this study, a typical CD122 in Nile tilapia (Oreochromis niloticus) was characterized by bioinformatics analysis, cloned to produce retrovirus infected NIH/3T3 cells for mouse immunization. After cell fusion and screening, we successfully developed a mouse anti-tilapia CD122 monoclonal antibody (mAb), which could specifically recognize CD122 and identify CD122-producing T cells of tilapia. Using the mAb to detect, CD122 was found to widely distribute in immune-related tissues, and significantly elevate post Edwardsiella piscicida infection or T-cell activation. More importantly, the expansion of CD122+ T cells and up-regulation of CD122 occurred both in total T cells and T-cell subsets during T-cell activation upon in vitro stimulation or in vivo infection. These results indicate that CD122 can be used as a T-cell activation marker in tilapia. Notably, CD122 mAb blocking blunted the activation of MAPK/Erk and mTORC1 pathways, and inhibited T-cell proliferation, suggesting a critical role of CD122 in ensuring proper proliferation of tilapia T cells. Therefore, this study enriches the knowledge of T-cell responses in fish and provides new evidence for understanding the evolution of lymphocyte-mediated adaptive immunity.

Granzyme B secreted by T cells is involved in anti-bacterial immune response of tilapia.

Secreted by natural killer cells and cytotoxic T lymphocytes, Granzyme B is involved in regulating the adaptive immune response in vertebrates and plays a pivotal role in resisting virus invasion and removing pathogens. Although it had been extensively studied in mammals, the involvement of Granzyme B in adaptive immune response of early vertebrates remained elusive. In this study, we investigated the Granzyme B in Oreochromis niloticus (OnGrB), found that its function domain was conserved. Additionally, OnGrB was widely expressed in various tissues and could respond to T-cell activation in vitro at the transcriptional level. Furthermore, we prepared the recombinant OnGrB (rOnGrB) as an immunogen to develop a mouse anti-OnGrB monoclonal antibody (mAb). Using this anti-OnGrB mAb as a tool, we explored the expression of OnGrB in the adaptive immune response of tilapia. Our findings revealed that T cell was a significant source of OnGrB production, the expression of OnGrB at the protein level and the proportion of OnGrB + T cells increased after both T cell activation in vitro and infection with Edwardsiella piscicida in vivo. More importantly, our findings also preliminarily illuminated that p65 could regulate the transcriptional activity of OnGrB. These results indicated that OnGrB was involved in the adaptive immunity of tilapia and played a critical role in T cell function in teleost. Our study provided theoretical support and new perspectives for understanding adaptive immunity in teleost.

Excellent ratiometric two-photon fluorescent probes for hydrogen sulfide detection based on the fluorescence resonance energy transfer mechanism.

Fluorescence resonance energy transfer (FRET) is an important mechanism to design ratiometric fluorescent probes that are able to detect analytes quantitatively according to the ratio of two well-resolved emission signals. Two-photon (TP) fluorescent probes can realize the detection in living cells and tissues with deeper penetration depth, higher resolution, and lower photodamage in contrast to one-photon fluorescent probes. However, to date, fabricating TP-FRET ratiometric fluorescent probes possessing large two-photon absorption (TPA), high fluorescence quantum yield and perfect FRET efficiency is still challenging. Consequently, to develop excellent TP-FRET ratiometric probes and explore the relationship between their molecular structures and TP fluorescence properties, in this paper, we designed a series of H2S-detecting TP fluorescent probes employing the FRET mechanism based on an experimental probe BCD. Thereafter, we comprehensively evaluated the TP sensing performance of these probes by means of time-dependent density functional theory and quadratic response theory. Furthermore, we determined energy transfer efficiency and fluorescence quantum yield. Significantly, through regulating benzene-fused positions, we successfully improved fluorescence quantum yield and TPA cross-section simultaneously. Large spectral overlap between energy donor emission and acceptor absorption was achieved and near perfect energy transfer efficiency was acquired for all the studied probes. We revealed that these probes exhibit two well-resolved TPA bands, which are contributed by FRET donors and acceptors, respectively. Especially, both the wavelengths and the cross-sections of the two TPA bands agree well with those of energy donors and acceptors, which is the unique TPA spectral profile of FRET probes and has never been previously reported. Moreover, we proposed an excellent TP-FRET probe BCD3 and its product molecule BCD3-H2S, which exhibit large Stokes (141 nm and 88 nm) and emission shifts (5931 cm-1), as well as greatly increased TP action cross-sections (24-fold and 60-fold) in the near-infrared region with respect to BCD and BCD-H2S. Our detailed study can give an insight into the efficient design of novel TP-FRET fluorescent probes.

