In this way, ACE inhibitory peptides derived from scale gelatin have the potential to be used as healthy ACE-inhibiting drug raw materials.Hypertension can cause coronary heart disease. Synthetic angiotensin-converting enzyme (ACE) inhibitors are effective antihypertensive drugs but often cause side effects. The aim of this study was to prepare potential ACE inhibitors from scales. Gelatin was extracted from lizardfish scales. Then, scale gelatin was enzymolyzed to prepare ACE inhibitory peptides using response surface methodology. Proteolytic conditions after optimization were as follows: pH 7.0, enzyme substrate ratio 3.2%, temperature 47 °C, and proteolysis lasting 2 h and 50 min. The experimental ACE inhibitory activity under optimal conditions was 86.0 ± 0.4%. Among the 118 peptides identified from gelatin hydrolysates, 87.3% were hydrophilic and 93.22% had a molecular weight <2000 Da. Gelatin peptides had high stability upon exposure to high temperature and pH as well as gastrointestinal tract enzymes. Gelatin peptides showed an antihypertensive effect in spontaneously hypertensive rats at a dosage of 2 g/kg in the long-term experiments. A new ACE inhibitory peptide was isolated from gelatin hydrolysates, and was identified as AGPPGSDGQPGAK with an IC Hypertension can cause cardiovascular, cerebrovascular, and renal diseases, as well as stroke . AngioteSynodus macrops) is widely distributed in tropical and subtropical waters. It is one of the most commercially important fish in China. Unfortunately, processing plants dump lizardfish scales as waste. The preparation of active peptides from lizardfish scales would not only improve the added value of fish products, but also reduce environmental pollution problems. Response surface methodology (RSM) is a statistical tool for evaluating the process parameters with the fewest experiments when many factors and interactions affect desired responses for a given process. For this reason, RSM is widely used in the industrialization of active peptides obtained from proteolysis [Wild-caught fish are important foods, accounting for approximately 50% of the total world fish production. More than 70% of this product is subjected to processing ,11,12. Dteolysis . Using tteolysis ,18. Currteolysis , whey prteolysis , lizardfteolysis . Howeverteolysis .The purpose of this investigation was to determine the optimal conditions of neutral protease hydrolysis of scale gelatin and characterize the resulting ACE inhibitory peptides, and to prepare new ACE inhibitory peptides from the protease hydrolysis of scale gelatin.p < 0.05) after 1–8 h of hydrolysis. The DH of neutral protease hydrolysate was the second highest, and the DH increased from 6.6 ± 0.0% to 10.3 ± 0.1% (p < 0.05) after 1–8 h of hydrolysis. The DHs of other protease hydrolysates were <5%, relatively stable after 1 h of hydrolysis, and generally did not change with time. The inhibition percentage (IP) of neutral protease hydrolysates remained relatively stable during 1−4 h of hydrolysis , and then decreased to 74.6 ± 0.6% (p < 0.05). Therefore, neutral protease hydrolysates had the most ACE inhibitory activity and relatively higher DHs, and these were selected for further studies.The gelatin was hydrolyzed using seven proteases, and the degree of hydrolysis and ACE inhibitory activity data are shown in The effect of E/S from 0.1% to 3% on the DH and IP of hydrolysates was determined with 50 °C and pH 7.0 for 3 h A. As theThe effect of hydrolysis temperatures from 30 °C to 70 °C on the DH and IP of hydrolysates was determined with E/S 2% at pH 7.0 for 3 h B. When tThe effect of pH from 5–9 on the DH and IP of hydrolysates was determined with E/S 2% at 50 °C for 3 h C. The hiThe effect of hydrolysis time from 1–5 h on the DH and IP of hydrolysates was determined with E/S 2% at 50 °C and pH 7 D. As hyd1) and IP (Y2) are shown in Equations (1) and (2), respectively.The central composite design and results are shown in 1, p < 0.05), E/S , temperature , and time  on DH were all significant. However, none of the interactions between any factors other than X3X4 < 0.05 were significant (p > 0.05), and the quadratic terms X12 (p < 0.001), X22 (p < 0.01), X32 (p < 0.0001), and X42 (p < 0.0001) were significant. By comparing the absolute values of coefficients of the linear terms in the equation, we were able to list the factors affecting the DH in descending order of magnitude: X3 > X2 > X4 > X1. The interaction between variables on the DH were shown in 1 (p < 0.05), X2 (p < 0.01), and X3 (p < 0.001) on IP were significant, and that there was a significant interaction between X2X3 (p < 0.05). X12 (p < 0.05) and X32 (p < 0.001) were the only significant quadratic terms. By comparing the absolute values of coefficients of the linear terms in the equation, we were able to place the factors affecting ACE inhibition percentage in descending order of magnitude: X3 > X2 > X1 > X4. The interaction between variables on the IP were shown in As shown in 1 = 0.13694, X2 = 1.08393, X3 = −0.49563, and X4 = 0.48944. That is, the optimal process conditions for the preparation of ACE inhibitory peptides with the DH as the indicator were as follows: pH 7.1, E/S 3.1%, temperature 45 °C, time 3 h and 30 min, and the DH was predicted to be 10% under the above conditions. By calculating the partial derivative of Equation (2), optimal levels of the four factors were obtained as follows: X1 = −0.04859, X2 = 1.19468, X3 = −0.33561, and X4 = −0.13719. That is, the optimal process conditions for the preparation of ACE inhibitory peptides with the ACE inhibition rate as the indicator were as follows: pH 7.0, enzyme dosage 3.2%, temperature 47 °C, reaction time 2 h and 50 min, and the ACE inhibition rate under the above conditions was predicated to be 86.3%. The optimal proteolysis conditions to produce ACE inhibitory activity peptides were different from the optimal proteolysis conditions to reach the highest DH. At the initial stage of proteolysis, ACE inhibitory peptides are released from the scale gelatin by protease, and the DH and ACE inhibitory activity are enhanced at this stage. As proteolysis progresses, some ACE inhibitory peptides are enzymolyzed by the protease into peptides with weaker ACE inhibitory activity. The time spent in proteolysis needed to produce the highest DH is longer than that needed to obtain the strongest ACE inhibitory activity [Data for the partial derivative of the equation for DH appear in Equation (1). The optimal levels of the four factors were as follows: XAs shown in A grand average of hydropathicity (GRAVY) index value was used to evaluate the hydrophilicity and hydrophobicity of peptides. When GRAVY > 0, the peptide is hydrophilic, and when GRAVY < 0, the peptide is hydrophobic. As shown in The molecular weight (MW) distribution analysis of the identified peptides B showed p > 0.05) across different temperature and pH treatments. Data show that the ACE inhibitory activity of peptides obtained from proteolysis has thermal and pH stability. These peptides can be easily transported and stored after they are developed into a product. As shown in p > 0.05), indicating that they had strong resistance to the digestive enzymes and were true inhibitor-type ACE inhibitory peptides [As shown in peptides . García 7d: 185.6 ± 14.9 mm Hg, Sample7d: 161.8 ± 12.8 mm Hg, p < 0.05, n = 10). The decreases in SBP caused by gelatin peptides were more pronounced than that caused by 2 mg/kg of captopril from day 7 through day 21. This suggests that the gelatin peptides produced an antihypertensive effect in SHR at a dosage of 2 g/kg per rat in the long-term experiments.As shown in TM CaptoTM Q chromatography into four fractions (The peptides present in gelatin hydrolysates were fractionated by HiTrapractions . Fractioractions . Fractio50 = 420 ± 20 μM) was higher than that of Ala-Val (IC50 = 956.3 μM) [50 = 570.8 μM) [50 > 1000 μM) [50 = 420 ± 20 μM) was lower than that of Ala-Lys-Lys (IC50 = 3.1 μM) [50 = 77 μM) [50 = 34.7 μM) [The amino acid sequence of purified peptide was found to be Ala-Gly-Pro-Pro-Gly-Ser-Asp-Gly-Gln-Pro-Gly-Ala-Lys (AGPPGSDGQPGAK). As shown in 56.3 μM) , Ala-Arg70.8 μM) , and Pro1000 μM) . The ACE 3.1 μM) , Trp-Leu= 77 μM) , and Leu34.7 μM) . These d34.7 μM) .Synodus macrops Tanaka) was purchased from a fishery factory in Zhangzhou, China. Scales were separated manually, washed with water, and then stored in polyethylene bags at −20 °C until use. High molecular weight markers, angiotensin I-converting enzyme (from rabbit lungs), hippuryl-l-histidyl-l-leucine (Hip-His-Leu), vitamin C, pepsin, trifluoroacetic acid (TFA), and formic acid were obtained from Sigma Chemical Co. . Pepsin (3 × 106 U/g) was purchased from Shanghai Generay Biological Engineering Co., Ltd. . Papain (8 × 106 U/g), trypsin (3 × 105 U/g), bromelain (1 × 106 U/g), chymotrypsin (1 × 105 U/g), neutrase (2 × 106 U/g), acid protease (1 × 105 U/g), and alcalase (2 × 106 U/g) were purchased from Nanning Pangbo Biological Engineering Co., Ltd. . LC-grade methanol and acetonitrile were purchased from Merck . All other reagents used in this study were reagent grade chemicals.Lizardfish  and the E/S ratio was 1:100 (w/w). Samples were hydrolyzed for 1, 2, 3, 4, 5, 6, 7, and 8 h. Subsequently, the hydrolysate was centrifuged at 20,000× g for 15 min. The supernatant was lyophilized and then stored at −20 °C until use.The gelatin sample was added to 200 mL of pure water and enzymolyzed with bromelain , alcalase , chymotrypsin , papain , acidic protease , trypsin , and neutral protein , separately. After the enzymatic reaction was completed, the temperature was raised to 100 °C for 5 min to deactivate enzymes. The gelatin/solution ratio was 4:100 .2). Taking the amount of free amino groups in gelatin hydrolysates minus the amount of free amino groups existing in gelatin, the number of peptide bonds produced by the hydrolysis of gelatin can be determined. Additionally, taking the amount of free amino groups in gelatin completely hydrolyzed by HCl minus the amount of free amino groups originally existing in gelatin, the total number of peptide bonds in gelatin can be determined. The DH of gelatin hydrolysate can be obtained by the ratio of the number of peptide bonds produced by the hydrolysis of gelatin to the total number of peptide bonds in gelatin. A mixture of 1 mL of 0.13 mg/mL gelatin hydrolysate sample solution, 1 mL of pure water, 1 mL of 0.5% ninhydrin solution, and 1 mL of 0.1% ascorbic acid solution was heated in boiling water for 15 min and allowed to cool at room temperature for 4 min. Subsequently, 7 mL of 45% (v/v) ethanol solution was added and the mixture was shaken rapidly. The optical density of the mixture was read at 580 nm and free amines in the sample were measured using a ninhydrin colorimetric curve. The DH was calculated using Equation (4).The method reported by Zhang et al.  was modi1 is the free amino groups in gelatin hydrolysates; B2 is the originally existing free amino groups in the gelatin; and B3 is total free amino groups in the gelatin hydrolyzed by 6 M HCl at 100 °C for 24 h.Here, Bv/v) containing 0.1% TFA for 10 min at a flow rate of 1 mL/min and monitored at 228 nm. ACE inhibitory activity (IP) was calculated, and A0 and A were chromatographic peak areas of HA with and without sample. ACE inhibitory activity was calculated using Equation (5).The method reported by Cushman and Cheung (1971)  was modi0 and A