Arabidopsis in combination with the chemical probes developed through chemical genetics to understand SL perception of parasitic plants. This work first shows that understanding SL signaling is useful in developing chemical probes that perturb SL perception. Second, it indicates that the chemical space available to probe SL signaling in both model and parasitic plants is sizeable. Because these parasitic pests represent a major concern for food insecurity in the developing world, there is great need for chemical approaches to uncover novel lead compounds that perturb parasitic plant infections.Strigolactones (SLs) are a collection of related small molecules that act as hormones in plant growth and development. Intriguingly, SLs also act as ecological communicators between plants and mycorrhizal fungi and between host plants and a collection of parasitic plant species. In the case of mycorrhizal fungi, SLs exude into the soil from host roots to attract fungal hyphae for a beneficial interaction. In the case of parasitic plants, however, root-exuded SLs cause dormant parasitic plant seeds to germinate, thereby allowing the resulting seedling to infect the host and withdraw nutrients. Because a laboratory-friendly model does not exist for parasitic plants, researchers are currently using information gleaned from model plants like With respect to plants, around ten small molecule hormones have been identified so far, and core signaling pathways for each of these hormones have been characterized3. Much of the success in understanding how small molecule hormones are perceived in plants comes from genetic analysis, which usually involves finding mutants with altered hormone sensitivity followed by molecular identification of the wild-type protein involved. The power of genetics in dissecting plant hormone signaling was impressive, especially since many players in plant hormone signaling were genetically redundant, which precluded their identification as simple recessive mutations6. Many saturated genetic screens, however, led to the identification of rare dominant mutations9 which served as toeholds in building many signaling pathways. The subsequent development of well-characterized mutant knockout collections inArabidopsis10 allowed more components to be validated through the construction of multiple loss-of-function mutant lines in redundant steps.Small organic molecules are important sources of signaling hormones in both plants and animals11. Simply defined, chemical genetics involves the development of chemical agonists and antagonists to probe biological processes. This approach by definition should be well suited for plant hormone signaling since plant hormones are small molecules and as such their receptors should be “druggable”12. However, geneticists often look suspiciously on chemical perturbation experiments because of concerns of off-target effects. Chemical geneticists have tried to address this criticism by defining criteria for what makes a good chemical probe14  to a mutant lacking the 5-AT target (histidine3 [his3]) resulted in dramatic reductions in the global gene expression signature observed in 5-AT-treated wild-type yeast cells16.The contribution of genetics to unravelling plant hormone biology is self-evident, which raises interest in where genetic analysis can make future contributions. A good example of the evolution of genetic approaches in plant hormone signaling is now occurring in the field of chemical geneticsrobe14 . From a 17. Pyrabactin was first identified as a general germination inhibitor in a chemical screen inArabidopsis, but its specific role in ABA signaling was suggested by the ability of ABA-insensitive mutants to germinate on pyrabactin. Second, the pyrabactin analog, apyrabactin, showed no biological activity. Finally, although a decoder approach was not applied, global gene expression experiments between seeds treated with ABA or pyrabactin were highly correlated, suggesting few off-target effects.In plants, perhaps the best example of chemical probe development involved the identification of pyrabactin as a selective agonist of the receptor for the hormone abscisic acid (ABA)17. Because pyrabactin was a selective ABA agonist, it activated only a subset of ABA receptors and particularly the major one involved in germination17. Thus, resistant mutants to pyrabactin circumvent genetic redundancy issues that cannot be resolved by traditional ABA screens. This result showed how a new approach like chemical genetics, which can identify more selective compounds, can uncover novelty when wedded to an old approach like a traditional forward genetics screen. The pyrabactin story also demonstrated how the identification of a specific chemical probe can have broader applications beyond basic biology. Information on the pyrabactin structure has led to the identification of chemical analogs that could be used to protect important agronomic crops from drought, a process that is mediated by ABA signaling19.The identification of pyrabactin allowed the development of genetic screens to identify mutations in an essential gene encoding an ABA receptor that is involved in germination21. SLs, like all small molecule plant hormones, have many roles in plant growth ranging from filament growth in nonvascular plants22 to shoot branching23 and root development24 in vascular plants. However, unlike most plant hormones, SLs also have ecological signaling roles. SLs are exuded from plant roots into the rhizosphere, where they attract arbuscular mycorrhizal (AM) fungi for a beneficial interaction that brings water and nutrients to the plant25. Unfortunately, root-exuded SLs also act as a cue to tell a number of parasitic plant species that a host is nearby26. In these cases, obligate parasites of theStriga,Phelipanche, andOrobanche genera have evolved strong seed dormancy, which is broken only when they sense host-derived SLs. After germination, the parasite infects the host with devastating consequences. The range of infestations by theStriga genera alone includes much of sub-Saharan Africa, and these infestations are considered to be a major impediment to poverty alleviation on the continent28. Thus, the identification of chemicals that probe SL perception not only will help to understand SL biology but also may lead to the development of new compounds to combat parasitic plant infestations in the developing world.This development of chemicals as both probes for plant hormone signaling and translational leads is clearly exciting. Perhaps there is not a more obvious application of chemicals than in the study of a recently identified collection of chemically related hormones called strigolactones (SLs)30. The ABC rings do show natural chemical variation, and to date approximately 20 SLs have now been identified30. SLs can be further categorized based on stereochemistry around the B and C ring into the strigol and orobanchol families  connected to a tricyclic lactone (ABC rings) via an enol-ether bridge 29,30. amilies . Finally35. Avenaol, for example, which is found in root exudates fromAvena strigosa (black oat) and shows SL activity, has the C and D moieties but lacks a B ring and has an additional carbon between the A and C rings , which form a lid encompassing the SL ligand-binding pocket . This event appears to result in small conformational changes that allow the recruitment of other SL signaling partners. The recruitment of these signaling partners in turn appears to cause a larger conformational change of the receptor to a closed state by trapping either a D-ring intermediate or the D ring itself43. Although many details on timing and order need to be worked out, the covalent bond between the D-ring moiety and D14-type receptors suggests that the metabolism of the SL ligand may be integral to perception and downstream SL signaling44. This also explains why D14-type receptors retain a canonical α/β hydrolase amino acid catalytic triad that shows very slow kinetics of hydrolysis44.The enol-ether linkage between the C and D rings also seems to be important. SL receptors in model plants, collectively called D14-type SL receptors after the rice DWARF14 (D14) receptor pocket 38–42. merging . As SL e45. The double-barreled name of HTL/KAI2 is because of two groups independently identifying loss-of-function mutations in this gene based on two phenotypes: 1) hyposensitivity to light (HTL)46 and 2) insensitivity to the smoke-derived germination stimulant karrikin (KAI2)47. The identification of HTL/KAI2 hydrolases as SL receptors in parasitic plants was surprising, as these proteins in model systems do not respond well to naturally occurring C2'-(R) SL isomers and at this time their natural ligand is unknown48. Analysis ofHTL/KAI2 genes fromStriga hermonthica (ShHTL/KAI2) revealed that these receptors have a range of sensitivity and specificity with respect to the SLs they recognize50. Functional analysis inArabidopsis demonstrated that one receptor,ShHTL7, was sufficient to increase the sensitivity of the SL response inArabidopsis to picomolar levels, which is within the concentration range observed for the response ofStriga hermonthica seed to SLs50. Recent biochemical analysis of ShHTL7 suggested that it may have a similar mode of action to D14-type receptors with respect to transducing an SL signal51. Although a mechanistic understanding of how parasitic plant receptors attain high levels of SL sensitivity has not been clearly elucidated, it does appear that parasitic plantHTL/KAI2 genes, unlike their nonparasitic plant counterparts, have evolved away from perceiving karrikins to sensing SLs50.So what about parasitic plant SL signaling? Unlike model plants, D14-type receptors do not appear to be the major player in the germination response of parasites to host-derived SLs. This function falls to a related D14 α/β hydrolase given the name HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE2 (HTL/KAI2)53. The logic involves synthesizing cheap SL analogs that are stable in the soil which would germinate parasitic seed in the absence of a host. These compounds are generically called “suicide germination compounds” because these obligate parasites likeStriga hermonthica will die after germination without a host. The first synthesized suicide germination compounds such as the popular derivative GR24  that showed good SL activity although they lacked the ABC ring structures and any enol-ether linkages , a number of these compounds have subsequently been shown to bind and activate AtHTL/KAI257. The phthalimide structure in some cotylimides (CTL-IV) is also found in Nijmegen-1, a potent SL mimic58. However, unlike Nijmegen-1, CTL compounds do not contain a D ring and certainly are not a hydrolysable substrate. Interestingly, other compounds built on phthalimide lactone scaffolds but lacking enol-ether linkages have good selectivity activity on variousOrobanche andPhilibanche species59. Finally, a screen for novel germination agonists developed specifically around the activation of the AtHTL/KAI2 receptor with one of its protein partners, MORE AXILLARY GROWTH2 (MAX2), identified more structurally unrelated compounds that have activity inStriga hermonthica germination assays   fromStriga hermonthica now exists50, and it would be interesting to see howin silico screening of this receptor may influence lead compound identification.Although chemical genetic screens for new SL agonists are viewed through the prism of SL chemistry, screening for SL antagonists has not had this bias. One of the first attempts to identify SL antagonists involved taking advantage of D14-type receptor structural biology. Compounds were first screened (2-MN) , and subArabidopsis rather than knowledge of the receptor structurea priori. This assay was based on the observation that SLs inhibitedArabidopsis hypocotyl elongation. Another advantage to usingArabidopsis was a genotype whose hypocotyl was sensitized to SL inhibition, which was used in the primary screen to identify compounds that interfered with SL-dependent inhibition of hypocotyl growth61. Seven compounds were found, one of which was a piperidine-based molecule called soporidine (SOP). In addition to binding AtHTL/KAI2, SOP bound theStriga hermonthica receptor ShHTL7 and inhibited its hydrolytic activity. Functionally, SOP inhibited the germination ofStriga hermonthica seed in the presence of SL61. Thus, hypocotyl-based assays inArabidopsis appear to be a good screening platform to