Unraveling the evolutionary patterns and phylogenomics of coronaviruses: A consensus network approach.

The COVID-19 pandemic emphasizes the significance of studying coronaviruses (CoVs). This study investigates the evolutionary patterns of 350 CoVs using four structural proteins (S, E, M, and N) and introduces a consensus methodology to construct a comprehensive phylogenomic network. Our clustering of CoVs into 4 genera is consistent with the current CoV classification. Additionally, we calculate network centrality measures to identify CoV strains with significant average weighted degree and betweenness centrality values, with a specific focus on RaTG13 in the beta genus and NGA/A116E7/2006 in the gamma genus. We compare the phylogenetics of CoVs using our distance-based approach and the character-based model with IQ-TREE. Both methods yield largely consistent outcomes, indicating the reliability of our consensus approach. However, it is worth mentioning that our consensus method achieves an approximate 5000-fold increase in speed compared to IQ-TREE when analyzing the data set of 350 CoVs. This improved efficiency enhances the feasibility of conducting large-scale phylogenomic studies on CoVs.

Rational design of HJH antimicrobial peptides to improve antimicrobial activity.

Antimicrobial peptides (AMPs) have attracted considerable interest in the past decade due to their advantages for tackling antibiotic resistance. They exhibit potential antimicrobial activity through unique cell membrane destruction mechanism based on their net charge, hydrophobic properties, and α-helix. In this work, a series of HJH peptides was rationally designed and synthesized. The antimicrobial activity and cytotoxicity assays indicated that HJH-5 and HJH-6 containing hydrophobic residues and helices displayed prominent antimicrobial activity and mild cytotoxicity, respectively. These peptides may be developed for combatting microbial infections.

Interleukin-12 induces IFN-γ secretion and STAT signaling implying its potential regulation of Th1 cell response in Nile tilapia.

As a pleiotropic cytokine consisting of IL-12p35 and IL-12p40, Interleukin-12 (IL-12) features in inflammation regulation and anti-bacterial immunity. While IL-12 homologs have been identified in non-mammalian species, the precise mechanisms by which IL-12 contributes to early adaptive immune responses in vertebrates remain incompletely understood. Herein, an evolutionary conserved Oreochromis niloticus IL-12 (defined as OnIL-12) was identified by synteny characterization, structural comparisons and phylogenetic pattern of IL-12p35b and IL-12p40a. IL-12p35b and IL-12p40a exhibited widespread expression in lymphoid-related tissues of tilapia, while their mRNA expression in head-kidney demonstrated a significant increase after Edwardsiella piscicida infection. Compared with other lymphocytes, recombinant OnIL-12 (rOnIL-12) displayed stronger affinity binding to T cells. Although stimulation of lymphocytes with the p35b or p40a subunit resulted in a significant induction of IFN-γ expression, rOnIL-12 showed stronger potential to promote IFN-γ expression than these subunits. rOnIL-12 not only elevated the mRNA expression level Th1 cell-associated transcription factor T-bet in lymphocytes, but also increased the proportion of CD4-1+IFN-γ+ lymphocytes. Moreover, the mRNA and phosphorylation levels of STAT1, STAT3, STAT4 and STAT5 were enhanced by rOnIL-12. These findings will offer previous evidence for further exploration into the regulatory mechanisms of Th1 cellular immunity in early vertebrates.

Exploring site-specific N application rate to reduce N footprint and increase crop production for green manure-rice rotation system in southern China.