are chromatographic peak areas of HA with and without sample. The IC50 value is defined as the concentration of inhibitor that can inhibit 50% of the ACE inhibitory activity.Here, IP is the ACE inhibition percentage (%) of the sample; A1), E/S , temperature , and time  were chosen as independent variables, and the range and center point values are shown in 1) and ACE inhibitory activity  were selected as dependent variables for combining independent variables, as shown in RSM was used to determine the optimal proteolytic conditions for the fish scale gelatin, and the experiment was optimized using a five-level four-factor Box-Behnken central composite design. pH . βv/v/v). In brief, 5 μL of samples were loaded onto a self-made 5-μm Reprosil C18 AQ column (75 μm × 150 mm) for separation at 25 °C. The mobile phase consisted of acetonitrile/formic acid/water  and acetonitrile/formic acid/water ; the gradient elution starting with 2% buffer B and ending with 30% buffer B lasted for 150 min. LTQ-Orbitrap operated in data-dependent acquisition mode and could automatically switch between the full scan (m/z 300–2000) in Orbitrap and the higher energy collisional dissociation (HCD) MS/MS scan in a linear ion trap. Helium was used as a collision gas for HCD. The normalized collision energy was 35% and the activation time was 30 ms. Data collection was controlled by Xcalibur 2.0.7 and Tune 2.4 software .The method reported by Liu et al. (2017)  was modiAll MS/MS data were processed using Xcalibur (version 2.0.7.) and PEAKS 7.5  was used. The spectra were also searched against an equal number of decoy sequences to estimate the false discovery rate, as previously described. When the variables were designed, “specified enzyme” was selected as a non-enzyme, allowing up to two restriction sites to be missed. Methionine oxidation (+15.99), protein N-terminal acetylation (+42.01), amidation (−0.98), deamidation (+0.98), formylation (+27.99), carbamylation (+43.01), methyl ester (+14.02), and carbamoyl methylation of cysteine (+57.02) were used as modifications. Default settings were used for other variables, including fragment ion tolerance set at 0.5 Da and maximum allowed mass deviation set at 6 ppm for the precursor ions after recalibration. The de novo sequence was used to identify the peptide sequence to produce the dataset, with an average local confidence (ALC) score >90%. The identified peptides were screened to create a dataset with a value of −10 logP greater than 60.−1 solution, which was allowed to stand at 4, 20, 40, 60, 80, and 100 °C for 2 h, respectively. After samples were cooled to room temperature and pH was adjusted to 8.3 with NaOH and H2SO4, ACE inhibitory activity was measured as described in The method reported by Wang et al.  was used−1 solution, whose pH was adjusted to 2, 4, 6, 8, 10, and 12, respectively. These samples were allowed to stand at 37 °C for 2 h. After samples were cooled to room temperature and the pH was adjusted to 8.3 with NaOH and H2SO4, ACE inhibitory activity was measured as described in The method reported by Wang et al.  was used−1 pepsin solution (pH 2.0), and phosphate buffer solution was used to prepare 0.05 mg·mL−1 trypsin solution (pH 8.0) and 0.05 mg·mL−1 chymotrypsin solution (pH 8.0). The gelatin hydrolysates were subjected to three digestion tests. Samples were incubated with pepsin solution with E/S 1:12 for 2 h at 37 °C. The pepsin was deactivated for 10 min at 100 °C and the pH was adjusted to 8.3. Then the mixture was centrifuged at 10,000× g for 10 min, and the supernatant was collected and lyophilized to measure the ACE inhibition percentage. After the pepsin treatment, samples were incubated with trypsin solution with E/S 1:12 for 2 h at 37 °C. The trypsin was deactivated for 10 min at 100 °C and the pH was adjusted to 8.3. The mixture was centrifuged at 10,000× g for 10 min, and the supernatant was collected and lyophilized to measure the ACE inhibition percentage. After the treatment of pepsin and trypsin, samples were incubated with chymotrypsin solution with E/S 1:12 for 2 h at 37 °C. The chymotrypsin was deactivated for 10 min at 100 °C and the pH was adjusted to 8.3, the mixture was centrifuged at 10,000× g for 10 min, and the supernatant was collected and lyophilized. The ACE inhibitory activity was measured as described in The stability of ACE inhibitory peptide in the presence of gastrointestinal proteases was assessed in vitro using a slightly modified version of the method described by Vercruysse et al. . KCl-HClSHRs  with tail SBP over 180 mmHg were obtained from Beijing Vital River Laboratory Animal Technology Co., Ltd. . SHRs were housed individually in steel cages in a room kept at 24 °C under controlled lighting from 8:00 to 20:00, and fed a standard laboratory diet. SBP was measured by the tail-cuff method with an Non invasive blood pressure (NIBP) system . Scale gelatin peptides prepared from lizardfish were dissolved in saline and administrated orally at a dose of 2 g/kg body weight. Captopril was used as a positive control and was administrated the same way as the scale gelatin peptides at a dose of 2 mg/kg body weight. Control rats were administrated orally with the same volume of saline solution. Oral administration was performed for 35 days. The SBP was measured on days 0, 7, 14, 28 and 35.TM CaptoTM Q ion-exchange column (7 × 25 mm), equilibrated with 20 mM Tris-HC buffer (pH 8.0), and eluted with a linear gradient of NaCl (0–1 M) in the same buffer at a flow rate of 1 mL/min. The absorbance of the eluent was monitored at 228 nm. Fractions showing ACE inhibitory activity were collected and lyophilized. One ACE inhibitory activity fraction was re-dissolved in super-pure water. After that, the solution was further purified on a self-made Sephadex G15 gel column (2.5 × 15 cm), and eluted with super-pure water at a flow rate of 1 mL/min. The absorbance of the eluent was monitored at 228 nm and 280 nm. The injection of the sample was 1 mL. The fraction showing ACE inhibitory activity was lyophilized. The ACE inhibitory activity fraction was re-dissolved in super-pure water. After that, 1 mL of solution was loaded onto a UNO Q1 ion-exchange column (7 × 35 nm), equilibrated with 50 mM Tris-HC buffer (pH 8.0), and eluted with a linear gradient of NaCl (0–2 M) in the same buffer at a flow rate of 4 mL/min. The absorbance of the eluent was monitored at 228 nm. The purified peptide was collected and lyophilized for further use.One milliliter of gelatin hydrolysates was loaded onto an HiTrapThe amino acid sequence of the purified peptide was determined by automated Edman degradation using a Shimadzu PPSQ-33A protein sequencer. The purity of peptide was determined by RP-HPLC on a SinoChrom ODS-BP C18 column (4.6 × 250 mm) using the linear gradient of acetonitrile (6–94%) containing 0.1% TFA in the same buffer at a flow rate of 1 mL/min, while monitoring the absorbance at 220 nm. The molecular mass of peptide was determined by Waters ZQ2000 mass.50 value of 420 ± 20 μM. This indicates that our prepared ACE inhibitory peptides may be suitable for further development into health products and drugs raw materials. RSM was used to optimize proteolytic conditions for gelatin hydrolysates. The optimal conditions were as follows: pH 7.0, E/S 3.2%, temperature 47 °C, and proteolysis time 2 h and 50 min. The experimental ACE inhibition rate of the gelatin peptides obtained under the optimal conditions was 86.0 ± 0.4%, which is consistent with the predicated value of 86.3%. The gelatin peptides, prepared from lizardfish scales, reached a yield of 10.8% (dry weight). There were 118 peptides identified from the gelatin peptides. Among them, 87.3% were hydrophilic peptides and 93.2% had an MW of less than 2000 Da. There were 26 peptides with C-terminal Arg residues and eight peptides with C-terminal Lys residues, and these 34 peptides may have ACE inhibitory activity. Gelatin peptides show strong high stability under exposure to extremes of high temperature and pH, and the digestive enzymes in the gastrointestinal tract, suggesting that gelatin peptides may pass through the membrane barrier in the form of whole peptides to be directly absorbed by the body. In the long-term experiments, the gelatin peptides showed an antihypertensive effect in SHR at a dosage of 2 g/kg per rat. The decrease in SBP caused by gelatin peptides was greater than that caused by 2 mg/kg of captopril from days 7–21. Although the dosage of gelatin peptides was larger than that of the captopril, gelatin peptides were prepared from food resources and could be eaten daily without toxic side effects. In addition, a new ACE inhibitory peptide was isolated from gelatin hydrolysates and identified as AGPPGSDGQPGAK, with an IC


Helicobacterpylori bacterium is one of the main causes of chronic gastritis, peptic ulcers, and even gastric cancer. It affects an average of half of the world population. Its difficult eradication depends upon multi-drug therapy. Since its classification as a group 1 carcinogenic by International Agency for Research on Cancer (IARC), the importance of H. pylori eradication has obtained a novel meaning. There is considerable interest in alternative therapies for the eradication of H. pylori using compounds from a wide range of natural products. In the present study, we investigated the antibacterial property of the isocoumarin paepalantine against H. pylori and it exhibited significant anti-H. pylori activity at a minimum inhibitory concentration (MIC) of 128 μg/mL and at a minimum bactericidal concentration (MBC) of 256 μg/mL. The scanning electron microscopy (SEM) revealed significant morphological changes of the bacterial cell as a response to a sub-MIC of paepalantine, suggesting a penicillin-binding protein (PBP) inhibition. Computational studies were carried out in order to study binding modes for paepalantine in PBP binding sites, exploring the active and allosteric sites. The data from the present study indicates that paepalantine exhibits significant anti-H. pylori activity, most likely by inhibiting membrane protein synthesis.The  Helicobacter pylori is a gram-negative bacterium that infects the gastric mucosa and is considered one of the main etiological agents of chronic gastritis, eventually leading to the development of peptic ulcers and gastric cancer pyran-1-one), isolated from the capitula of Paepalanthus bromelioides (SILV.) [The isocoumarin paepalantine  , has pro (SILV.) ,29, anti (SILV.) ,31, anti (SILV.) , and cyt (SILV.) .H. pylori, conducting in vitro and in silico analysis of possible molecular targets.Thus, based on the previous studies with paepalantine which showed antibacterial and antioxidant potential, the objective of the present study was to investigate the effect of paepalantine on the viability and morphology of H. pylori; paepalantine showed a MIC of 128 μg/mL and a minimum bactericidal concentration (MBC) of 256 μg/mL. The vehicle used (propylene glycol) had no inhibitory effect on the micro-organisms at the concentrations used in the assay. Since the bacterial strain used in this assay was amoxicillin sensitive and metronidazole resistant, only amoxicillin presented MIC and MBC values, which were 0.25 μg/mL and 2 μg/mL, respectively.H. pylori growth and viability led us to study bacterial morphology after exposure to a MIC and sub-MIC. The treatment of H. pylori with CIM and ½ MIC (64 μg/mL) of paepalantine resulted in morphological