identify lead compounds that may perturbStriga germination. None of the antagonists identified so far have significant structural similarity to known naturally occurring SLs. Similar to the case of SL agonists, it is possible that these antagonists bind somewhere else on the receptor to inhibit activity. This again begs for more studies on the structure–function relationships between the SL receptors and the plethora of compounds identified from chemical screens.Recently, a second SL antagonist screen has been performed using whole plant SL-dependent assays inThe rapid increase in our knowledge of the fundamental biology of SL signaling based on studies on model plants is allowing researchers to test the mechanism by which SLs are perceived in parasitic plants. Furthermore, this information is currently being used to identify new chemical probes that perturb SL perception in both nonparasitic and parasitic plants. Although it is not surprising that small molecules resembling SLs have activity in SL-based plant assays, the identification of a large number of synthetic lead compounds from chemical screens that do not possess canonical SL structures is intriguing. This could simply reflect different binding sites and modes of action. It could also mean that SL receptors, whether from parasitic or nonparasitic plants, are more promiscuous than previously thought with respect to small molecule activation, particularly at higher concentrations.in vivo. This explanation, however, has never really explained functional low-affinity ligandsin vivo and many times these compounds are written off as nonspecific and biologically irrelevant. In some sense, this hand-waving argument is akin to early beliefs about “sticky proteins” found in large-scale protein–protein interaction networks. Once thought to be biochemical artifacts, most of these proteins are now viewed as essential components in scale-free signaling networks63. It is now becoming clear from evolutionary studies on small molecule hormone receptors from animals that many small molecule receptors were initially low-affinity sensors for a range of metabolites and later evolved to become high-affinity receptors of particular chemicals66. These models are consistent with suggestions that the ancient HTL/KAI2 receptors may have had reduced chemical specificity that later evolved into high-specificity SL receptors67. Such a model may explain why HTL/KAI2-type receptors were selected over D14-type receptors by parasitic plants, since these species must be able to readily evolve to new host SL ligands and compositions in order to move to new hosts68.Biologists are usually taught that compounds that bind a receptor with high affinity are most likely the most relevant69. Once compounds have been identified, the methods of medicinal chemistry can be used to increase their potency orders of magnitude. In this scenario, lead compounds identified with SL activity would serve as chemical scaffolds for the development of more potent compounds. Applying the same logic, perhaps the addition of D-ring structures to leads implicated in SL function, would increase their efficacy.The lack of high-affinity agonists and antagonists could suggest chemical genetics will not yield useful probes and interesting insights. However, although traditionally searches for drugs have been based on finding high-affinity compounds, the pharmaceutical industry has changed its strategy to perform “fragment-based screening”, which is designed to find low-molecular-weight compounds that work in the high micromolar range initially2, which are well defined developmentally and easy to handle experimentally, were instrumental in dissecting how small molecule hormones are perceived and signal. By contrast, plant hormone signaling was led by phenotypic screening typically on whole organisms such asArabidopsis70. Chemical genetic analysis of hormone signaling in these two kingdoms will most likely follow the same route, and animal researchers are now more frequently using phenotypic screening as an effective method of drug discovery71. In plant systems, compounds will continue to be identified through some phenotypic screen and the resulting compounds will be the basis for mutational analysis to further understand the mode of action as was seen with pyrabactin17. The grounding of plant chemical genetics in phenotypic screening bodes well for new insights, since phenotypic screening is gathering momentum as a way to re-energize animal drug research72. In this sense, plant biologists are already there.Finally, insights into plant versus animal hormone signaling have come from different approaches. Animal cell culture systems


In conclusion, liver lipid accumulation, fatty acids (FA) composition and inflammation were modulated by the dietary composition of lipids and carbohydrates. The HCD group had more potent lipogenic and inflammatory effects in comparison with HFD.Both high-carbohydrate diet (HCD) and high-fat diet (HFD) modulate liver fat accumulation and inflammation, however, there is a lack of data on the potential contribution of carbohydrates and lipids separately. For this reason, the changes in liver fatty acid (FA) composition in male Swiss mice fed with HCD or HFD were compared, at the time points 0 (before starting the diets), and after 7, 14, 28 or 56 days. Activities of stearoyl-CoA desaturase-1 (SCD-1), ∆-6 desaturase (D6D), elongases and de novo lipogenesis (DNL) were estimated. Liver mRNA expression of acetyl-CoA carboxylase 1 (ACC1) was evaluated as an additional indicator of the de novo lipogenesis. Myeloperoxidase activity, nitric oxide (NO) production, and mRNA expressions of F4/80, type I collagen, interleukin (IL)-6, IL-1β, IL-10, and tumor necrosis factor-α (TNF-α) were measured as indication of the liver inflammatory state. The HCD group had more intense lipid deposition, particularly of saturated fatty acids (SFAs) and monounsaturated fatty acids (MUFAs). This group also showed higher DNL, SCD-1, and D6D activities associated with increased NO concentration, as well as myeloperoxidase activity. Livers from the HFD group showed higher elongase activity, stored more polyunsaturated fatty acids (PUFAs) and had a lower omega-6/omega-3 fatty acid ( Non-alcoholic fatty liver disease (NAFLD) is a condition whereby there is high hepatic lipid accumulation even under low alcohol intake . NAFLD iNAFLD is the most common cause of liver diseases since this organ has a limited capacity for lipid storage. It is associated with obesity, insulin resistance, type 2 diabetes, hypertriglyceridemia, plasma non-high density lipoprotein cholesterol (HDL) high levels and cardiovascular disease ,6,7. TheIn spite the fact that liver fat accumulation and liver inflammation are the main characteristics of NAFLD , the conWe compared herein the changes in the liver FA composition in mice fed with HCD or HFD for 7, 14, 28 or 56 days as compared to the values before starting the diets (day 0). Activities of stearoyl-CoA desaturase-1 (SCD-1), ∆-6 desaturase (D6D), elongases and de novo lipogenesis (DNL) were estimated and gene expression of acetyl-CoA carboxylase 1 was evaluated. Nitric oxide content, myeloperoxidase activity and inflammatory gene expressions were measured as indication of the liver inflammatory state that is closely associated with fat deposition ,10 and iMus musculus), weighing about 35 g (six weeks of age), were obtained from the State University of Maringá Breeding Center and used in the experiments. Swiss mice are resistant to development of diet-induced obesity [Male Swiss mice  and had free access to water and food.The experimental protocol used (protocol 002/2014) was approved by the Animal Ethics Committee of The State University of Maringá (CEUA).®, Curitiba, PR, Brazil) before the initiation of the experimental protocol.Mice were fed with standard rodent chow  of protein, carbohydrate and total fat in the HCD were 14.2, 73.8 and 4, respectively. Quantities (g/100 g) of protein, carbohydrate and total fat in the HFD were 20.3, 36.5 and 35.2, respectively.The diets were prepared with highly refined ingredients purchased from Rhoster Company  and lard was used as a major source of fat. The HCD composition was based on purified diets for maintenance of laboratory adult rodents proposed by the American Institute of Nutrition (AIN-93-M) . This lan = 5) or HFD (n = 6) for 56 days.Mice  were fed with a HFD or HCD for 0, 7, 14, 28 or 56 days. After receiving HCD or HFD, mice fasted from 17:00 to 08:00 h, were killed by decapitation (to avoid any interference of anesthesia) and the livers removed and stored in liquid nitrogen. The inflammatory state of the liver was evaluated only in animals fed with HCD (n-6/n-3) ratio, i.e., 15/1.FA composition of the diets was measured . The maiTotal lipid contents of diets and livers were extracted using the method of Bligh and Dyer , in redu−1 carrier gas (hydrogen), 30 mL∙min−1 make-up gas (nitrogen), 350 mL∙min−1 synthetic air and 35 mL∙min−1 hydrogen flame gas. The injected sample volume was 2.0 µL with split injection ratio 1:80. The injector and detector temperatures were 200 °C and 240 °C, respectively. The column temperature was maintained at 165 °C for 7 min, followed by a heating rate of 4 °C per min until reaching 185 °C, which was maintained for 4.67 min. After that, a new heating rate of 6 °C per min was applied until reaching 235 °C, which was maintained for 5 min, totalling 30 min of analysis.The homogenized sample was weighed (1.000 ± 0.001 g) in 10 mL glass tubes and extraction performed by adding 2 mL methanol, 2 mL chloroform and 1 mL distilled water in different steps. The tubes were vortexed for 9 min and, after extraction, centrifuged at 3000 rpm for 5 min. The residual chloroform phase, containing total lipid extracted, was separated and the solvent removed using nitrogen gas flow. Fatty acid methyl esters (FAME) of diet and liver homogenates were prepared by ultrasound assisted total lipid methylation as described by Santos et al. . FAME seRetention times and peak areas were determined using the Chrom-Quest™ software (Thermo Scientific™). For identification of FA, retention times were compared to those of standard methyl esters. FA contents in the diets and livers were expressed as mg/100 mg sample. n-7/16:0 and 18:1n-9/18:0; D6D as the ratio of 18:3n-6/18:2n-6; elongase as the ratio of 18:0/16:0; and DNL as the ratio of 16:0/18:2n-6.Enzyme activities and DNL were estimated as the product/precursor ratio of individual FA as follows: SCD-1 activity as the ratios of 16:1N-(1-naphtyl) ethylenediamine in milli-Q water (0.1 g), was added at room temperature. Absorbance was measured at 550 nm using an ELISA plate reader. NO production was calculated using a standard curve of sodium nitrite and results expressed as μM.NO gas released from cells into the media reacts with water to produce nitrite and nitrate . NitriteThe livers were homogenized in phosphate buffered saline (PBS) and the homogenate was stirred in a vortex and centrifuged (2500 rpm) for 5 min.o-dianisidine dihydrochloride (4.2 mg), double-distilled water (22.5 mL), potassium phosphate buffer , and H2O2  was also added. The enzyme reaction was stopped by addition of 30 μL sodium acetate (2.23 g in 20 mL of double-distilled water).The supernatant (10 μL) was added to each well in triplicate. PBS (0.2 mL), containing Myeloperoxidase activity was determined at 460 nm using a microplate spectrophotometer .Total RNA was extracted from 50 mg of liver using TRIzol reagent  and reverse transcribed to cDNA . Gene expression was evaluated by real-time PCR using SYBR Green as fluorescent dye (Invitrogen Life Technologies). The primer sequences were: F4/80, NM_010130.4, sense CCTGAACATGCAACCTGCCAC, antisense GGGCAT GAGCAGBCTGTAGGATC, Type I collagen, NM_009931.2, sense CTCTATGTCCAAGGCAACGAG, antisense TCACAAACCGCACACCTG, IL-6, NM_001314054.1, sense, antisense, IL-1β, NM_008361.4, sense GGCAGCTACCTGTGTCTTTCCC, antisense ATATGGGTCCGACAGCACGAG, TNF-α, NM_001278601.1, sense TCTTCTCATTCCTGCTTGTGGC, antisense CACTTGGTGGTTTGCTACGAC G, IL-10, NM_010548.2, sense TGCCAAGCCTTATCGGAAATG, antisense AAATCGATGACAGCGCCTCAG, ACC1 NM_133360.2, sense GAGAGGGGTCAAGTCCTTCC, antisense AAAACATCCACTTCCACACACGA. The analysis of gene expression was carried out using a previously described method  with theThe IMI was calculated by the sum of expressions of pro inflammatory factors divided by the sum of expressions of anti-inflammatory factors as follows: interleukin (IL)-6 + IL-1β + tumor necrosis factor-α (TNF-α) + F4/80 + type 1 collagen/IL-10.t-test using Graph-Pad Prism Version 5.0 software  were used to assess differences between means. p-values < 0.05 indicate statistical significance.Results are reported as means ± standard deviation of the means and were analyzed by one-way ANOVA. Tukey and Student In agreement with the diet composition , livers p < 0.05) during the experimental period for both HCD and HFD. The HCD group had high (p < 0.05) FA accumulation, particularly with respect to SFA and MUFA (p < 0.05) levels of PUFAs (n-3 and n-6), due to more intense reduction of linoleic acid, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) levels during the 56-day period  content was increased during the 56-day period in the livers of both groups but the changes were more pronounced in HFD mice . Decreasy period . The PUFy period .