Milk vetch (Astragalus sinicus L.) is leguminous green manure (GM) which produces organic nitrogen (N) for subsequent crops and is widely planted and utilized to simultaneously reduce the use of synthetic N fertilizer and its environmental costs in rice systems. Determination of an optimal N application rate specific to the GM-rice system is challenging because of the large temporal and spatial variations in soil, climate, and field management conditions. To solve this problem, we developed a framework to explore the site-specific N application rate for the low-N footprint rice production system in southern China based on multi-site field experiments, farmer field survey, and process-based model (WHCNS_Rice, soil water heat carbon nitrogen simulator for rice). The results showed that a process-based model can explain >83.3% (p < 0.01) of the variation in rice yield, aboveground biomass, crop N uptake, and soil mineral N. Based on the scenario analysis of the tested WHCNS_Rice model, the simple regression equation was developed to implement site-specific N application rates that considered variations in GM biomass, soil, and climatic conditions. Simulation evaluation on nine provinces in southern China showed that the site-specific N application rate reduced regional synthetic N fertilizer input by 29.6 ± 17.8% and 65.3 ± 23.0% for single and early rice, respectively; decreased their total N footprints (NFs) by 23.4% and 49.3%, respectively; and without reduction in rice yield, compared with traditional farming N practices. The reduction in total NF was attributed to the reduced emissions from ammonia volatilization by 35.2%, N leaching by 28.4%, and N runoff by 32.7%. In this study, we suggested a low NF rice production system that can be obtained by combining GM with site-specific N application rate in southern China.

Sortase A-mediated cyclization of novel polycyclic RGD peptides for ανß3 integrin targeting.

Cyclic arginine-glycine-aspartic (RGD) peptides that specifically bind to integrin ανß3 have been developed for drug delivery, tracers, and imaging for tumor diagnosis and treatment. Herein, a series of polycyclic RGD peptides containing dual, tri, and tetra rings were designed and synthesized through sortase A-mediated ligation. An in vitro test on cell adhesion inhibition indicated that the RGD peptide containing tricylic structure exhibited outstanding potency and selectivity for ανß3 integrin.

Activated T cells are the cellular source of IL-22 that enhances proliferation and survival of lymphocytes in Nile tilapia.

As a pleiotropic cytokine mainly secreted by CD4+ T cells, interleukin (IL)-22 plays an important role in immune regulation and infection elimination. Despite IL-22 homologues have been identified in non-mammal, whether and how IL-22 participates in the adaptive immune response of early vertebrates have not been fully addressed. In this study, we identified an evolutionarily conserved IL-22 from Nile tilapia Oreochromis niloticus (defined as OnIL-22), proved by its properties regarding sequence, gene structure, functional domain, tertiary structure and phylogeny. IL-22 was broadly expressed in lymphoid-related tissues of tilapia, and with relatively higher levels in skin, gill, intestine and liver. The expression of OnIL-22 in spleen lymphocytes was markedly induced at the adaptive immune stage after Streptococcus agalactiae infection. Moreover, once lymphocytes were activated by PMA plus ionomycin or T-cell specific mitogen PHA in vitro, OnIL-22 expression was obviously up-regulated at both mRNA and protein levels. These results thus suggest that activated T cells produce IL-22 to take part in the adaptive immune response of tilapia. Furthermore, treatment of lymphocytes with recombinant OnIL-22 increased the expression of genes related to proliferation and survival, and further promoted the proliferation and reduced the apoptosis of lymphocytes during bacterial infection or T-cell activation. These cellular effects of IL-22 seem to be associated with JAK1/STAT3 axis downstream of IL-22, because IL-22 application not only elevated the mRNA expression of JAK1 and STAT3, but also enhanced their phosphorylation in lymphocytes. Altogether, we suggest that activated T cells produce IL-22 to promote lymphocyte proliferation and survival probability via JAK1/STAT3 signaling pathway, thus participating in adaptive immune response of Nile tilapia. Our study therefore provides helpful perspective for understanding the function and mechanism of adaptive immune system in teleost.

Evolutionary history and functional characterization of Lj-TICAM-a and Lj-TICAM-b formed via lineage-specific tandem duplication in lamprey (Lampetra japonica).