changes in the membrane that were not observed in the control. In the control treatment, the H. pylori cells presented regular shapes with even surfaces, whereas the cells treated with paepalantine were damaged and exhibited altered or irregular shapes, such as spheres and cell surface blebs .The effect of paepalantine on ce blebs . These mThe development of the docking model was based on the analysis of the crystallographic chemical structures of the available penicillin-binding proteins complexed with betalactamic inhibitors, obtained from the Protein Data Bank (PDB) database ,37 Tabl. The PDBPenicillin-binding proteins (PBPs) have an important hole in the maintenance of the bacterial cell wall. The penicillin-binding proteins with an allosteric site binding mechanism promotes a conformational change during its binding that results in a slight opening of the active site, making it more accessible for substrate binding. Finding inhibitors with affinity for both the active and allosteric site is a promising strategy for drug discovery. In order to properly evaluate the affinity of paepalantine for PBPs, it was necessary to investigate both the active and allosteric binding mechanism, since this investigation could result in an additional and unique inhibition pattern ,38.Molecular docking calculations were performed using the Molecular Operating Environment suite (MOE) . For docThe pose prediction accuracy of the docking software was evaluated by redocking analysis. The redocking process consists of a high-quality structural model of the protein structure, usually extracted from an X-ray crystal of a protein-ligand complex. Then, the co-crystal ligand is extracted from this file, leaving only the protein that is prepared for docking calculation. Once the calculation is performed, the docking poses were compared to the protein-ligand complex. The measure of this deviation is represented by the root mean square deviation (RMSD) measure. Values below 2 Å deviation are considerable acceptable ,42. The The developed model was based on the inhibitor cefuroxime, since it is the smallest co-crystallized ligand available amongst the other two ligands, tebipenem and biapenem . As a reThe docking calculations for the active site consisted of the use of a 3D shape-based volume surrounding the core interactions combined with a hydrogen bond acceptor feature . Then, tA pharmacophore model contains Boolean expressions to encode concepts representing a hydrogen bond donor, hydrogen bond acceptor, hydrophobic, aromatic, anionic, etc., indicated by colored spheres named pharmacophore features ,44. One The docking score values for paepalantine were in a range similar to the reference inhibitor cefuroxime: S = −5.7 to −5.4 for the active site and S = −6.4 to −5.3 for the allosteric site. The values are in accordance with the reference values, S = −7.8 to −6.5 for the active site and S = −6.8 to −5.8 for the allosteric site, evidencing that the proposed model was able to describe the main interactions. For more information about the docking score values, access the available H. pylori, the ideal model for treatment against this infection is yet to be discovered [H. pylori. The micro-environment created by this bacterium is a further factor that influences the eradication rate. Specifically, the most severe clinical manifestation associated with certain H. pylori strains is most likely due to a higher degree of inflammation caused by these bacteria [Despite many years of experience in fighting scovered . Mechanibacteria ,46,47.In this context, alternative treatment approaches must be developed and the research of natural products is a valuable resource in drug discovery . The useH. pylori effects of paepalantine, which exhibited significant activity, with a MIC of 128 µg/mL and a MBC of 256 µg/mL. Structure-activity studies with coumarins suggest that the presence of antimicrobial activity is directly related to the lipophilicity and planar structure of the molecule [In the present study, we evaluated the anti-molecule ,51. Its molecule . Additiomolecule ,29. H. pylori caused by subinhibitory concentrations of substances with antibiotic activity. The analysis of β-lactam antibiotics via transmission electron microscopy (TEM) has shown a strong link of bacterial penicillin-binding proteins (PBPs), characterized by large spherical cells with few cytoplasmic elements and eventual membrane rupture, thus suggesting PBP63 inhibition as the main mechanism responsible for these changes [Some authors have observed morphological changes in  changes ,34. OtheH. pylori subjected to the sub-MIC of paepalantine revealed spherical, irregular shapes, which were related to membrane protein inhibition.The SEM morphological analysis of H. pylori after exposure to the sub-MIC of paepalantine included swelling of the bacillary forms, bleb formation on the cell surface, and the emergence of spherical shapes. These observations suggest that the target of paepalantine activity may be located at the cell surface, acting as a permeability barrier. The bactericidal mechanism of paepalantine against H. pylori might then be found in the disruption of the permeability barrier within cell membranes.The morphological changes observed in Based on the results from the experimental tests (Scanning electron microscopy), in silico studies comprised by docking analysis were conducted to investigate the potential interactions of the metabolite paepalantine in the binding site of the penicillin-binding proteins (PBP) ,53. The The analysis of the allosteric site evidences that paepalantine can be buried deep inside its pocket and still accesses the main core interactions of the inhibitor cefuroxime in its non-covalent binding pattern. The hydroxyl groups of paepalantine are near a region of interaction with the key amino acids Pro424 and Arg463. The carbonyl group interacts with the nitrogen atom of Arg426 .H. pylori [Comparing the antibacterial activity of the metabolite paepalantine with other active compounds and analogues , the doAnalyses concerning the allosteric site suggests that the interaction of the naphthoquinone derivative  a with thPaepalanthus bromelioides was collected at Serra do Cipo, Minas Gerais State, Brazil, and identified by Dr. Paulo Takeo Sano from the Institute of Biosciences, University of Sao Paulo. The voucher specimen  is on file at the herbarium in the Department of Botany, Institute of Biosciences, University of Sao Paulo, Brazil. Dimethyl sulfoxide (DMSO), peroxidase (77332), and deoxyribose (121649) were purchased from Sigma Chemical Co. . Propylene glycol was purchased from Neon Comercial Ltda. . Paepalantine was obtained according to the procedure previously reported by Vilegas et al. (1990) , and theH. pylori activity was assessed by determining the minimum inhibitory concentration (MIC) using the spectrophotometric method of broth micro dilution according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI) . Microplate wells were filled with 100 µL of liquid growth medium  containing different concentrations of test agent (16–1024 µg/mL). Another 100 µL containing H. pylori (ATCC 43504) suspension (approximately 106–107 bacteria per mL) was added to each well. Absorbance was read in a spectrophotometer at a wavelength of 620 nm, and the microplates were then incubated at 36–37°C in the presence of 10% CO2 for 72 h. After incubation, the plate well contents were homogenized, and a new spectrophotometric reading was performed under the abovementioned conditions. The tests were performed in triplicate and repeated at least three times, together with growth controls (absence of sample) and negative controls (culture medium containing different concentrations of test agent to control for color). The MIC was determined graphically and was defined as the lowest concentration of antibacterial agent at which there was a sharp decline (90%) in absorbance. Amoxicillin and metronidazole were used as standards.Anti-H. pylori bacteria after exposure to the MIC and a sub-MIC was performed by scanning electron microscopy (SEM). For sample preparation, culture medium containing bacteria exposed to paepalantine was aliquoted and centrifuged at 4000 rpm for 5 min. The supernatant was discarded, and 1 mL of 0.1 M sodium cacodylate buffer (pH 7.2) was added. After centrifugation, the cell pellet was resuspended with 200 µL of 0.1 M sodium cacodylate buffer (pH 7.2). One aliquot was placed in the center of a cover slip and allowed to dry. The material was then fixed with 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.2) for 30 min. Finally, the sample was dehydrated with alcohol, and the material was sputter coated for analysis in a JEOL® JSM-6610LV scanning electron microscope  at an accelerating voltage of 10 kV [A morphological analysis of of 10 kV ,57,58.Molecular docking calculations were performed using the Molecular Operating Environment suite (MOE) .The protThe docking model for the active site was developed to investigate the potential of binding for small inhibitors, such as the investigated metabolite, and thus an appropriate protein-ligand complex with a small ligand inside was selected. Since the active site comprises an acylation mechanism, this chemical bond was undone, restoring the original hydroxyl group of the serine amino acid (Ser337) and applying the described methods for protein preparation and hydrogen bond optimization . The oriFor the allosteric site, the placement method was set to “triangle matcher”. The selected scoring function was set to “London dG” and the free energy of binding was estimated using the force field-based scoring function “GBVI/WSA” for both models. The three-dimensional structures of the penicillin-binding proteins were obtained from the RCSB Protein Data Bank . The chep < 0.05.Data are expressed as the mean ± standard deviation and were subjected to analysis of variance (ANOVA) and linear regression. The significance level was set at binding site. The investigation of natural products as a source of new scaffolds for drug development is an interesting strategy and must be investigated.Beta-lactam antibiotics are a class of broad-spectrum antibiotics introduced in the later stages of World War II and represents one of the most important contributions to medical science in recent history. The beta-lactams remain the most widely utilized antibiotics for several reasons, such as high effectiveness, low costs, ease of delivery, and minimal side effects . One of H. pylori activity, most likely by inhibiting membrane protein synthesis. Thus, paepalantine may be a promising molecule for the treatment and prevention of diseases caused by H. pylori.The data from the present study indicates that paepalantine exhibits significant anti-H. pylori agents based on molecular modeling techniques are important strategies to implement during the drug design process since the actual multi-therapy with amoxicillin and clarithromycin associated with a proton pump inhibitor such as omeprazole is no longer effective due to the prevalence of antibiotic resistance [The initial molecular modeling docking calculations suggests that the isocoumarin paepalantine should be investigated for PBP inhibition, since one of the main mechanisms that could help its effectiveness against the PBPs, based on the results of the electron microscopy, is that it will not contain the beta-lactamic ring, and thus the beta-lactamases will be ineffective ,63,64. Isistance ,66.H. pylori colonization by the inactivation of bacterial DNA gyrase, which is a common mechanism among coumarin derivatives and urease inhibition.However, other mechanisms of action cannot be ruled out, such as the interruption of