(The MUFA/SFA ratio was increased in livers from the HCD group from day 7 .p < 0.05) of SCD-1 and D6D activities during the 56-day period were observed in the livers of HCD mice (t-test for p < 0.05.Elevations (HCD mice . Liver Ap < 0.05) and decreased (p < 0.05) in the liver of the HFD and the HCD groups, respectively, during the experimental period. DNL was increased throughout the study either by HCD or HFD but the changes were more pronounced in HCD mice.The elongase activities were increased (p < 0.05) after 56 days of receiving HCD as compared to the HFD group  indicating a more intense inflammatory state in this group.However, the IMI was increased in the liver of HCD animals (In agreement with previous studies in mice  and humap < 0.05) deposition of FA in the HCD group, which was inferred from the sum of all FA, was due to MUFAs being the main contributors  expressions, and stimulation of FA oxidation [The mechanisms by which DNL increases due to high carbohydrate diet involve SREBP-1c and ChREBP, which influence the expression of key genes involved in DNL such as acetyl-CoA carboxylase. Acetyl-CoA, generated from glucose, activates the transcription factors SREBP1c and ChREBP in the liver, which stimulate DNL . In contxidation .p < 0.05) hepatic levels of linoleic acid and α-linolenic acid and their products of elongation and desaturation were found in HFD mice  levels of EPA and DHA  elongase activity, also had higher stearic acid levels (p < 0.05) in the HCD group, in spite of the fact that the HCD group had lower elongase activity  D6D and SCD-1 activity  levels of n-3 PUFAs (p < 0.05) n-6/n-3 fatty acid ratio .p < 0.05) inflammatory state in HCD mice (p < 0.05) activity of myeloperoxidase (Increased myeloperoxidase activity and NO contents in the liver indicate a more prominent (HCD mice . Myeloperoxidase  in the lp < 0.05) NO production (p < 0.05) in the liver from the HFD group (p < 0.05) in the livers of the HCD group (Pro-inflammatory cytokines such as IL-1 β, TNF-α, IFN-γ  and nucloduction  in HCD aFD group , decreasCD group , enhanceThe F4/80 is one of the most specific cell-surface markers for macrophages. F4/80 is highly and constitutively expressed in resident tissue macrophages, including Kupffer’s cells in the liver . There wn-6/n-3 fatty acid ratio, indicates an increased inflammatory state in livers from mice fed with HCD for 56 days, as compared with the HFD group. In accordance with these results, high postprandial blood glucose levels induced by high carbohydrate food intake increase NO generation, that in turn can combine with superoxide to produce peroxynitrite, a potent long-lived pro-oxidant molecule, contributing either to acute or chronic low-grade inflammation [There were no differences in liver cytokine mRNA expression between the HCD and HFD groups. However, the IMI was increased in livers from the HCD group mainly because IL-10 was poorly expressed in this group . This inammation .The results in livers from mice fed with HCD or HFD are summarized .n-3 and n-6 PUFAs. Increased lipid deposition particularly of SFAs and MUFAs, higher SCD-1 and D6D activities and ACC1 expression and DNL were reported in the liver from the HCD group. These changes were associated with a more intense liver inflammation state (The proportion of fat and carbohydrates in the diet modulated the deposition of lipids and composition of liver fatty acids. The liver from the HFD group had higher elongase activity and stored more on state .


In addition, the G4 ligand PhenDC3 prevents NCL binding on EBNA1 mRNA and reverses GAr-mediated repression of translation and antigen presentation. This shows that the NCL-EBNA1 mRNA interaction is a relevant therapeutic target to unveil EBV-carrying cancers to the immune system and that the yeast model can be successfully used for uncovering drugs and host factors that interfere with EBV stealthiness.The oncogenic Epstein-Barr virus (EBV) evades the immune system but has an Achilles heel: its genome maintenance protein EBNA1. Indeed, EBNA1 is essential for viral genome replication and maintenance but also highly antigenic. Hence, EBV evolved a system in which the glycine-alanine repeat (GAr) of EBNA1 limits the translation of its own mRNA at a minimal level to ensure its essential function thereby, at the same time, minimizing immune recognition. Defining intervention points where to interfere with EBNA1 immune evasion is an important step to trigger an immune response against EBV-carrying cancers. Thanks to a yeast-based assay that recapitulates all the aspects of EBNA1 self-limitation of expression, a recent study by Lista  Hence, in order to avoid the immune system to detect and kill EBV-infected cells, the virus has evolved a mechanism to limit EBNA1 production to the minimal level to sustain viral genome replication and maintenance and, at the same time, to minimize the production of EBNA1-derived antigenic peptides. The central glycine-alanine repeat (GAr) of EBNA1 plays a critical role in this immune evasion strategy as it inhibits the translation of its own mRNA in cis, thereby limiting the production of EBNA1-derived antigenic peptides . This inhibitory effect of GAr is length-dependent as a longer domain displays a stronger inhibitory effect on both mRNA translation and antigen presentation. Of note, a polymorphism in the length of GAr exists, which may reflect a way for EBV to adapt to its host. GAr-based EBNA1 immune evasion has been considered a relevant therapeutic target to treat EBV-related cancers, hence there is a need to develop cell-based assays that allow high throughput screenings. As the mechanism of GAr-mediated mRNA translation suppression in cis was not known, nor the cellular factors involved, a yeast (Saccharomyces cerevisiae) assay that recapitulates all the aspects of the GAr-based inhibition of translation, including the GAr-length dependency, has been developed. Briefly, this assay is based on the construction of yeast strains that express individually fusions between GAr of various lengths and the Ade2p reporter protein as sole source of Ade2p enzyme. Yeast cells which express Ade2p at a functional level form white colonies, whereas cells that do not express Ade2p readily form red colonies and cells that express any intermediate level of Ade2p form pink colonies whose coloration intensity is inversely proportional to the level of Ade2p expressed. Because GAr is also able to self-limit in a length-dependent manner the translation of its own mRNA in yeast, this leads to a reduction in Ade2p level which is proportional to GAr size and can be easily monitored by looking at the color of the yeast colonies. A yeast strain that endogenously expressed a fusion between a 43 amino acids GAr and Ade2p was chosen because it leads to a partial inhibition of Ade2p expression, thus leading to pink colonies expressing an intermediate level of Ade2p. Hence, this 43GAr-ADE2 strain allows to look for modifiers (drugs or cellular factors) that either suppress or exacerbate the inhibitory effect of GAr on translation . This 43GAr-ADE2 strain was successfully used to identify small molecular weight compounds that can stimulate 43GAr-Ade2p expression in yeast, hence leading to whiter colonies that express higher level of 43GAr-Ade2  whereas having no effect on the control strain expressing Ade2p alone . This yeast model for EBNA1 stealthiness was validated since these drugs, which include doxorubicin, were later shown to also stimulate EBNA1 expression in EBV-infected human cells and to relieve GAr-based limitation of antigen presentation. These results also suggested that potential host cell genes involved in EBV immune evasion were conserved in yeast.Yeast-based chemobiological models for a number of human diseases involving functionally conserved physiopathological factors have been quite largely and successfully developed for about fifteen years. However, yeast assays based on the expression of key physiopathological players that do not exist in yeast are also possible (e.g.: yeast models for Huntington and Parkinson diseases). Here we briefly describe how we developed a yeast model to identify drugs and host cell factors that interfere with the capacity of the Epstein-Barr virus (EBV) to evade the host immune system. EBV is the first oncogenic virus discovered in human and has been linked to various cancers that include Burkitt and Hodgkin lymphomas, the nasopharyngeal carcinoma and 10% of gastric cancers. EBV evades the host immune system but, fortunately, has an Achilles heel: its genome maintenance protein EBNA1 which is, on the one hand, essential for EBV genome replication and maintenance and as such expressed in all dividing EBV-infected cells. On the other hand, EBNA1 is highly antigenic and CD8et al. (2017) took advantage of the same yeast assay and of a multicopy plasmid-based yeast genomic library to identify these host cell genes involved in the GAr-mediated inhibition of translation. This way, the yeast NSR1 gene, which encodes the orthologue of human nucleolin (NCL), was isolated. Indeed, when overexpressed in the pink 43GAr-ADE2 strain, Nsr1p led to the formation of redder yeast colonies that express even less 43GAr-Ade2p, whereas the knockout of the NSR1 gene (which is viable in yeast) in the same strain led to white colonies that express high level of 43GAr-Ade2p, showing that Nsr1p is involved in GAr-based limitation of protein expression . The effect of NSR1 is GAr-dependent since its overexpression or its inactivation had no effect on Ade2p expression in the control strain . Importantly the human NCL was able to complement the 43GAr-ADE2 nsr1Δ strain that then turned back to its initial pink color and expressed lower levels of 43GAr-Ade2p. Therefore, based on these yeast results, the role of NCL in EBNA1 immune evasion was deciphered in EBV-infected human cells, leading to the formal demonstration that human NCL is involved in GAr-based limitation of translation and antigen presentation.Hence, Lista NCL was known to interact with G-quadruplexes (G4) which are non-canonical secondary structures of guanine-rich nucleic acids formed by the stacking of guanine-quartets which correspond to a planar arrangement of four guanines connected by hydrogen bonds. In the study highlighted here, it was shown that NCL binds directly to the G4s formed in the GAr-encoding sequence of EBNA1 mRNA and that this interaction is crucial for EBNA1 self-limitation of translation and antigen presentation. Importantly, this interaction is druggable since the G4 ligand PhenDC3 prevents NCL from binding to G4 formed in the GAr mRNA sequence of EBNA1 mRNA and stimulates GAr-limited translation and antigen presentation . Hence, NCL, the first host cell factor involved in EBNA1 immune evasion, has been uncovered thanks to the yeast model for EBNA1 stealthiness. In addition, this led to the discovery of a pertinent and druggable therapeutic target to trigger an immune response against EBV-related cancers: the interaction between NCL and the G4s of GAr-encoding sequence of the EBNA1 mRNA. Therefore, yeast-based chemo¬biological models for diseases involving key physiopathological actors that do not exist in yeast are definitively worth a trial as, should they work, they potentially allow to easily setup drug and genetic screenings aiming at uncovering both candidate drugs and new therapeutic targets. Of note, doxorubicin, which has been isolated thanks to the yeast assay, has recently been shown to interact with G4, hence potentially filling the gap with nucleolin.