Toll/interleukin-1 receptor domain-containing adaptor molecule (TICAM) genes respond to infections. We identified TICAM-a and TICAM-b in Lampetra japonica and investigated their evolutionary history and potential function via comparative genomics and molecular evolution analyses. They are arranged in tandem and evolved from a multi-exon to a single-exon structure. Lj-TICAM-a and Lj-TICAM-b might be the ancestral gene of the vertebrate TICAM genes. Lj-TICAM-b arose via a lamprey-specific tandem duplication event. Both genes are expressed in many tissues during an immune response, and exhibit different responses to peptidoglycan, indicating their functional divergence. Simultaneous overexpression of both proteins activated nuclear factor κB expression and co-immunoprecipitation assays indicated that they might form a complex for signal transduction. However, unlike in mammals, the TICAM-dependent signaling pathway in lamprey might rely on TRAF3 rather than on TRAF6. These results suggest that both Lj-TICAM-a and Lj-TICAM-b play a role in host defenses.

MSClustering: A Cytoscape Tool for Multi-Level Clustering of Biological Networks.

MSClustering is an efficient software package for visualizing and analyzing complex networks in Cytoscape. Based on the distance matrix of a network that it takes as input, MSClustering automatically displays the minimum span clustering (MSC) of the network at various characteristic levels. To produce a view of the overall network structure, the app then organizes the multi-level results into an MSC tree. Here, we demonstrate the package's phylogenetic applications in studying the evolutionary relationships of complex systems, including 63 beta coronaviruses and 197 GPCRs. The validity of MSClustering for large systems has been verified by its clustering of 3481 enzymes. Through an experimental comparison, we show that MSClustering outperforms five different state-of-the-art methods in the efficiency and reliability of their clustering.

Density functional theory calculations and thermodynamic analysis of bridgmanite surface structure.

Bridgmanite, a high temperature and pressure form of MgSiO3, is believed to be Earth's most abundant mineral and responsible for the observed seismic anisotropy in the mantle. Little is known about surfaces of bridgmanite but knowledge of the most stable surface terminations is important for understanding various geochemical processes as well as likely slip planes. A density functional theory based thermodynamic approach is used here to establish the range of stability of bridgmanite as well as possible termination structures of the (001), (010), (100) and (011) surfaces as a function of the chemical potential of oxygen and magnesium. The vibrational contribution to the Gibbs free energy is found to be essential for obtaining a stability region of bridgmanite in the phase diagram. The most stable surface termination of bridgmanite varies between three different atomic structures depending on the chemical potential of oxygen and magnesium. The results presented provide a basis for further theoretical studies of the chemical processes on bridgmanite surfaces in the Earth's mantle and slip plane analysis.

Effects of gas reservoir configuration and pore radius on shale gas nanoflow: A molecular dynamics study.

We calculated methane transport through cylindrical graphite nanopores in cyclical steady-state flows using non-equilibrium molecular dynamics simulations. First, two typical gas reservoir configurations were evaluated: open (OS) and closed (CS) systems in which pores connect to the gas reservoir without/with a graphite wall parallel to the gas flow. We found that the OS configuration, which is commonly used to study nanoflows, exhibited obvious size effects. Smaller gas reservoir cross-sectional areas were associated with faster gas flows. Because Knudsen diffusion and slip flow in pores are interrupted in a gas reservoir that does not have walls as constraints, OSs cannot be relied upon in cyclical nanoflow simulations. Although CSs eliminated size effects, they introduced surface roughness effects that stem from the junction surface between the gas reservoir and the pore. To obtain a convergent nanoflow, the length of a side of the gas reservoir cross-section should be at least 2 nm larger than the pore diameter. Second, we obtained methane flux data for various pore radii (0.5-2.5 nm) in CSs and found that they could be described accurately using the Javadpour formula. This is the first direct molecular simulation evidence to validate this formula. Finally, the radial density and flow-velocity distributions of methane in CS pores were analyzed in detail. We tested pores with a radius between 0.5 nm and 2.5 nm and determined that the maximum ratio (∼34%) of slip flow to overall flow occurred in the pore with a radius of 1.25 nm. This study will aid in the design of gas reservoir configurations for nanoflow simulations and is helpful in understanding shale gas nanoflows.