Individuals with tears collected during spring or summer were 3.9 times (p = 0.028) more likely to have high IgE compared to those sampled at other times of year. Subjects born in the US were 3.45 times (p = 0.010) more likely to have high IgE compared to individuals born outside the US. To conclude, a majority of individuals with DE symptoms and/or signs had detectable IgE levels in their tears. High tear IgE levels were correlated with allergy season and exposures in the home linked with allergy.Dry eye (DE) and allergic conjunctivitis may present similarly, and it remains unclear whether some individuals have an underlying allergic component to their DE. To better understand this relationship, we performed a cross-sectional study in 75 individuals with DE symptoms and/or signs. Immunoglobulin E (IgE) levels in tear samples were quantified and home environmental exposures assessed via standardized survey. Tears were collected by Schirmer strip, and total tear IgE levels were quantified using enzyme-linked immunosorbent assay (ELISA). Data were analyzed using descriptive statistics and linear and logistic regressions. The main outcome measures were total tear IgE levels and their association with environmental exposures. The mean age of the subjects was 66.2 ± 7.8 years. Sixty-two individuals had dry eye symptoms (Dry Eye Questionnaire-5 ≥ 6), and 75 had one or more signs of DE. Detectable total tear IgE levels were observed in 76% of subjects, and 17.3% had high levels (>1 ng/mL). Individuals with exposure to pet(s) (odds ratio (OR) 11.5,  Dry eye (DE) is a multifactorial condition broadly defined by inflammatory and/or neurosensory changes at the ocular surface, accompanied by derangements of the tear-film . AffecteImmunoglobulin E (IgE) is an antibody that plays a critical role in type 1 hypersensitivity reactions and has been used as a biomarker to assist in the diagnosis of allergic disease ,23. TotaStudy Population: Male and female subjects were prospectively recruited from the Miami VA Medical Center eye clinic between October 2010 and December 2011. Written informed consent was obtained from all subjects after verbal explanation of the nature and possible consequences of the study.Inclusion criteria included the presence of DE symptoms and/or signs, grossly normal eyelid and corneal anatomy, no evidence of ocular surface pathology beyond DE , and no characteristic findings of AC . Based on review of the medical record, patients were excluded from participation if they had ocular or systemic conditions that could confound DE, including contact lens wear, a history of refractive surgery, use of ocular medications except for artificial tears, active external ocular pathology, cataract surgery in the last 6 months, or a history of glaucoma or retinal surgery. Systemically, individuals with human immunodeficiency virus, sarcoidosis, Sjögren’s, graft-versus host disease, collagen vascular disease, or primary immunodeficiency with elevated serum IgE  were excluded. Subjects with a history of omalizumab use, symptomatic AC, or specific allergic sensitization confirmed by skin or conjunctival provocation test or serum-specific IgE were also excluded.The Miami VA institutional review board approved the examination of patients for this study, which was conducted in accordance with the principles of the Declaration of Helsinki (IRB Protocol #3011.02) and complied with the requirements of the United States Health Insurance Portability and Accountability Act.Clinical assessment and home environment survey: Each individual filled out a standardized questionnaire regarding DE symptoms  and an IgE Analysis: Schirmer strips from the right eye of each patient were used. The Schirmer strips were defrosted and total tear proteins were eluted from the wetted area of the strip into 30 µL BSS by first vortexing for 10 s, followed by gentle shaking for 1 h at room temperature, followed by an additional vortexing for 10 s. The sample was centrifuged for 1 min, and the solution was removed and kept on ice. An additional 30 µL BSS was added to the strips, and the samples were processed as above. The eluates were pooled and then stored at −80°C until analysis.ELISA was performed on the eluted tear samples using Human IgE ELISAPRO kit , following the manufacturer’s directions. Each eluted sample was diluted to a final volume of 200 µL using BSS, and equivalent volumes of 100 µL of each sample were tested in duplicate. These precautionary volume adjustments were performed to ensure standardization of tear fluid volumes in the samples. Negative controls (assay background control and blanks) were included to measure background signals from the plate and the positive control was a standard IgE stock solution provided with the kit. The limit of total tear IgE detection was 0.069 ng/mL. The ELISA was quantified using a microplate reader at 450 nm (BMG Labtech Omega microplate reader). The data were fit to a 4-parameter standard curve, and the average of each sample was calculated after subtracting the blank from the standard for each run . We chose not to measure allergen-specific tear IgE because potential allergens were unknown at the time of analysis.Statistical Analysis: All statistical analyses were performed using STATA Version 14 . Descriptive analyses were employed to examine study variables. We examined the contribution of independent variables on total tear IgE, both as a continuous variable using linear regression and as an ordinal variable using ordinal logistic regression (function ologit) and treating IgE level below the detection limit as the base. For the categorical analysis, we binned our subjects into 3 groups: <0.69 ng/mL (non-detectable (ND)), ≤1 ng/mL (Low), and >1 ng/mL (High). We chose 1 ng/mL (0.417 IU/mL) as the cutoff between Low and High IgE status based on prior studies ,28,29.A total of 75 individuals with DE symptoms and/or signs, but without a diagnosis of AC were included. Mean age was 66.2 years ± 7.7, and the majority of subjects were male (87%), white (60%), and non-Hispanic (67%). Sixty-two individuals had dry eye symptoms (DEQ5 ≥ 6), and 75 had one or more signs of DE. Mean DE symptoms (via DEQ5) were in the moderate range (11.04 ± 5.32), and most individuals had evaporative DE  with adequate tear production . Fifty-seven individuals (76%) had detectable IgE levels in their tears (≥0.069 ng/mL) and 13 (17.3%) had High total tear IgE, defined as a value >1 ng/mL based on prior literature ,28,29 T.Total tear IgE levels were not found to correlate with DE measures, both when considering symptoms and signs of disease. For example, High tear IgE was observed in 17% of individuals with negligible DE symptoms (DEQ5 < 6), 27% of individuals with mild-moderate symptoms (DEQ5 ≥ 6 and <12), and 11% of individuals with severe symptoms (DEQ5 ≥ 12) . While np = 0.003). Likewise, individuals who smoked  or were exposed to second hand smoke at home  had a higher frequency of High IgE compared to individuals not exposed to smoke at home. Finally, tear collection during local allergy season (spring and summer) associated with a higher frequency of High IgE (29.0% vs. 9.1%) and lower frequency of ND IgE (12.9% vs. 31.8%) compared to samples collected during fall and winter (p = 0.032) (p = 0.008) and pet(s) in home  showed significant association both with total tear IgE at a log scale and total tear IgE categories. Measurement during spring or summer   was a significant risk factor, while birthplace outside the US was a significant protective factor  based on categorical analyses. In the analysis of total tear IgE as a continuous variable, patients below the limit of detection were excluded, restricting the analysis to 52 patients. IgE levels among smokers and patients with a pet at home were 2.86 (p = 0.033) and 2.16 (p = 0.049) times higher than among non-smokers and patients without a pet, respectively. Even exposure to second-hand smoke had a significant positive association with total tear IgE levels (p < 0.01).The presence of environmental allergens and/or irritants inside the home positively associated with detectable total tear IgE levels . Specifi= 0.032) . These f= 0.032) . SmokingWe found that a majority of individuals with clinical features of DE but no diagnosis of AC had detectable total tear IgE levels. Seventeen percent had High levels, defined as >1 ng/mL. Individuals with High tear IgE levels were more likely to be exposed to allergens and irritants in their home environment. Our data provide indirect evidence that some individuals with symptoms and/or signs of DE have an allergic component underlying their disease. However, we did not find that one specific manifestation of DE associated with tear IgE, as levels were distributed equally amount the different DE measures. The only trend noted was between Schirmer score and IgE levels, with high tear production positively associating with total tear IgE. Interestingly, elevated tear production has been well described in AC .Overall, the mean total tear IgE level (for those with detectable levels) in our population was 0.363 IU/mL (0.873 ng/mL), which is higher than prior reports from healthy controls , but lower than what has been reported in patients with AC  ,31,32,33n = 2817) [There is biologic plausibility for the observed relationships between our noted exposures and ocular surface parameters. Pet dander is already known to drive several IgE-mediated conditions, including AC , so it i = 2817) ,38.As with all studies, our findings must be considered in light of the study limitations. First, there are technical concerns in total tear IgE measurement, as sample volumes differed between subjects with variable Schirmer wetting lengths. We attempted to mitigate this limitation by normalizing tear fluid volumes for analysis and then by controlling for tear volume in the multivariable analysis. Indeed, Schirmer length did not significantly associate with IgE, suggesting that his was not a confounding factor in our data. Second, tear IgE levels may be elevated for reasons other than ocular allergy. For example, it is possible that some of our cohort had increased conjunctival vessel permeability secondary to ocular surface inflammation, which could lead to increased total tear protein and IgE content. As such, future studies are needed that evaluate other potential biomarkers of allergy, such as trypase , eosinopDespite these limitations, our study findings suggest that a proportion of individuals with clinical features of DE have an underlying component of ocular allergy. While it is not known if tear IgE levels will be the best biomarker to identify these patients, we found several modifiable exposures associated with total tear IgE levels in a heterogeneous group of individuals without a formal AC diagnosis. Identification of such exposures is crucial for the development of preventative and targeted interventions that are more cost effective and less risky than traditional therapies. Future studies, in diverse and well defined populations (such as those with data on skin testing), will be needed to clarify the role of IgE as a biomarker of allergy in individuals with DE.