This data presented herein are the research summary of “mechanical behavior and durability performance of concrete containing recycled concrete aggregate”   Specifications TableValue of the data•The influence of replacement of recycle concrete aggregate in production of new structural concrete is discussed here•This data set can also be used as a guideline for others to scrutinize the properties of recycle concrete aggregate•The research data presented herein may be useful to manufacture different commercial elements using recycle concrete aggregate•This data set may also encourage to recycle other wastes to minimize the dependency on natural resources1This dataset reported herein were obtained from the experimental studies conducted on the different percentages of recycle concrete aggregate (RCA) replacement in the concrete mix and relate them to the influence of physical, mechanical and durability properties. The detailed of the dataset can also be found in 2The materials compositions for different RCA mixes used in the research are shown in The RCA was collected from the different sources of construction and demolition waste (C&DW) sites in the Western Cape region of South Africa. Concrete was then made by replacing different percentages  of NA by RCA as they are assigned RCA0 to RCA100 in The physical properties of aggregates such as relative density of aggregates, aggregate crushing value, flakiness index, and water absorption capacity were also measured and presented in


Klebsiella pneumoniae associated with β-lactamase and efflux pump activities.Bacteria treated with different classes of antibiotics exhibit changes in susceptibility to successive antibiotic treatments. This study was designed to evaluate the influence of sequential antibiotic treatments on the development of antibiotic resistance in K. pneumoniae grown at 37 °C by adding initial (0 h) and second antibiotics (8 or 12 h). Treatments include control , no initial antibiotic addition followed by 1 MIC ciprofloxacin addition (CON-CIP), no initial antibiotic addition followed by 1 MIC meropenem addition (CON-MER), initial 1/4 MIC ciprofloxacin addition followed by no antibiotic addition (1/4CIP-CON), initial 1/4 MIC ciprofloxacin addition followed by 1 MIC ciprofloxacin addition (1/4CIP-CIP), and initial 1/4 MIC ciprofloxacin addition followed by 1 MIC meropenem addition (1/4CIP-MER).The antibiotic susceptibility, β-lactamase activity, and efflux activity were determined in K. pneumoniae throughout the incubation period. The ciprofloxacin treatments (CON-CIP and 1/4CIP-CIP) showed significant reduction in the number of K. pneumoniae cells compared to meropenem (CON-MER and 1/4CIP-MER). The 1/4CIP-CIP achieved a further 1 log reduction of K. pneumoniae, when compared to the 1/4CIP-CON and 1/CIP-MER. The increase in sensitivity of K. pneumoniae to cefotaxime, kanamycin, levofloxacin, nalidixic acid was observed for CON-CIP. Noticeable cross-resistance pattern was observed at the 1/4CIP-CIP, showing the increased resistance of K. pneumoniae to chloramphenicol, ciprofloxacin, kanamycin, levofloxacin, nalidixic acid norfloxacin, sulphamethoxazole/trimethoprim, and tetracycline. The levels of β-lactamase activities were estimated to be 8.4 μmol/min/ml for CON, 7.7 μmol/min/ml for 1/4CIP-CON and as low as 2.9 μmol/min/ml for CON-CIP. Compared to the absence of phenylalanine-arginine-β-naphthylamide (PAβN), the fluorescence intensity of EtBr was increased in K. pneumoniae cells treated at the CON, CON-CIP, and CON-MER in the presence of PAβN. However, the efflux pump activity remained in K. pneumoniae cells treated at the 1/CIP, 1/CIP–CIP, and 1/CIP-MER in the presence of PAβN.Compared to the CON, the initial addition of 1/4 MIC ciprofloxacin inhibited the growth of The results suggest that the pre-exposed antibiotic history, treatment order, and concentrations influenced the development of multiple antibiotic resistant associated with β-lactamase and efflux pump activities. This study highlights the importance of antibiotic treatment conditions, which would be taken into consideration when new antibiotic strategy is designed to prevent antibiotic resistance. Klebsiella pneumoniae, an opportunistic pathogen, is mainly responsible for nosocomial infections with high morbidity and mortality [K. pneumoniae has significantly increased the risk of developing serious nosocomial and community-acquired infections worldwide [K. pneumoniae strains can cause treatment failure with current antibiotic therapy [The overuse, underuse, and misuse of antibiotics have become major causes of the development of antibiotic resistance in bacteria . The emeortality . Recentlorldwide , 4. Furt therapy .K. pneumoniae against different classes of antibiotics include release of antibiotic-inactivating enzymes, modification of antibiotic target sites, change in membrane permeability, activation of efflux pump systems, and alteration of metabolic pathways [K. pneumoniae [K. pneumoniae strains produce enzymes, including extended-spectrum β-lactamases, metallo-β-lactamases, oxacillinases, and carbapenemases, that can degrade β-lactam antibiotics. The efflux pumps, belonging to the resistance-nodulation-division (RND) family, can extrude amphiphilic and charged antibiotics such as β-lactams, fluoroquinolones, and aminoglycosides [The proposed resistance mechanisms of pathways , 5. Amoneumoniae . K. pneuycosides .K. pneumoniae in association with β-lactamase and efflux pump activities.The sequential and combination antibiotic therapies have currently been used to reduce not only the evolution of multidrug resistant bacteria but also the levels of antibiotics used in the treatment of bacterial infection –9. The dK. pneumoniae ATCC 23357 was purchased from American Type Culture Collection . The strain was cultured in trypticase soy broth  at 37 °C for 18 h. After cultivation, the culture was centrifuged at 3000×g for 20 min at 4 °C. The harvested cells were washed twice with phosphate-buffered saline  and diluted to 108 CFU/ml for assays.Strain of K. pneumoniae to ciprofloxacin and meropenem was evaluated according to the Clinical Laboratory Standards Institute (CLSI) procedure with slight modification [5 CFU/ml and incubated at 37 °C for 18 h. Minimum inhibitory concentrations (MICs) of ciprofloxacin and meropenem were defined as the lowest concentration of each antibiotic at which no visible cell growth was observed.The susceptibility of fication . All antK. pneumoniae using a sequential treatment scheme as described in Table 5 CFU/ml) of K. pneumoniae was inoculated at 37 °C in TSB treated with  and without antibiotic . After being reached an optical density (OD) of 0.5, the non-adapted and ciprofloxacin-adapted K. pneumoniae cells were further treated with no (Treatments 1 and 4) and the increased concentrations of ciprofloxacin (Treatments 2 and 5) and meropenem (Treatments 3 and 6). The survival curves were measured at 600 nm at every 4 h interval throughout the incubation period. The cultured cells were collected for further analyses, including bacterial enumeration, disk diffusion, lactamase activity, and cartwheel tests.Time-kill curve assay was carried out to determine the antibiotic activities of ciprofloxacin and meropenem against NTRT/NCON) was calculated for each treatment as compared to the control (CON). NCON and NTRT represent the numbers of control and treatments after incubation, respectively.The collected cells were serially diluted (1:10) with PBS and then plated on trypticase soy agar (TSA) using an Autoplate Spiral Plating System . The plates were incubated at 37 °C for 24–48 h to enumerate viable cells using a QCount Colony Counter . Log reduction , including cefotaxime , chloramphenicol , ciprofloxacin , kanamycin , levofloxacin , nalidixic acid , norfloxacin , sulphamethoxazole/trimethoprim , and tetracycline , were placed on the surface of the agar using sterilized forceps. After incubation at 38 °C for 24 h, the diameters of inhibition zone were measured using a metric ruler.The antibiotic susceptibility of cultured g for 20 min at 4 °C to collect cell-free supernatants. The cell-free supernatants were mixed with 20 μl of 1.5 mM nitrocefin  and incubated at 37 °C. The absorbance was measured every 5 min up to 1 h at 515 nm [The β-lactamase activity was evaluated using a nitrocefin hydrolysis assay . The cult 515 nm .K. pneumoniae cells were centrifuged at 3000×g for 20 min at 4 °C, rinsed twice with PBS, and then suspended in PBS with and without efflux pump inhibitor  [The cultured 8 μg/ml) , 16. TheAll analyses were conducted in duplicates for three replicates. Data were analyzed by the Statistical Analysis System (SAS) software. The General Linear Model (GLM) and least significant difference (LSD) procedures were used to compare treatments at p < 0.05.K. pneumoniae was observed during the sequential antibiotic treatments  compared to the ciprofloxacin-adapted cells (1/4CIP-CIP and 1/4CIP-MER). The number of viable K. pneumoniae cells under 1/4CIP-CIP was significantly reduced by 1.8 log, followed by CON-CIP and 1/4CIP-MER , while no significant reduction was observed for the 1/4CIP-MER switched from CIP to MER at the second treatment. The antibiotic resistance acquisition was evaluated in K. pneumoniae exposed to sequential antibiotic treatments . The enhanced susceptibility to all antibiotics tested was observed at the CON-CIP treatment, while the decreased susceptibilities to CIP and TET were observed at the treatments, CON-MER, 1/4CIP-CON, and 1/4CIP-MER. The treatments, CON-MER and 1/4CIP-MER, showed the similar resistance pattern as the CON.The MIC values of ciprofloxacin and meropenem were 0.03 and 0.06 μg/ml, respectively. The survival of nts Fig. . The groK. pneumoniae cultured at sequential antibiotic treatments  was evaluated on the EtBr-agar plates , piperazine (MIC > 16 μg/ml), cephalotin (MIC > 32 μg/ml), and cefoxitin (MIC > 16 μg/ml), resulted from the production of β-lactamases [K. pneumoniae cells exposed to CON-CIP showed the increased susceptibility to CTX .The strain of ctamases . No signctamases . K. pneuin MIC > 2 μg/ml, CTX Fig. . The mec MIC > 25 μg/ml, pin MIC > 2 μg/ml, K. pneumoniae treated at 1/4CIP-CIP and 1/4CIP-CIP may include CHL, TET, NOR, NAL, and LEV, corresponding the enhanced multiple antibiotic resistance in disk-diffusion assay in Fig. The efflux pump systems competitively expel substrates, which plays an important role in the development of multidrug resistance . The speK. pneumoniae. The difference in antibiotic activity against K. pneumoniae might be due to the time- and concentration-dependent modes of action of meropenem and ciprofloxacin, respectively. The antibiotic resistance mechanisms can occur within a short time in association with lactamase production, membrane permeability, and efflux pump activity. The efflux pump activities in K. pneumoniae cells treated at 1/4CIP-CON, 1/4CIP-CIP, and 1/4CIP-MER were still observed in the presence of PAβN, indicating that various types of substrate-dependent efflux pumps exist to reduce the intracellular concentrations of antibiotics. The induced efflux pump activity are responsible for the increased antibiotic resistance in K. pneumoniae. The most significant finding in this study was that K. pneumoniae treated with sequential antibiotics, specifically 1/4CIP-CIP, showed reduced susceptibility towards CHL, NOR, NAL, and LEV, leading to the multiple antibiotic resistance. Therefore, antibiotic therapies should take into account the history of pre-exposed antibiotics to prevent the development of antibiotic resistance. Further studies are needed to assess the risk of antibiotic resistance in sequential and combination antibiotic therapies, which is essential to design an effective strategy for controlling multiple antibiotic resistant bacteria.This study highlights the influence of using sequential antibiotic treatment on the development of antibiotic resistance in