Increased summer cloudiness triggers rapid ablation zone expansion in north Greenland, amplifying the mass loss. Since the early 1990s, the Greenland ice sheet (GrIS) has been losing mass at an accelerating rate, primarily due to enhanced meltwater runoff following atmospheric warming. Here, we show that a pronounced latitudinal contrast exists in the GrIS response to recent warming. The ablation area in north Greenland expanded by 46%, almost twice as much as in the south (+25%), significantly increasing the relative contribution of the north to total GrIS mass loss. This latitudinal contrast originates from a different response to the recent change in large-scale Arctic summertime atmospheric circulation, promoting southwesterly advection of warm air toward the GrIS. In the southwest, persistent high atmospheric pressure reduced cloudiness, increasing runoff through enhanced absorption of solar radiation; in contrast, increased early-summer cloudiness in north Greenland enhanced atmospheric warming through decreased longwave heat loss. This triggered a rapid snowline retreat, causing early bare ice exposure, amplifying northern runoff. Since the 1990s, mass loss from the Greenland ice sheet (GrIS) has significantly accelerated (−1 for 1958–2017). Simulated GrIS climate and SMB components are evaluated using in situ SMB measurements collected in the accumulation  and ablation zones ; meteorological observations recorded at 37 automatic weather stations (AWSs) from the Programme for Monitoring of the Greenland Ice Sheet (PROMICE), Institute for Marine and Atmospheric Research Utrecht (IMAU), and Greenland Climate Network (GC-Net) networks (figs. S2 and S3); and meltwater discharge measured at the Watson River in west Greenland (fig. S4B). The results show that RACMO2.3p2 realistically represents near-surface temperature, specific humidity, wind speed and air pressure (0.73 < R2 < 0.98), cloud conditions through shortwave and longwave radiation components (0.85 < R2 < 0.96), SMB (R2 = 0.78), and Watson River meltwater discharge (R2 = 0.91). We distinguish seven Greenland sectors . The model evaluation is discussed in more detail in Materials and Methods.To address this, here, we combine SMB of a dedicated, high-resolution (5.5 km) run from the Regional Atmospheric Climate Model , statistically downscaled to 1 km , with ba sectors —NW, NO , in south Greenland (red) and significantly positive (P = 0.001) in north Greenland .−1 (fig. S6C). In the South, the ELA migrated by ~100 m from ~1350 masl (fig. S6D) to ~1450 masl (fig. S6E), increasing runoff by ~17 Gt year−1 (red line in fig. S6F). We conclude that the faster snowline retreat in the North is responsible for its enhanced contribution to Greenland runoff totals  and up to 20% locally (29 ± 4 g m−2). In contrast, northerly advection of relatively dry inland air reduces cloud content over the southern GrIS by 2 to 6% on average (−3 to −6 ± 4 g m−2), in line with (−2 on average) and NO (−3.2 ± 1.9 W m−2) sectors, while it mostly increases elsewhere (−2 on average (−2) and in NO (5.3 ± 1.7 W m−2) sectors, where increased clouds further enhance LWd compared to, e.g., SW Greenland (2.6 ± 1.7 W m−2). Therefore, the near-surface temperature increase is amplified in northern Greenland  relative to other sectors . Note the marked longitudinal contrast in cloud content change between NO-NW and NE sectors, where cloud content is reduced by 2% on average and down to 14% locally (−1 g m−2 down to −10 ± 4 g m−2). This is due to a large-scale Foehn effect, resulting in relatively warm and dry air flowing down the lee side of the main northern ice divide, after having generated enhanced orographic precipitation at the upwind slopes of the NW and NO sectors. The average circulation and radiative flux anomalies shown in We relate the latitudinal contrast in runoff increase to the recent change in the large-scale summertime atmospheric circulation. The latter change, observed in the Arctic from the late 1990s onward . As a relsewhere . In cont average  due to o−2) and refreezing capacity , i.e., the fraction of meltwater and rainwater that is retained and/or refrozen within the firn pack. Figure S8 shows similar results for the South region. In the South, the decrease in runoff contribution, i.e., the difference between the red and orange solid lines in −1 or 8% to North Greenland runoff totals . In late summer (September), rainfall in North Greenland occasionally even equals runoff over bare ice, in line with  and ablation zones (+46%), twice as fast as in the south (+14 and +25%). As a result, North Greenland has significantly increased its relative contribution to total GrIS mass loss through enhanced runoff. Superimposed on increased melt, rising temperatures and increased cloudiness in the North also led to a summer rainfall increase of ~42%, contributing 5 Gt yearWe used a new run at 5.5-km horizontal resolution of the polar (p) version of RACMO2.3p2 for the period 1958–2017. For detailed description of the model and recent updates, we refer to  , i.e., 2-m air temperature and specific humidity, 10-m wind speed and surface pressure, and radiative fluxes (fig. S3), i.e., short/longwave down/upward radiation , to daily measurements collected at 37 AWS (green dots in fig. S1A). These AWSs are operated by the PROMICE (18 sites for 2007–2016)  and LWd (2.7 W m−2) radiation  instead of 41% previously, with a smaller bias and RMSE reduced by 230 and 140 mm we or 38 and 11%, respectively. This is due to the better resolved SMB patterns over narrow glaciers and marginal ablation zones at a resolution of 5.5 km (fig. S1A) compared to 11 km [figure 1 of . For instance, at stake QAS_L (291 masl), i.e., the lowest site of the 30-km-long Qagssimiut transect at the southern tip of the GrIS, strong winter precipitation and summer ablation gradients are not well resolved, resulting in SMB biases of up to 8 m we for summer 2010 (fig. S1C). In these rugged marginal regions, a spatial resolution of 5.5 km remains insufficient to resolve narrow glaciers and confined ablation zones that are prime contributors to GrIS runoff totals (R2), with a positive bias of 70 mm we, suggesting a small runoff underestimation. In addition, a comparison between modeled and observed meltwater discharge from the Watson River catchment in SW Greenland (gray contour in sector SW of −1 (or km3) (1976–2016) and up to ~2 Gt year−1 for exceptional melt episodes that occurred in summers of 2010 and 2012 (fig. S4B). The average runoff bias reaches 5% for the period 1976–2016, peaking at ~20% for extreme runoff years (2010 and 2012), in line with (R2 < 0.91). Last, a recent comparison between the period 2003–2017 mass changes derived from the Gravity Recovery and Climate Experiment (GRACE) and from a combination of modeled RACMO2.3p2 SMB at 1-km resolution and estimated solid ice discharge showed good agreement on both GrIS-wide and individual basin scales . To discard invalid albedo records, we masked erratic albedo grid cells from the GrIS-wide, daily 16-day MCD43A3 MODIS product (2000–2018), resorting to full Bidirectional Reflectance Distribution Function (BRDF) inversions. Valid daily MODIS data were classified as ice- or snow-covered grid cells using an upper threshold for shortwave albedo of 0.60 (SW0.60), slightly higher than the assumed maximum albedo of bright bare ice (0.55). The justification for this is to ensure that the maximum bare ice area is fully captured by MODIS. Subsequently, the daily ice/snow cells were converted to annual bare ice area assuming that the bare ice is exposed in summer if (i) the current pixel is classified as ice at least 5 days in that year (5th percentile) and (ii) that pixel is located within the long-term modeled ablation zone of the GrIS , i.e., including both the bare ice and lower percolation zone below the long-term ELA of ~1450 masl. These criteria allow the elimination of spurious bare ice conditions, e.g., meltwater lakes, meltwater runoff streams, and superimposed ice, that are detected at higher elevations in the percolation zone of the GrIS. For instance, in summer 2012, MODIS detected bare ice conditions at AWS S9 (1520 masl) located close to the equilibrium line along the K-transect in western Greenland (67°N), whereas fieldwork reported superimposed ice conditions resulting from firn saturation. Masked pixels were then filled on the basis of the recurrence of ice/snow observations over 2000–2018 for that cell. In brief, masked pixels exposing ice more than 50% of the time for the period 2000–2018 are assumed bare ice. This method may result in bare ice area overestimation in early years of MODIS operation, which includes more invalid albedo estimates. Alternatively, daily bare ice conditions could have been extrapolated onto masked pixels based on surrounding grid cells. However, we discarded this approach because (i) local MODIS pixels may not be representative of surrounding areas and (ii) a large number of MODIS records were masked due to, e.g., a large solar zenith angle and local cloud cover, deteriorating the quality of local MODIS estimates. Extrapolation may thus strongly overestimate remotely sensed (observed) bare ice area for the entire 2000–2018 period. That is why we used the ice recurrence method to obtain a spatially continuous annual BIE product covering the whole of the GrIS for the period 2000–2018 (fig. S5A). The MODIS product is sensitive to the shortwave albedo threshold used to discriminate between ice and snow conditions. To reflect this, we estimated an uncertainty in maximum MODIS BIE by repeating the analysis described above using the assumed maximum albedo of bright bare ice of 0.55 (SW0.55) as an upper threshold. The BIE uncertainty was estimated GrIS wide and for individual sectors as followsRemotely sensed annual bare ice area was derived from broadband shortwave clear sky albedo data from the MCD43A3 MODIS 500-m daily albedo product (The latter uncertainty was used to derive a symmetric error band around the MODIS bare ice area estimates in 2 (fig. S5C) peaking for exceptional melt years, e.g., 2012, 2014, and 2016. This negative bias, i.e., up to 16,100 km2 in 2014, mainly originates from the CE (~25%) and SE (~25%) sectors of the GrIS (fig. S5D). One possible explanation is that, in CE and SE sectors, RACMO2.3p2 simulates too much snowfall in winter, delaying or preventing the exposure of bare ice in summer. At the same time, these two sectors also experience frequent overcast conditions, negatively affecting the quality of the MODIS bare ice product. In other sectors, RACMO2.3p2 agrees well with observed bare ice area  with a small negative bias (140 km2).For model evaluation that is consistent with the MODIS product, we estimated a 10-day bare ice area in northern sectors  and a 5-day one for southern regions. Daily modeled bare ice area was estimated as the integrated area of pixels showing a surface albedo of ≤0.55 on the original 5.5-km grid, bilinearly interpolated to the 1-km GIMP ice mask. The 10 and 5 largest daily estimates of bare ice area are averaged for North and South Greenland, respectively, to obtain a maximum annual bare ice area comparable to MODIS data. This is deemed reasonable as MODIS records 16-day albedo and because less valid MODIS albedo estimates are available for the North  compared to the South (79%). This difference stems from large solar zenith angles at high latitudes, negatively affecting the quality of the satellite measurements. Figure S5A shows minimum and maximum annual BIE derived from MODIS for the period 2000–2018, i.e., summer of 2000 (black) and 2012 (red). Modeled GrIS bare ice area (orange) agrees well with MODIS , although with a negative bias of ~8100 km