Diabetes Mellitus (DM) is a huge burden for human health. Recent studies show the close relationship between DM and T cells. We investigated the trend in DM and T cells research.Using the Web of Science database, we searched the publications on DM and T cells in 1997–2016, and studied the source data using bibliometric methodology. Excel 2016, GraphPad Prism 5, and VOSviewer software were used to analyze the publication trend in DM and T cells research.We found a total of 1077 publications with 38109 citations up to January 23, 2017. The highest contribution came from the United States, with 48.38% of the publications, 61.44% of the citations and the highest H-index (74). China had the 5th place for total publications, but ranked 11th both for citation frequency (604) and H-index (13). The inflection point of the global DM and T cells publications was in 2000. Journal of Immunology published the most related articles (164). Santamaria P. was the leading scholar in this field with the most publications (35). The keywords “regulatory T cell” and “autoimmune diabetes” were mentioned more than 300 times. Furthermore, type 2 (T2)DM, T cell immunoglobulin and mucin domain (TIM) and obesity are becoming popular research topics in this field.The quantity of publications on DM and T cells grew rapidly around year 2000, but has relatively decreased recently. The United States had the leading position in global research. There was a discrepancy between productivity and quality of publications from China. Latest progress is most likely first published by the Journal of Immunology. Santamaria P., Roep B.O. and Peakman M. were the pioneer scholars in this field. Most researchers have focused on “regulatory T cell” and “autoimmune diabetes” research. In future, T2DM, TIM and obesity may be the popular areas. Diabetes mellitus (DM) is a glucose metabolism disease characterized by chronic hyperglycemia resulting from defects in insulin secretion, insulin action, or both. Type 1 Bibliometric analysis provides information about the trend in research activity over time and compares the contributions of scholars, journals, institutes and countries. It usesOur research attempted to provide all-round insights on the current state of global DM and T cells research. The distribution of the publications was analyzed as well as keywords and references to better understand the global trend of research and to discover the popular topics in this field.The Science Citation Index-Expanded (SCI-E) of Thomson Reuters’ Web of Science provides prepared and comprehensive data of publications and is considered the optimal database for bibliometric analysis. The litThe data of all eligible publications including title, year of publication, authors' names, nationalities, affiliations, name of publishing journal, keywords, abstract, times of citation per year and H-index per country were downloaded from the internet and extracted carefully and independently by two authors (Ye Gao and Yiran Wang). The data were imported into Microsoft Excel 2016, GraphPad Prism 5 and VOSviewer and analyzed both quantitatively and qualitatively.f(x) = c/[1+a×exp(-b×(x-1996))]. In this formula, the independent variable x refers to the year and the f(x) refers to the cumulative amount of publications. The formula T = lna/b+1996 was used to calculate the inflection point defined as the time the growth rate of publications changes from positive to negative. Furthermore, we used VOSviewer to analyze the relations between highly cited references and productive authors as well as to create the knowledge maps of cited references and keywords related to DM and T cell research.The intrinsic function of Web of Science was used to describe the aforementioned basic characteristics of eligible publications. The H-index was designed as a measure of scientific research impact. The index of H indicates that a scholar or country has published H papers and each of which has been cited in other publications at least H times. Therefore, the H-index reflects both the number of publications and the number of citations per publication. Relative research interest (RRI) was defined as the number of publications in a certain field divided by all-field publications per year. Impact factor (IF) of all journals were obtained from the Journal Citation Reports of 2016. GraphPad Prism 5 was used to draw the time curve of publications using the logistic regression model: Both the quantity and quality of publications were measured to evaluate the contributions of different countries and regions. A total of 1077 articles from 1997 to 2016 met the search criteria. The United States had the first place when ranking for the quantity of publications , followed by Canada  and the United Kingdom . China had the 5th place with 71 publications accounting for 6.59% of all publications. The overall number of citations of publications was 38109 since 1997 (32772 times without self-citations). The cited frequency per publication was 35.38 times. The United States had the most citations with a number of 23413 (61.44%) and an H-index of 74. Canada ranked 2nd with the citation number of 7484 and an H-index of 43. Though the number of publications of China ranked 5th, the number of citations and the H-index both ranked 11th . Most reMore than half of the publications in this field were published in the top 15 journals . The Journal of Immunology  published the most studies with 164 publications. There were 4 articles in Nature , 5 articles in Nature Immunology  and 1 article in Lancet Diabetes & Endocrinology  on DM and T cells research. The top 15 journals that published the most studies are listed in Publications from top 15 institutes accounted for 43.64% of all literature on DM and T cells research. The University of Colorado had the highest number of publications with a total of 59, thereby accounting for 5.48% of all published literature in this field. In the list of the top 15 institutes, 8 are from the United States, 2 are from Canada, 2 are from the United Kingdom, 2 are from France and 1 is from the Netherlands .The logistic regression model was used to create the time curve of the cumulative number of publications from which the inflection point could be derived and a future trend could be predicted. The inflection point (the time point that the growth rate of the number of publications changed from positive to negative) of global publications was in 2000 as derived from logistic model fitting . The infA total of 242 publications were from the top 10 authors, accounting for 22.47% of all publications related to the field. The 3 authors who published the most research were Santamaria P. with 35 publications on DM and T cells, followed by Roep B.O. with 30 publications and Peakman M. with 28 publications .Analysis of references was performed to examine the relations between references and to classify them into different categories. The top 245 references with more than 20 citations were selected for analysis of references by VOSviewer. The references included in the analysis were divided into 3 clusters. The first cluster included 90 publications and mainly focused on the autoimmune process in DM. The second cluster included 82 publications and focused on regulatory T cells in DM. The third cluster contained 73 publications on treatment  S1 Tabl.The purpose of analysis of keywords is to discover directions and popular topics in research and has proven to be important for monitoring the development of science and programs. KeywordKeywords were color coded by VOSviewer based on the average time they appeared in the 1077 related publications . The bluThe United States and Canada made the highest contributions to the global publications in terms of total number of publications as well as citation frequency and H-index. The Unites States and Canada started relatively early with studying DM and T cells. As shown in China was 5th when ranked for the total number of publications, but 11th for both citation frequency and H-index. The total number of citations and H-index of a country represent its academic impact and quality of publications. This discrepancy between the academic impact and quantity of publications may be due to the following reasons. First, studies on DM and T cells started late (in 2004) in China with the inflection point in 2008. Discoveries are difficult when following the steps of others. Besides, it takes years for the citation frequency of a publication to increase to a considerable number, even if the reported findings are important. Second, the academic evaluation system in China puts an excessive pressure on the quantity of publications, which fThe logistic model fitting curves showed that the inflection point of China (2008) was late because of the late start of research in 2004, compared to 1997 for the United States and Canada. When the cumulative number of publications reaches the inflection point, the rate of the number of publications changes from positive to negative. The global inflection point was in 2000, therefore the rate of increase in literature in this field has gradually slowed down since then. There are fewer studies published every year and new discoveries in this field are more difficult than before.The Journal of Immunology, Diabetes, Journal of Autoimmunity, Proceedings of National Academy of Sciences USA, and Clinical Experimental Immunology published the most studies on DM and T cells. Future discoveries in this field are likely to be reported by the aforementioned journals.Institutes in the United States were the leading organizations in DM and T cells research, which was consistent with the leadership of the United States in global publications. More than half (8) of the top 15 institutes were located in the United States. Meanwhile, Canadian and European institutes were also influential with 2 and 5 institutes in the top 15 list, respectively. This suggests that creating first-class research institutes is fundamental to improve the academic level of a country.Santamaria P., Roep B.O. and Peakman M. were the top 3 authors who published the most studies in the field of DM and T cells. Santamaria P. focused on the pathogenesis of autoimmune DM, especially on the role of regulatory T cells and CD8Click here for additional data file.S2 Table(PDF)Click here for additional data file.


D-methionine (Met) relative to L-Met for nursery pigs using the slope-ratio assay. A total of 50 crossbred barrows with an initial BW of 13.5 kg (SD = 1.0) were used in an N balance study. A Met-deficient basal diet (BD) was formulated to contain an adequate amount of all amino acids (AA) for 10–20 kg pigs except for Met. The two reference diets were prepared by supplementing the BD with 0.4 or 0.8 g L-Met/kg at the expense of corn starch, and an equivalent concentration of D-Met was added to the BD for the two test diets. The pigs were adapted to the experimental diets for 5 d and then total but separated collection of feces and urine was conducted for 4 d according to the marker-to-marker procedure. Nitrogen intakes were similar across the treatments. Fecal N output was not affected by Met supplementation regardless of source and consequently apparent N digestibility did not change. Conversely, there was a negative linear response (P < 0.01) to Met supplementation with both Met isomers in urinary N output, which resulted in increased retained N (g/4 d) and N retention (% of intake). No quadratic response was observed in any of the N balance criteria. The estimated bioavailability of D-Met relative to L-Met from urinary N output (g/4 d) and N retention (% of intake) as dependent variables using supplemental Met intake (g/4 d) as an independent variable were 87.6% and 89.6%, respectively; however, approximately 95% of the fiducial limits for the relative bioavailability estimates included 100%. In conclusion, with an absence of statistical significance, the present study indicated that the mean relative bioequivalence of D- to L-Met was 87.6% based on urinary N output or 89.6% based on N retention.This experiment was conducted to determine the bioavailability of  D- and L-Met which is produced through chemical synthesis  are commonly used to provide indispensable AA , which limit growth of pigs when protein sources are marginally used to reduce feed cost as well as N excretion. Methionine is essential for protein synthesis and is one of the most limiting AA for the growth of nursery pigs fed diets containing dried blood products (plasma and cells) and dried whey . Methionynthesis , but onl in pigs . Thus, iurements . The claurements  and is mreported  to the aThe Institutional Animal Care and Use Committee of Konkuk University reviewed and approved all protocols (KU13188) used in the present study.D- to L-Met during five consecutive periods. In each 9-d period, 10 pigs were allotted to five dietary treatments with two replicates per treatment in a randomized complete block design based on the initial BW.A total of 50 crossbred ((Landrace × Yorkshire × Duroc) × Duroc) nursery barrows at 35 ± 3 d of age were selected based on BW which was similar across the pigs. For adaptation to metabolism cages, the pigs were individually placed in the cages for five days. After adaptation, the pigs (13.5 ± 1.0 kg) were used to estimate the relative bioavailability of L-Met/kg at the expense of corn starch, and an equivalent concentration of D-Met was added to the BD for two test diets. To avoid any possible variation in the experimental diets caused by feed ingredients, the experimental diets were made of the same batch of ingredients, mixed at the same time and at the same location. To minimize orts, daily feed allowance was calculated as 3.5% of the BW of each animal at the beginning of each period. The feed was divided into two equal meals and fed to the pigs at 900 and 1,700 h. The pigs were provided ad libitum access to water.A Met-deficient basal diet (BD) was formulated to meet or exceed the estimated requirements of all nutrients except for Met. The BD contained total Met at 18.1 g/kg which was about 66% of the Met requirement for 10–20 kg pigs ; Table 1For the N-balance study, the pigs were adapted to the experimental diets for 5 d and then total but separated collection of feces and urine was conducted for 4 d according to the marker-to-marker procedure . The colAt the completion of the study, the frozen fecal samples were dried in a forced-air oven at 55 °C and finely ground prior to chemical analyses. The experimental diets, fecal and urine samples were determined for crude protein (CP) content (N × 6.25) by the Kjeldahl method .I is the amount of N ingested (g); NF and NU are the amount of N voided via the feces (g) and urine (g), respectively.Apparent total tract N digestibility and retention were calculated using the following equations:D-Met to L-Met was estimated using a multiple regression model and the slope-ratio analysis described by a is intercept; e is random error; sb and tb are the slopes for L- and D-Met, respectively; sx and tx are the concentrations of L- and D-Met intake, respectively. An individual pig served as the experimental unit and statistical significance was determined at P < 0.05.Experimental data were analyzed using the MIXED procedures of SAS . The independent variables in the model included diet as a fixed effect and period and block nested within period as random effects. The orthogonal polynomial contrast was used to examine the relationship between N balance response criteria and graded concentrations of Met isomers. The relative bioavailability of L- or D-Met on N balance are shown in P < 0.05) to Met supplementation from L- or D-Met in urinary N output, which resulted in increased (P < 0.01) retained N and N retention. No quadratic response was observed in any of the N-balance criteria. The estimated bioavailability of D-Met relative to L-Met from urinary N output (g/4 d) and N retention (% of intake) as dependent variables using supplemental Met intake (g/4 d) as an independent variable were 87.6 and 89.6%, respectively  and N retention (% intake) as dependent variables, are 87.6% and 89.6%, respectively, and the 95% fiducial limits included 100%. To obtain accurate values from bioavailability assays, the assumptions for slope-ratio assay should be validated  was used as the response criterion. This was in agreement with L- or DL-Met and reported no difference in utilization between Met isomers. Furthermore, no differences in BW gain or plasma urea levels were observed when three-week-old pigs received low-protein-liquid diets containing either DL- or L-Met (0.51%) for seven days  and N retention (% of intake) were 87.6% and 89.6%, respectively; but 95% fiducial limits for the relative bioavailability estimates included 100%.In conclusion, the relative bioavailability values for 10.7717/peerj.2368/supp-1Supplemental Information 1Click here for additional data file.


Antibiotics are among the most widely prescribed medications. The geographic variation in antibiotic prescribing patterns and associated costs among Medicare Part D recipients have not been described. The purpose of this study was to assess the regional variation in antibiotic prescriptions and costs among Medicare Part D enrollees in 2013.Retrospective cohort review of all Medicare Part D enrollees in 2013, using the Medicare Provider Utilization and Payment Data: Part D Prescriber Public Use File. All original or refill prescription claims for antibiotics as listed in the Part D Prescriber Public Use File were included. Our primary outcomes were total antibiotic claims and antibiotic cost per Medicare Part D Enrollee. Data were analyzed descriptively by state and by geographic region as defined by the United States Census Bureau. Antibiotic claims were described overall and by antibiotic class.Over 54 million outpatient antibiotic claims were filed for Part D enrollees in 2013, representing more than $1.5 billion in total antibiotic expenditures. Antibiotic use was highest in the South , followed by the Midwest , Northeast , and West . Average antibiotic costs per enrollee in each region were as follows: South $46.58, Northeast $43.70, Midwest $40.54, and West $36.42. Fluoroquinolones were the most commonly prescribed class overall (12.2 million claims).Antibiotic use among elderly Medicare enrollees in the United States was highest in the South region. Fluoroquinolones were the most common antibiotics used in all regions. These patterns could be utilized in the development of targeted antimicrobial stewardship efforts.The online version of this article (doi:10.1186/s12879-016-2091-0) contains supplementary material, which is available to authorized users. The advent of clinically useful antibiotics revolutionized the practice of medicine; however, the emergence of antibiotic-resistant organisms has posed a unique challenge to the utilization of these drugs. Despite advances in drug development and infection control measures, antibiotic resistance continues to be a great concern . Each yeThe overuse of antibiotics is a major factor in the development of resistant organisms . It is ePrevious studies have examined the geographic variation of antibiotic prescribing in the United States in an effort to identify areas where antibiotic stewardship efforts are most needed. Utilizing data from 2011, Hicks et al. found that outpatient antibiotic use was highest in the South and lowest in the West . At leasThe objective of this study was to describe regional variation in antibiotic prescribing and costs overall and by antibiotic class among Medicare Part D enrollees in 2013.This study utilized retrospective data from the 2013 Medicare Provider Utilization and Payment Data: Part D Prescriber Public Use File. The methodology for the development of this dataset has been described previously . In brieClostridium difficile . The corresponding brand names for each identified generic antibiotic were then again searched manually to remove all non-systemic antibiotics. Any antibiotic that could not be definitively identified as systemic use only  was excluded from the final analysis. Additionally, antibiotics that have significant use for non-infectious indications  were also excluded. A listing of each included generic and brand name is included in Additional file We limited the dataset to all original and refill prescription claims for antibiotics. The PDE Standard Analytic File was searched manually by all authors for generic names corresponding to systemic antibiotics  or antibiotics used for the treatment of Over 54 million outpatient antibiotic claims were filed for Part D enrollees in 2013, representing 1,452 claims per 1,000 enrollees and more than $1.5 billion in total antibiotic expenditures. Across regions, antibiotic claims per 1,000 enrollees  and complications from drug-drug interactions, such as QT interval prolongation [Appropriate antibiotic use is of paramount importance in elderly patients because of their higher risk for antibiotic-related adverse reactions  were among the most commonly prescribed agents in our study, with a prescribing rate of 326 claims per 1,000 enrollees, or 22% of all claims. This class of antibiotics is notable for its association with infrequent, yet severe, adverse effects which may be more likely to occur in elderly patients , 17. FurOur findings indicate the need for stewardship interventions to promote appropriate antibiotic use where the burden is highest. Prescribing rates in regions with lower antibiotic use could serve as useful benchmarks when determining goals for reduction. Several types of interventions to reduce inappropriate antibiotic prescribing have been studied in the outpatient setting, including provider and patient education, provider feedback, communication skills training, restriction policies, and integration of rapid diagnostics . All typThis study has several potential limitations. First, the dataset only contains patients enrolled under the Medicare Part D prescription drug program , and may not represent antibiotic prescribing patterns for all Medicare beneficiaries or elderly patients. Furthermore, we are unable to determine the appropriateness of antibiotic prescribing using this dataset, and it is important to note that lower prescribing rates do not necessarily reflect more appropriate prescribing. Next, the data are limited to medications covered by Medicare Part D or those covered through supplemental benefits; therefore, the results might underestimate antibiotic prescribing in this population. Lastly, this study included a single year of data, which would not capture annual changes in prescribing patterns.In summary, antibiotic use among elderly Medicare Part D enrollees in the United States was highest in the South region, confirming the regional variation seen in prior years and in other populations. Fluoroquinolones were the most common antibiotics used in all regions. These patterns could be utilized in the development of systematic antimicrobial stewardship efforts.