Mangifera Indica L., has been investigated extensively because of its remarkable pharmacological effects. In vitro recombinant UGTs-catalyzed glucuronidation of 4-methylumbelliferone (4-MU) was used to investigate the inhibition of mangiferin and aglycone norathyriol towards various isoforms of UGTs in our study, which evaluated the inhibitory capacity of MGF and its aglycone norathyriol (NTR) towards UDP-glucuronosyltransferase (UGT) isoforms. Initial screening experiment showed that deglycosylation of MGF into NTR strongly increased the inhibitory effects towards almost all the tested UGT isoforms at a concentration of 100 μM. Kinetic experiments were performed to further characterize the inhibition of UGT1A3, UGT1A7 and UGT1A9 by NTR. NTR competitively inhibited UGT1A3, UGT1A7 and UGT1A9, with an IC50 value of 8.2, 4.4, and 12.3 μM, and a Ki value of 1.6, 2.0, and 2.8 μM, respectively. In silico docking showed that only NTR could dock into the activity cavity of UGT1A3, UGT1A7 and UGT1A9. The binding free energy of NTR to UGT1A3, 1A7, 1A9 were −7.4, −7.9 and −4.0 kcal/mol, respectively. Based on the inhibition evaluation standard , an in vivo herb–drug interaction between MGF/NTR and drugs mainly undergoing UGT1A3-, UGT1A7- or UGT1A9-catalyzed metabolism might occur when the plasma concentration of NTR is above 1.6, 2.0 and 2.8 μM, respectively.Mangiferin (MGF), the predominant constituent of extracts of the mango plant  Extensical dose ,50. To tcal dose . As demoGF < 3%) . HoweverIn conclusion, the specific inhibition of MGF and its aglycone NTR was demonstrated in this manuscript. The deglycosylation of MGF into NTR strongly increased the inhibitory effects towards others tested UGT isoforms except UGT1A10. In silico prediction model was successfully employed to explain the different inhibition effects of MGF and NTR on UGT1A3, UGT1A7 and UGT1A9. The inhibition kinetic type and behavior were determined for the inhibition of NTR towards UGT1A3, UGT1A7 and UGT1A9, and using IVIVE to clarify the possibility of herb–drug interaction between MGF/NTR and drugs. Based on these results, clinical close monitoring the utilization of norathyriol is very important and necessary.d-glucuronide (4-MUG), Tris-HCl, 7-hydroxycoumarin, and uridine-diphosphoglucuronic acid (UDPGA) . All other compounds were of high-performance liquid chromatography (HPLC) grade or of the highest grade commercially available. For enzyme sources, recombinant UGT isoforms expressed in baculovirus-infected insect cells were purchased from BD Gentest Corp. , including UGT1A1, 1A3, 1A6, 1A7, 1A8, 1A9, 1A10, 2B4, 2B7, 2B15, and 2B17.Mangiferin and norathyriol were purchased from BioBioPha Co. , and the purity of these two compounds was demonstrated to be above 95%. Sigma-Aldrich  provided the following constituents in the incubation mixture: 4-methylumbelliferone (4-MU), 4-methylumbelliferone-β-® BEH C18  at a flow rate of 0.2 mL/min and UV detector at 316 nm. The mobile phase was consisted of ultrapure water containing 0.5% formic acid (A) and acetonitrile (B). The following gradient condition was used: 0–3.5 min, 10–65% B; 3.5–4.0 min, 65% B; 4.0–9.0 min, 10% B. To calculate the standard curve 0.1–100 μM of 4-MUG was used through drawing the peak area ration of 4-MUG/internal standard towards the concentration range of 4-MUG. The curve was linear over this concentration range, with an r2 value > 0.99.4-MU, a nonspecific probe substrate for all the UGT isoforms, was employed to investigate the inhibition of MEF and NIR towards the activity of UGT isoforms. The incubation and analytical methods have been previously described ,54,55. AComparative homology modeling with MODELLER9v14 program was carried out to elucidate the three-dimensional structure of UGT1A, UGT1A7 and UGT1A9, respectively ,57. The Ki value. The determination of IC50 was calculated using Probit analysis in SPSS11.5 . The Student’s t-test was adopted at a significance level of p < 0.05 to determine statistically significant differences among experimental groups. In vitro-in vivo extrapolation (IVIVE) was performed using the following equation:The inhibition type and Kinetics were determined for the inhibition of NTR towards UGT1A3, UGT1A7 and UGT1A9. The glucuronidation velocity of 4-MU was determined at multiple concentrations of 4-MU and NTR. Dixon and Lineweaver–Burk plots were employed to determine the inhibition type, and the second plot of the slopes from the Lineweaver–Burk plot versus the compound concentrations was utilized to calculate the in vivo is the in vivo exposure concentration of CAG, and the Ki value was in vitro inhibition constant. The evaluation standard was as follows: [I]/Ki < 0.1, low possibility; 0.1 < [I]/Ki < 1, medium possibility; [I]/Ki > 1, high possibility.The terms are defined as follows: AUCi/AUC is the predicted ratio of in vivo exposure of xenobiotics or endogenous substances with or without the co-exposure of CAG. [I]


The western gray whale population is endangered with approximately 175 individuals and 33 known reproductive females. Photo-identification studies were conducted from 2002–2017 during the gray whale foraging season off northeastern Sakhalin Island, Russia. Despite abundant prey resources, significant variation in whales’ body condition, inter-birth intervals and calf survival have been documented with limited understanding of factors that account for the observed variability. We examine sea ice concentrations at their known foraging grounds to define the maximum duration of a “foraging season”. We explore the relationship between foraging season length during a female’s pregnancy and post-weaning calf survival and reproduction. Approximately 77% of the variation in calf survival, which ranged annually from 10–80%, was associated with the duration of the feeding season while the mother was pregnant. Poor body conditions and prolonged inter-birth intervals of western gray whales have also been documented to coincide with shorter duration in feeding seasons found in this study. These results imply that shorter foraging seasons are associated with reduced energy intake by physically limiting the number of days gray whales can forage, and thus sea ice conditions may be one limiting factor affecting growth rates of this endangered population of baleen whales. Eschrichtius robustus) that reside only in the North Pacific Ocean5. On the eastern side of the North Pacific Ocean, gray whale recovery from commercial whaling has been a conservation success story with population sizes reaching pre-whaling numbers6. However, gray whales on the western side of the North Pacific remain a small remnant of their historical population estimates7 and are currently listed as endangered by IUCN with a population size of 175 individuals, with only 33 known reproductive females9. In the past two decades, the western population has increased in size at an average rate of 2–4% per annum with recruitment almost exclusively from calf production with little to no immigration8. Despite the overall positive growth rate, several demographic parameters suggest that the western population are not realizing their maximum productivity levels with low post-weaning calf survivability and prolonged inter-birth intervals for reproductive females11. In addition, body conditions of western gray whales have notably varied over time with significantly worse conditions prevalent in some years compared to others12. Although anthropogenic activity has been hypothesized to limit pregnancy and calf survival rates8, other natural factors such as prey availability and/or access to their foraging habitat have also been suggested to explain the variability in population growth rates and reproductive success12.There are two genetically distinct populations of gray whales  with the number of feeding days that their mothers experienced during the previous foraging season when they were pregnant  or two years prior to the observed calf arriving on the feeding grounds . Sensitivity analyses indicated that significant associations between calf survival and the number of feeding days during the mother’s pregnancy were found with median ice concentration values for feeding days being defined as 10–40% concentration with the strongest relationship occurring with 40% median concentration criteria. The definition of feeding days using higher median ice concentrations (e.g. >40%) showed no significant association with calf survivability, which could suggest that gray whale feeding may have been inhibited at these higher median ice concentration levels.At the end of a foraging season, a fasting and pregnant female relies solely on stored energy she acquired during the foraging season when she was pregnant  to successfully bring a calf back to the feeding grounds the following year. Examining post-weaning calf survival as a function of sea ice coverage during pregnancy provides a potential link between possible poor energy intake during pregnancy due to ice, reduced female condition and lower calf survival even after weaning has occurred. Calf survival was significantly correlated  was linearly correlated to sea ice conditions during the previous year. Perryman et al.31 examined the number of calves that had survived to reach central California. Our calf counts represent the number of calves that survived to reach the foraging grounds, after potential additional mortality, such as predation, may have occurred. In addition, the small number of females that may be pregnant in any given year means individual stochasticity plays an important role in calf counts. The total number of calves off Sakhalin was not associated with the number of feeding days in the previous foraging season. In fact, the highest number of observed calves (15) was recorded in 2011, but only five of those calves were observed to continuously return to Sakhalin foraging grounds in subsequent years. The number of calves in this study was not likely representative of true calf production due to variable photo-identification research effort on the foraging grounds34 and mortality events during their first 6–7 months of life. It is possible that calf production was impacted by shorter feeding seasons prior to individuals reaching the feeding grounds while further decreased survival of the calves continued even after calves reached the feeding grounds with their mothers. In addition to providing less nutrients to calves during the migration, struggling lactating females