Necator americanus infection.IgG4-related disease is a fibroinflammatory systemic condition characterized by tumefactive lesions, lymphoplasmacytic infiltrate rich in IgG4-positive plasma cells, storiform fibrosis, and elevated serum IgG4 concentrations. It has been described in virtually every organ system. Autoimmunity and infectious agents are potential immunologic triggers in IgG4-related disease. Herein, we describe a peculiar case of effusive-constrictive pericarditis in an 18-year-old boy with polyserositis and concomitant IgG4-related disease is a systemic fibroinflammatory disease with tissue infiltration of IgG4-positive plasma cells. The histopathological features show similarities across organs; however clinical features may vary considerably. Cardiac manifestations may include pericarditis, often in a constrictive form.3/mm3), high erythrocyte sedimentation rate (ESR—87 mm/h), C-reactive protein (CRP—10.22 mg/dl), and gamma globulins , while procalcitonin (PCT) was 0.9 ng/ml (n.v. < 0.5 ng/ml). All these findings suggested an inflammatory/infectious disease.An 18-year-old boy immigrant from Guinea was admitted to our hospital because of fever and abdominal pain starting about three weeks before. His past medical history was unremarkable. Physical examination revealed diminished lung breath sounds consistent with pleural effusion and hepatomegaly. Lab tests showed abnormal blood count . The patient was empirically treated with Ibuprofen (600 mg tid) and Doxycycline (100 mg bid), until further microbiological tests (faeces and urine) revealed an Ancylostoma duodenale/Necator americanus infection. The patient was thus successfully treated with mebendazole (100 mg bid). His clinical conditions progressively improved, despite the persistence of abdominal discomfort. A new echocardiogram after 6 weeks showed an evolution to constrictive pericarditis  with significant respiratory variations  of mitral inflow and inferior vena cava plethora . All the Figures  and 5. cm with At this point, due to the lack of a definite aetiology for the polyserositis with effusive-constrictive pericarditis we suspected an IgG4-related disease. Our diagnostic hypothesis was confirmed by elevated serum IgG4 concentrations . The patient was thus treated with prednisolone at a dose of 75 mg per day for 2 weeks. The echocardiogram showed a progressive resolution of the pericardial disorder without constrictive physiology. The prednisolone was tapered to 25 mg per day and the patient was discharged. Last follow-up echocardiogram (three weeks later) showed an almost complete resolution of the pericardial disease with mild residual thickening of both layers and residual deposits of fibrin .IgG4-related disease is an increasingly recognized systemic immune-mediated condition. Its features include tumor-like swelling of involved organs, a lymphoplasmacytic infiltrate rich in IgG4-positive plasma cells, and a variable degree of fibrosis that has a characteristic “storiform” pattern . FurtherSince there is still no definitive serological test to diagnose IgG4-related disease, a combination of histological, immunophenotypic, clinical, radiographic, and laboratory investigations has been proposed to perform the diagnosis . For exaIn a recent large cohort study the histopathological evaluations were more frequently performed in the most easily accessible organ such major salivary glands, kidney, lacrimal glands, and lung; in contrast, only a minority of patients underwent pancreas or retroperitoneal/periaortic biopsy because these deeper structures are much more difficult to access .The same authors showed elevated serum levels of IgG4 in more than 95% of Japanese patients with IgG4-related disease [IgG4-related disease is now a well, although rare, recognized cause of pericardial disease  with several reports over the last years , 7.We would like to draw attention to certain key aspects of our report, first of all the clinical course of the pericardial disease in our patient. Pericardial constriction due to IgG4-related disease, once clinically manifested, often requires biopsy followed by surgical pericardiectomy. In our patient we were planning these procedures due to the worsening of pericardial pathology; however, we observed an impressive and prompt response to glucocorticoids with an almost complete resolution of the pericardial constriction after 5 weeks, as documented by the instrumental echocardiographic follow-up. Helicobacter pylori [ Paragonimus westermani infection [ Necator Americanus in our patient) causing the IgG4-related disease.The second aspect is related to the pathophysiology of IgG4-related disease in our patient. The pathogenesis of IgG4-related disease is poorly understood. Autoimmunity and infectious agents are potential immunologic triggers in IgG-4-related disease . Some str pylori . More renfection . Elevate Necator americanus infection.We conclude by recalling that IgG4-related disease, even if rare, should be considered in the diagnostic workout of patients with polyserositis and effusive-constrictive pericarditis and that there are strict relationships between parasitic infections and elevated serum IgG4 levels. To the best of our knowledge this is the first reported case of constrictive pericarditis in a patient with concomitant elevated serum IgG4 levels and


AbstractOreoglanishponkanensis. It is a member of the Oreoglanissiamensis species group and can be distinguished from its congeners in having a unique combination of the following characters: lower lip with median notch and posterior margin entire, caudal fin emarginate, nasal barbel reaching about half the distance to eye, tip of maxillary barbel rounded, posterior margin of maxillary barbel entire, absence of pale elliptical patches on sides of body below adipose fin, absence of patch on base of first dorsal fin ray, caudal fin brown with two round, bright orange patches in middle, branched dorsal fin rays 5, branched anal fin rays 2, vertebrae 40, pectoral fin surpassing pelvic fin origin, pelvic fin length 21–26% SL, caudal peduncle length 25–33% SL, caudal peduncle depth 3–5% SL, adipose fin base length 34–39% SL, and dorsal to adipose distance 12–16% SL.During a survey of the Mali Hka River drainage in Hponkanrazi Wildlife Sanctuary in December 2015, a new species was collected and is described herein as  Sisoridae is the largest family of Asian catfish, with more than 200 species and 22 genera . The Glyptosterninae includes 10 genera in one tribe (Glyptosternini).The Glyptosterninae is well-supported as a monophyletic group with 15 synapomorphies, within which Oreoglanis is monophyletic and considered to be a sister group of Pseudoexostoma and Exostoma, with five synapomorphies . Head length and body parts were measured as proportions of standard length (SL). Counts and measurements followed Kunming Institute of Zoology (KIZ), Chinese Academy of Sciences (CAS), Kunming, China, and the Southeast Asia Biodiversity Research Institute (SEABRI), Chinese Academy of Sciences, Nay Pyi Taw, Myanmar.Measurements were made point to point with dial calipers and recorded to 0.2 mm. Counts and measurements were made on the left side of the specimens when possible. Subunits of the head were measured as proportions of PageBreakTaxon classificationAnimaliaSiluriformesSisoridaehttp://zoobank.org/A539FAAD-34D9-4370-ABCA-D6048BC54CA6SL; Myanmar: Kachin State, Hponkanrazi Wildlife Sanctuary, Zeyar Stream near Zeyar Dan Village, 27°34.2'N, 97°06.05'E; XY. Chen, T. Qin and SS. Shu, 14 Dec. 2015.KIZ2015006376 (CXY20150125), 102.14 mm SL; data as for holotype. SEABRI-CXY20150143, 1 ex., 110.68 mm SL; Myanmar: Kachin State, Hponkanrazi Wildlife Sanctuary, Ponyin Stream near Zeyar Dan Village, 27°33.86'N, 97°05.42'E; XY. Chen, T. Qin and SS. Shu, 14 Dec. 2015. SEABRI-CXY20150104, SEABRI-CXY20150106, KIZ2015006378, 3 ex., 70.6–120.64 mm SL; Myanmar: Kachin State, Hponkanrazi Wildlife Sanctuary, Zeyar Stream near Zeyar Dan Village, 27°34.2'N, 97°06.05'E; XY. Chen, T. Qin and SS. Shu, 9 Dec. 2015. SEABRI-CXY20150078, 1 ex., 88.78 mm SL; Kachin State, Hponkanrazi Wildlife Sanctuary, Monlar Stream near Warsar Dan Village, 27°29.82'N, 97°11.34'E; XY. Chen, T. Qin and SS. Shu, 7 Dec. 2015.KIZ2015006375 (CXY20150124), KIZ2015006377 (CXY20150126), 2 ex., 78.88–99.26 mm Oreoglanishponkanensis is a member of the Oreoglanissiamensis species group, and can be distinguished from its congeners in having a unique combination of the following characters: lower lip with median notch and posterior margin entire, caudal fin emarginate, nasal barbel reaching about half the distance to eye, tip of maxillary barbel rounded, posterior margin of maxillary barbel entire, absence of pale elliptical patches on sides of body below adipose fin, absence of patch on base of first dorsal fin ray, caudal fin brown with two round, bright orange patches in middle, branched dorsal fin rays 5, branched anal fin rays 2, vertebrae 40, pectoral fin surpassing pelvic fin origin, pelvic fin length 21–26% SL, caudal peduncle length 25–33% SL, caudal peduncle depth 3–5% SL, adipose fin base length 34–39% SL, and dorsal to adipose distance 12–16% SL.Morphometric data are listed in Table PageBreakDorsal fin without spine and with i, 5 (7) rays. Adipose fin with long base. Anal fin with i, 2 (7) rays. Caudal fin emarginate, with 6/6 (7) rays. Pelvic fin greatly enlarged, with convex distal margin and i, 5 (7) rays; first ray flattened, with numerous plicae on ventral surface; tip of pelvic fin surpassing anus, and anus at midpoint between posterior end of pelvic fin base and tip of pelvic fin. Pectoral fin greatly enlarged, without spine and with i, 16 (4) or i, 17 (3) rays; first ray flattened, with numerous plicae on ventral surface. Tip of pectoral fin reaching beyond pelvic fin origin; Vertebrae 25+15=40 (3), or 26+14=40 (1).Males with small genital papilla located immediately posterior to anus Figure . FemalesColor. In life: brown on dorsal and lateral surfaces of head and body, light yellow on ventral region. Dorsal surfaces of head and body with series of small, light yellow PageBreakpatches: two ovoid patches on occipital region, elliptical patches on anterior and posterior bases of adipose fin. Ovoid patch on base of first dorsal fin ray absent, and elliptical patch on lateral surface of body below middle part of adipose fin base absent. Dorsal fin brown, dorsal surfaces of pectoral and pelvic fins brown, anal fin and ventral surfaces of pectoral and pelvic fins light yellow. Adipose fin light yellow. Caudal fin brown with two round, bright orange patches in middle. Pectoral fin base occasionally with round yellow patch on inner and outer anterior sides, respectively. Dorsal surface of barbels brown, ventral surface light yellow.PageBreakPageBreakKnown from high mountain streams of Mali Hka River drainage (upper Irrawaddy River drainage) in Hponkanrazi Wildlife Sanctuary, Kachin State, north Myanmar Figure .Cyprinidae: Danioaequipinnatus, Bariliusbarnoides, Torqiaojiensis, Neolissochilus sp., Garrasalweenica, Garrabispinosa, Placocheilusdulongensis, Schizothoraxmeridionalis; Nemacheilidae: Paracanthocobitisadelaideae, Schisturamalaisei; Siluridae: Pterocryptisberdmorei; Amblycipitidae: Amblycepsmurraystuarti; Sisoridae: Exostomavinciguerrae; Channidae: Channaburmanica.Fast flowing mountain streams with stone, cobble, and sand beds Figure . Other aFrom Hponkanrazi Wildlife Sanctuary, adjectival.Oreoglanishponkanensis is a member of the Oreoglanissiamensis species group .PageBreakPageBreakPageBreakOreoglanishponkanensis is further distinguished from Oreoglanissiamensis in having the posterior margin of lower lip entire , the posterior margin of the maxillary barbel entire (vs. lobulated), longer pelvic fin , absence (vs. presence) of pale patches on the sides of the body below the adipose fin, fewer branched dorsal fin rays (5 vs. 6), longer and more slender caudal peduncle , shorter nasal barbel (16–25% HL vs. 28–37), and larger interorbital distance (28–34% HL vs. 23–27).Oreoglanishponkanensis can be further distinguished from Oreoglanissudarai in having the posterior margin of lower lip entire (vs. with lobulate projections), longer pelvic fin (greatly surpassing anus vs. slightly surpassing), absence (vs. presence) of pale patches on sides of body below adipose fin absent, longer caudal peduncle (length 25–33% SL vs. 17–23), shorter prepelvic length (29–35% SL vs. 37–43), shorter anal fin base length (3–6% SL vs. 6–9), longer pelvic fin length (21–26% SL vs. 13–17), and shorter nasal barbel (16–25% HL vs. 27–41).Oreoglanishponkanensis can be distinguished from Oreoglanisheteropogon in having the nasal barbel reaching midway between its base and anterior orbital (vs. reaching anterior margin of orbit), the posterior margin of the maxillary barbel entire (vs. with crenulate projections), more branched dorsal fin rays (5 vs. 6), more vertebrae (40 vs. 38), shorter predorsal length (24–31% SL vs. 35), shorter prepectoral length (11–16% vs. 19), shorter prepelvic length (29–35% SL vs. 42), shorter preanal length (61–69% SL vs. 75), shorter dorsal to adipose distance (12–16% SL vs. 20), longer caudal peduncle (length 25–33% SL vs. 18), shorter post-adipose distance (8–12% SL vs. 13), longer adipose fin base (34–39% SL vs. 29), longer pelvic fin (21–26% SL vs. 13), shorter anal fin base (3–6% SL vs. 7), larger interorbital distance (28–34% HL vs. 22), and shorter nasal barbel (16–25% HL vs 33).Oreoglanishponkanensis differs from Oreoglanisjingdongensis and Oreoglanissuraswadii in having the caudal fin emarginate (vs. lunate). It can be further distinguished from Oreoglanisjingdongensis in having the posterior margin of the maxillary barbel entire , pelvic fin surpassing anus for a longer distance (vs. just surpassing), less blunt snout, and fewer vertebrae (40 vs. 42–43). It can be further differentiated from Oreoglanissuraswadii in having the pelvic fin greatly surpassing anus (vs. just surpassing), more vertebrae (40 vs. 36–38), absence (vs. presence) of pale patches on sides of body below adipose fin, longer caudal peduncle (25–33% SL vs. 19–25), shorter prepelvic length (29–35% SL vs. 36–40), and shorter dorsal fin base (8–11% SL vs. 11–14).Oreoglanishponkanensis can be distinguished from Oreoglanislaciniosus in having the posterior margin of the lower lip entire (vs. with lobulate projections), absence (vs. presence) of pale patches on sides of body below adipose fin, shorter predorsal length (24–31% SL vs. 35–37), shorter prepelvic length (29–35% SL vs. 38–42), and longer adipose fin base (34–39% SL vs. 32–33).Oreoglanishponkanensis shares a similar color pattern with Oreoglanisimmaculatus, but differs in having the posterior margin of the lower lip entire (vs. with lobulate projections), the posterior margin of the maxillary barbel entire , more vertebrae (40 vs. 37–38), longer pelvic fin (21–26% SL vs. 18–21), pelvic fin far surpassing anus (vs. just surpassing), much slenderer caudal peduncle , longer adipose fin base (34–39% SL vs. 26–33), and shorter dorsal to adipose distance (12–16% SL vs. 16–23).Oreoglanishponkanensis differs from Oreoglanismacropterus in having the posterior margin of the maxillary barbel entire , and shorter dorsal to adipose distance (12–16% SL vs. 18–19).Oreoglanishponkanensis can be distinguished from Oreoglanismajusculus in having the posterior margin of the maxillary barbel entire (vs. with villiform projections), absence (vs. presence) of patches on sides of body below adipose fin, fewer branched pectoral fin rays (16–17 vs. 20), fewer caudal fin rays (6/6 vs. 7/8), and much slenderer caudal peduncle (with length 25–33% SL vs. 18–21 and depth 3–5% SL vs. 6).Oreoglanishponkanensis differs from Oreoglanispangenensis in having the posterior margin of the maxillary barbel entire (vs. with lobulate and laciniate projections), fewer branched dorsal fin rays (5 vs. 6), fewer caudal fin rays (6/6 vs. 7/8), absence (vs. presence) of pale patches on sides of body below adipose fin, slenderer caudal peduncle (with length 25–33% SL vs. 23 and depth 3–5% SL vs. 5), shorter head (17–23% SL vs. 26), shorter predorsal length (24–31% SL vs. 33), shorter preanal length (61–69% SL vs. 75), shorter prepelvic length (29–35% SL vs. 36), longer adipose fin base (34–39% SL vs. 31), and shorter dorsal to adipose distance (12–16% SL vs. 21).Oreoglanishponkanensis can be differentiated from Oreoglanissetiger in having the posterior margin of the lower lip entire (vs. with laciniate projections), the posterior margin of the maxillary barbel entire , shorter predorsal length (24–31% SL vs. 32–37), shorter prepelvic length (29–35% SL vs. 37–39), and larger eye diameter (8–12% HL vs. 7–8).Oreoglanishponkanensis can be distinguished from Oreoglanisinsignis in having the distance between anal fin origin and caudal fin base almost equal to distance between pelvic and anal fin origins , anus much closer to snout tip than caudal fin base , absence (vs. presence) of pale patches on sides of body below adipose fin, black (vs. yellow) tip of caudal fin, fewer branched dorsal fin rays (5 vs. 6), fewer caudal fin rays (6/6 vs. 8/7), more vertebrae (40 vs. 36–39), shorter predorsal length (24–31% SL vs. 31–35), shorter prepelvic length (29–35% SL vs. 36–40), and longer adipose fin base (34–39% SL vs. 29–34).Oreoglanissiamensis group, Oreoglanissiamensis and Oreoglanissudarai only occur in the Chao Phraya River drainage, Oreoglanissuraswadii, Oreoglanissetiger, and Oreoglanisjingdongensis only occur in the Mekong River drainage, Oreoglanisheteropogon, Oreoglanislaciniosus, and OreoglanisimmaculatusPageBreakare found only in the Salween River drainage, Oreoglanismajusculus and Oreoglanispangenensis only occur in the Brahmaputra River drainage, Oreoglanismacropterus occurs in the Salween and Irrawaddy river drainages, and Oreoglanisinsignis and Oreoglanishponkanensis are only found in the Irrawaddy River drainage. Oreoglanisinsignis was distributed in the “upper Irrawaddy and Salween (Nu Jiang) river drainages in northern Myanmar and southwestern China” based on specimens from René Malaise’s 1934 collection “Qushi, Baoshan, Yunnan, China from the Kambawti area, Kachin state, Myanmar and Tengchong area, Yunnan, China”. Kambawti, Tengchong, and Qushi (a Tengchong Township) are located in the Irrawaddy drainage. Thus, Oreoglanisinsignis should be confined to the Irrawaddy drainage, as clarified by Within species of the Oreoglanisimmaculatus. Holotype: KIZ 200261015, 54.2 mm SL, Paratypes: KIZ200261010, 012, 014, 016, 4 ex., 57.4–63.9 mm SL, Nanjing River , Yongde County, Yunnan, China; KIZ 794762, KIZ794763, 2 ex., 52.0–54.3 mm SL, Nangun River , Cangyuan County, Yunnan, China.Oreoglanisinsignis. Holotype: KIZ 9810191, 66.7 mm SL, Longchuanjiang River (a tributary of the upper Irrawaddy), Tengchong County, Yunnan, China. KIZ 2006010198-217, 220-229, 30 ex., 41.–85.3 mm SL, upper Binglang River , Tengchong County, Yunnan, China.Oreoglanisjingdongensis. Holotype: KIZ 200104003, 109.1 mm SL, Paratypes: KIZ 200104001-002, 004-008, 7 ex., 87.1–115.2 mm SL, upper Mengpian River (a tributary of the upper Mekong), Jingdong County, Yunnan, China.Oreoglanismacropterus. KIZ 2004000749-754, 825-840, 22 ex., 43.8–85.1 mm SL, a tributary of upper Dulong River (a tributary of the upper Irrawaddy), Gongshan County, Yunnan, China.Oreoglanissetiger. KIZ 2016000859, 868, 870, 874, 883, 5 ex., 74.7–95.0 mm SL, Nanbi River (a tributary of the upper Mekong), Mengsa Township, Gengma County, Yunnan, China.


Statistical learning (SL) is the extraction of the underlying statistical structure from sensory input  and the order of SL (can higher-order transitions be effectively learned in a short exposure). The triplet learning paradigm is unable to provide insight into either of those questions because it mixes first- and second-order transitions and does not sample a range of probabilities. Other approaches such as the transition matrix paradigm (Durrant et al., Another important limitation of the triplet paradigm, which is particularly relevant for this study, is the role of prior preferences for particular triplets. Probably all participants will have had extensive exposure to Western tonal music, which results in the development of cognitive schemata (Krumhansl, Combining auditory and visual SL with a comparison of musicians and non-musicians is the main contribution of this paper. The results of this study may be interpreted as evidence of a domain-specific component to SL in keeping with other findings (Conway and Christiansen, FM, PP, and SD conceived the ideas in the article, discussed the specific arguments to be presented, and wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


L being a continuous lattice, two functors from the category of convex spaces (denoted by CS) to the category of stratified L-convex spaces (denoted by SL-CS) are defined. The first functor enables us to prove that the category CS can be embedded in the category SL-CS as a reflective subcategory. The second functor enables us to prove that the category CS can be embedded in the category SL-CS as a coreflective subcategory when L satisfying a multiplicative condition. By comparing the two functors and the well known Lowen functor , we exhibit the difference between (stratified L-)topological spaces and (stratified L-)convex spaces.Consider  Abstract convexity theory is an important branch of mathematics -convexity -convexity is precisely an M-fuzzifying convexity; when L, M)-convexity is precisely an L-convexity; and when L, M)-convexity is precisely a convexity. Similar to  topology, the categorical relationships between convexity and latticed-valued convexity is an important direction of research. When L being a completely distributive complete lattices with some additional conditions, Pang and Shi (L-convex spaces as a coreflective subcategory.It is easily seen that there is some similarity between convexity and topology (a family of subsets of a set closed for union and finite intersection). Thus, similar to lattice-valued topologies  to the category of stratified L-convex spaces (denoted by SL-CS). The first functor enables us to prove that the category CS can be embedded in the category SL-CS as a reflective subcategory when L being a continuous lattice. The second functor enables us to prove that the category CS can be embedded in the category SL-CS as a coreflective subcategory when the continuous lattice L satisfying a multiplicative condition. The second functor is an extension of Pang and Shi’s functor topological spaces and (stratified L-)convex spaces from the categorical sense.In this paper, we shall continue to study the categorical relationships between convex spaces and stratified  functor  from theCS in the category SL-CS. In “CS in the category SL-CS. Finally, we end this paper with a summary of conclusion.The contents are arranged as follows. In “a is way below b (2)(3)such that(4)for some: Maruyama  A subsetL-convex spaces is called L-convexity-preserving  provided that L-convex spaces and whose morphisms are L-CP mappings will be denoted by SL-CS.A mapping Finally, we recall some categorical notions from Adámek et al. .A and B are concrete categories; F, G) is called a Galois correspondence if B-morphism; and A-morphism. is a Galois correspondence, then F is a left adjoint of G , hence F and G form an adjunction If . It is obvious.(LC2). It is easily seen that N is the set of all choice functions For any Let (LC3) Let 2.The set It is easily seen that 3.For any We have proved that for any By a combination of (1)–(3), we have proved that Let be a convex space. Then iffThe sufficiency is obvious. We check the necessity. Let is CP iffisL-CP.L-CP. Let L-CP. Then for any Conversely, let It is easily seen that the correspondence Let be a stratifiedL-convex space. Then the setforms a convex structure onXand the correspondencedefines a concrete functor For any For any Let be directed. Thenfor anyLet L is called multiplicative  and (LC2) are obvious. We only check (LC3) below.C being closed for intersection and directed union. Let L-convex space. Then Pang and Shi convex space from (L-) topological spaces.At last, we give two interesting examples to distinguish –(2), we can define (stratified) L-convex structure on M.Let L being a continuous lattice, an embedding functor from the category CS to SL-CS is introduced, then it is used to prove that the category CS can be embedded in the category SL-CS as a reflective subcategory. When L being a continuous lattice with a multiplicative condition, Pang and Shi’s functor topological spaces and (stratified L-) convex spaces from categorical sense.When  functor  is gener