that arrive with a calf could potentially wean calves earlier due to insufficient energy reserves to further support the calf. Early weaning could result in decreased calf growth and survival and, thus, could impact overall population growth.Annual sea ice conditions were found to account for 77% of the annual variability in calf survival in western gray whales. These results are comparable to the effects of sea ice conditions found for eastern gray whales. Perryman 17. Therefore, these shorter than average foraging seasons could explain lower than average survival of calves in 2006–2008 and 2011. Pregnant females who experience limited access to the foraging habitat early in the season could potentially “compensate” by either foraging more efficiently or staying later in the foraging season. However, when ice conditions early in the feeding season are prolonged and ice formation builds up earlier in the same feeding season off northeastern Sakhalin, this limits compensation opportunities. Females could also risk reproductive success by extending their foraging period as they could give birth outside of the breeding grounds which could increase predation of their calf by killer whales (Orcinus orca) as well as place thermoregulatory stress on the newborn calf. In recent years, eastern gray whale calves were recorded to have been born north of their breeding grounds which has been suggested as a result of delayed southbound migration due to increased competition for food resources or limited early access to prey due to sea ice conditions35. The results of this study suggest gray whales will compensate to some degree as the shorter feeding seasons were a result of both prolonged ice conditions early in the season coupled with early arrival of sea ice in the same year. In other years, such as 2013, sea ice was extended early in the feeding season to mid-June, but a rather mild onset of ice arrival in December could have allowed females to extend their foraging opportunities and thus increased the probability of survival of their calf. This study further suggest that gray whale compensation during prolonged ice conditions early in the feeding season is more likely a result of staying on the foraging grounds longer rather than by the possibility of individuals feeding more efficiently. While this study only considered calf survival if they returned to the Sakhalin foraging grounds, the unobserved calves could have survived and emigrated. In fact, limited data collected off Kamchatka, Russia in 2012–2013 identified three calves from 2011 that have never been observed to have returned to Sakhalin in subsequent years37. Although these calves may have survived, they ultimately did not contribute to growth rates of the western gray whale population.This study is the first to examine the potential duration of the gray whale foraging season off Sakhalin, as determined by sea ice conditions. The observed ice conditions on their feeding grounds may have limited access to prey resources for prolonged periods of time in some years. In particular, pregnant females feeding in 2005–2007 and 2010 had less than a 200 day foraging season. Gray whale energetic models assume a pregnant female would need approximately 200 foraging days to sustain them and their calfet al.12 found significant annual variation in body condition in western gray whales which they hypothesized to be related to environmental limitation(s) or density-dependent regulation. Body conditions of western gray whales were significantly better in 2004 and worse in 1999, 2006 and 200712. Poorer body conditions in 2006–2007 coincide with shorter foraging seasons in the previous year as estimated in this study, suggesting females return from fasting in a poorer nutritive state if the foraging season prior was shorter in duration. Conversely, foraging opportunities in 2003–2004 represented longer foraging seasons, which could explain the significantly improved body conditions observed in 2004. Although this study was limited to photo-identification data obtained from 2002–2017, sea ice concentrations in 1998 (219 feeding days) do not appear to account for poor body conditions observed in 1999 (213 feeding days) by Bradford et al.12. Eastern gray whales experienced an unusually high mortality event in 1999/2000 which was hypothesized to be related to prey availability or alternatively climate effects in the North Pacific Ocean39, although there are no prey data from that time period. Of the three calves identified by Bradford et al.12 in 1999, none were recorded to have returned to the Sakhalin foraging grounds . Therefore, sea ice conditions alone do not appear to account for the anomalous calf survival or the whales’ body condition in 1999.Bradford 8. Cooke et al.8 hypothesized that low recruitment could be related to anthropogenic activity (pile driving) that was conducted in 2008. This hypothesis seems unjustified for calf survival as pile driving activity in 2008 was localized and short in duration with relatively low sound exposure levels compared to pile driving activity in the subsequent year, which was more extensive. In addition, other known activities, such as seismic surveys, would have likely had more of an impact on calf survival due to the larger spatial extent, longer duration and higher sound exposure levels47. Restricted access to their feeding area due to ice conditions in 2005–2007 would be a more plausible explanation to the unusually low calf survival and pregnancy rates in 2007–2008 found in population models.Western gray whale population modelling efforts have further documented an exceptionally low pregnancy rate in 2008 as well as low calf survival/annual return of calves in 200711. Based on photo-identification data from 2002–2017, a total of 24 two-year and 9 three-year inter-birth intervals have been recorded to date34. Seven of the nine (77%) of the longer three-year birth intervals occurred to mothers known to have given birth after a shorter-than-usual feeding season during their pregnancy. This could suggest that a pregnant female’s reproductive success may not only decrease due to a lower probability of calf survival, but shorter foraging seasons may also result in a delay of successive pregnancies due to a compromised body condition and the need to replenish reserves.In addition to calf survival and body condition, western gray whales have prolonged and highly variable inter-birth intervals48, other studies suffer from limited sample size and inability to address cumulative impacts for the entire foraging season. For example, behavioral and distribution changes during a seismic survey in 2010 found relatively minor changes in gray whale movement, respiration and habitat use relative to sound exposures46. However, these studies had limited sample size due to the timing and duration of the seismic survey and did not account for impacts of a subsequent seismic survey that occurred in the same feeding season that had a longer duration and likely greater exposure to the population. The impacts of these activities cannot be excluded as a potential source of influence relative to calf survival and growth rates of this population.Impacts from anthropogenic activity related to oil and gas development near the western gray whale feeding grounds could have also contributed to the observed calf survival. Foraging seasons with lower calf survival, worse body conditions and prolonged inter-birth intervals were also confronted with human activities related to pile driving, seismic surveys, platform and pipeline installations and dredging activities. Although impact studies have found changes in behavior, abundance and distribution of gray whales relative to some of these activities within some feeding seasons49. Our results imply that the timing of sea ice cover can predict the post-weaning survival rate, which affects population growth and, thus, recovery of this endangered population of whales. Alternative feeding areas within the Sea of Okhotsk are currently unknown. Given the high site fidelity and residency on the Sakhalin foraging grounds, there might be no other comparable prey resources available in situations when ice or other environmental and/or anthropogenic factors prevent access to their annual foraging areas.Compared to their eastern counterparts, the western gray whale population is likely more susceptible to ice conditions due to their spatially static and geographically small foraging grounds. In addition, the western gray whale small population size, high site fidelity and limited observations of the population feeding in alternative areas would make them less resilient to environmental changes50. Pigmentation patterns on the whales body was the primary feature used to identify individuals, with scars and barnacle patches being secondary identification markings. Annual photo-identification efforts varied in timing , platforms  and number of teams. Photo-identification surveys were also conducted annually in both offshore and nearshore feeding areas with varying effort 51, were extracted for a feeding season. These remote sensing data were downloaded from the National Snow and Ice Data Center . The gridded resolution of each raster file was 25 km2. The median value of grids overlapping the historical known western gray whale nearshore feeding grounds52 was calculated on a daily basis for the period of 15 May–15 December in 2002–2017 (Figs. 54.To examine the number of foraging days for a given feeding season, daily ice concentration data, based on passive microwave sensor data from the SSMR and SSMI satellites17 Figs. . Median


Pinctada imbricata radiata pearl oysters made during this study revealed this species’ ability to implement active movement. Byssal threads were secreted in a sequence of attachment and detachment phases, which resulted in the active displacement of the oyster. The oyster was observed, in the laboratory over a 9 day period, travelling a distance of 28cm in a horizontal path. After horizontal displacement, a vertical climbing phase was observed until the oyster reached the water surface at which point the byssus was discarded and the animal dropped, drifting in accordance with water current intensity. It is possible that these adaptations of byssal use are a result of environmentally induced evolutionary change within P. i. radiata.A number of molluscs within the Class Bivalvia are defined by their ability to secrete fine silk like threads known as byssus which are used to anchor themselves to solid substrates. With relatively few exceptions the majority of these species remain in a sedentary state throughout their life attached via their byssal threads. However, observations of adult  Pinctada imbricata radiata  [The Arabian-Persian Gulf is renowned globally as one of the most challenging ecological marine environments due it arid conditions ,2. Annuah, 1814) . These rh, 1814) ,6. HowevP. i. radiata belongs to the family Pteriidae and as a byssal attached bivalve is not considered a “true” oyster by malacologists [Pinctada, is relatively unique amongst oysters as the functional byssal gland remains active throughout the life cycle [The pearl oyster ologists –9. Durinologists –13. Settologists ,15. The ologists  and the ologists  maximisefe cycle  and simife cycle . With thfe cycle –21.P. i. radiata whereby byssal threads are used as a means of active transport and not only as an attachment material. Documented observations explain how the byssus can be secreted, used and discarded in a successive action to actively drag the bivalve horizontally over substrate and climb vertical structures.This research documents a unique observation in adult P. i. radiata samples were collected by divers during September 2018 from an inshore site close to the coastal port of Semaisma on the eastern coast of Qatar. The oysters were located on a sand/shell substrate at a depth of 5m close to sea grass beds in a water temperature of 32 oC. The specimens were transported from the survey site to the laboratory using an in-situ water sample. There is no specific permission required to collect oysters in this site and this study does not involve endangered or protected species.On return to the laboratory, two large individuals (7cm) were placed in a 10lt aerated aquarium containing water from the sample site. The oysters were positioned 28 cm apart independently of each other at the lateral end of the aquarium . Observain-situ information is also presented. Specimens were photographed in-situ and in the laboratory using an underwater camera (Canon Mark-ii and Fantasea housing FG7X-II).A descriptive account of Daily Laboratory observations of the Semaisma samples began 24hrs after the oysters had been positioned in the aquarium. The first observational session recorded the visible displacement of one specimen moving towards the other. It was noted that the oyster was secreting a single byssal thread and contracting the muscle in the foot to drag itself along the aquarium. The byssal thread was fixed to the lateral side of the aquarium and discarded after movement had taken place, directly after this event another thread was secreted and attached to repeat a further displacement phase. A recognized sequence of byssal secretion, attachment, drag and discard was observed throughout the sessions. The oyster discarded a linear sequence of horizontally attached byssal threads which followed the path of movement along the aquarium glass . A totalOn reaching the second specimen the oyster was observed undertaking a vertical climb of the aquarium wall . It was in-situ (Um-Bab), oysters were observed climbing all vertical substrate, including the branches of red algae. Initially the specimens were observed attached in clumps within crevices to among rocks  using byssus identifies the species within the recognized locomotion-attachment group of the seventh ecological category of bivalves [P. radiata during this study can be considered unique as it represents the fastest benthic displacement for a byssaly attached species within the group. Comparable studies have recorded bivalve displacement at maximum speeds of < 25cm in one month therefore 28 cm in 9 days represents a significant increase in maximum locomotive speed [The basic biology, anatomy and attachment-displacement strategies of bivalves, can be directly related to their modes of life . The musbivalves  and is sbivalves . Howeverve speed . This prPinctada. A genus which contains 19 species distributed worldwide, several of which have important economic and environmental significance [Pinctada i radiata is an isolated evolutionary behaviour or is it a phylogenetic trait within the Pinctada species. It is probable that the foot is also connected to the observed movements as the threads are produced by the byssal gland located in the foot [P. i radiata may have applications in the wild, possibly as an escape strategy from disagreeable environmental conditions or as a means of re-attachment to an oyster reef matrix after dislodgment.This study adds a new factor in regards to the displacement mechanisms for oyster species within the genus ificance . Displacificance ,21,23. Hthe foot  however Pinctada imbricata radiata is a recognized bioengineering ecosystem-builder responsible for the formation of numerous seascapes within the semi-enclosed region of the western Arabian-Persian Gulf [P. i. radiata into a situation where evolutionary adaptation is necessary for survival [in-situ climbing above the sea floor using rocks, algal stipe and fronds Click here for additional data file.


To effectively forage in natural environments, organisms must adapt to changes in the quality and yield of food sources across multiple timescales. Individuals foraging in groups act based on both their private observations and the opinions of their neighbours. How do these information sources interact in changing environments? We address this problem in the context of honeybee colonies whose inhibitory social interactions promote adaptivity and consensus needed for effective foraging. Individual and social interactions within a mathematical model of collective decisions shape the nutrition yield of a group foraging from feeders with temporally switching quality. Social interactions improve foraging from a single feeder if temporal switching is fast or feeder quality is low. When the colony chooses from multiple feeders, the most beneficial form of social interaction is direct switching, whereby bees flip the opinion of nest-mates foraging at lower-yielding feeders. Model linearization shows that effective social interactions increase the fraction of the colony at the correct feeder (consensus) and the rate at which bees reach that feeder (adaptivity). Our mathematical framework allows us to compare a suite of social inhibition mechanisms, suggesting experimental protocols for revealing effective colony foraging strategies in dynamic environments. The e sources ,7, and u sources  or curb  sources . While w sources . Thus, b sources ,12.Honeybee colonies live in dynamic environments, in which the best adjacent nest or foraging sites can vary across time ,14. BeesTo study how social inhibition shapes foraging yields, we focus on a task in which the nectar quality of feeders is switched periodically. Related situations probably occur in nature due to the dynamics of competitor and predator prevalence, crowding by nest-mates, and weather fluctuations –25. PrecForaging colonies appear to be able to adapt to change. In prior studies ,16,20, cconsensus (steady-state fraction of bees at the high-yielding feeder) and adaptivity (the rate of switching from low- to high-yielding feeders). This provides a clear means of determining the impact of social interactions on a colony’s foraging efficacy.We quantify the contribution of these positive and negative feedback interactions within a mathematical model of a foraging colony. Our study focuses on four potential inhibitory social interactions—discriminate and indiscriminate stop signalling ,26, dire2.The mathematical model of bee colony foraging decisions assumes potential foragers may be uncommitted or committed to one of the possible feeders . UncommiA and B) systems, in which the fraction of the foragers committed to either feeder is described by a pair of nonlinear differential equations in the limit of large groups (see figure 1a for a schematic)αA,B(t) are time-dependent food qualities at feeders A, B (see figure 1b for examples); β min−1 is the rate bees recruit nest-mates to their feeder via waggle dancing; γ min−1 is the rate bees spontaneously abandon a feeder3; and uA,B ≤ 1 and 0 ≤ uA + uB ≤ 1, the commitment  and recruitment (β) provide positive feedback and the abandonment (γ) and inhibition , αX(t) − c. Integrating this product and scaling by time yields the effective RR.αA,B(t) and total foraging time Tf, colonies with more efficient foraging strategies J.We assume feeders are large enough to accommodate all the bees in the colony and hence we neglect the effects of crowding. Foraging efficacy is thus quantified by the group reward rate (RR), assuming net nutrition is proportional to the fraction of the colony at a feeder Before studying how social inhibition shapes a bee colony foraging in two-feeder environments, we analyse the single-feeder model, finding that commitment and negative feedback from either abandonment or inhibition are usually sufficient for the group to rapidly adapt to feeder quality switches.2.1.self-inhibition, by which a foraging bee stops another based on a detected change in food quality (a). Since transit from the hive to the feeder takes time, we incorporate a delay of τ min, so the fraction of foraging bees u evolves asα(t) is the food quality schedule of the feeder that switches at time intervals T (min) between α(t) = 0 and b), Inhibitory social interactions in a single-feeder model can only take the form of  quality a. Since  α(t)=α¯ ,20 (figuα(t) and abandonment γ) and interactions (recruitment β and inhibition ρ) [c,d). Equation (α(t) = 0) the non-foraging (β < γ). When food becomes available (c and appendix A(a)). This fraction consensus of the group [adaptivity.Colony’s adaptivity and consensus is shaped by both individual behaviour changes (commitment ition ρ) . PeriodiEquation  admits ohe group , and the2.1.1.T. Colonies with stronger rates of abandonment γ and self-inhibition ρ more quickly leave the feeder once there is no food .The performance of a colony’s interaction strategies strongly depends on the feeder quality y feeder d.γ, recruitment β and self-inhibition ρ, we compute the long-term RR of the colony, measuring the foraging yield over a single period (2T min) once the group equilibrates to its periodic switching behaviour (see appendix A(b))To quantify the effect of abandonment γ, recruitment β and stop signalling ρ) within our set of possible strategies (see appendix A(c)) that maximizes the RR J (e). Here, private information is sufficient for individual bees to commit to foraging ), and recruitment does not benefit the colony (β = 0). Reinforcing the majority opinion via recruitment is detrimental once the environment changes, as opposed to static environments [T) or low food quality (ρ) is needed to swap group commitment when the environment changes . This nonlinear mechanism increases the adaptivity of the group, but tempers the initial stage of consensus after the feeder is switched on (see appendix A(a) for details). On the other hand, when food is plentiful (high e), inhibition should be weak . In intermediate environments, the best strategies interpolate these extremes.For each feeder quality level, γ, β, ρ) e. Here, ronments ,31,32. IT) with small delays (τ → 0), we can linearly approximate the evolving foraging fraction (see appendix A(d))λon/off are the rates the group arrives/departs the feeder once food is switched on/off. Plugging equation ). Clearly, J increases with γ tends to decrease consensus, so the most robust foraging strategies cannot use abandonment that is too rapid (appendix A(a)).Linearizing solutions to the model equation  providesequation , we estiT) and profitability : (a) direct switching: bees foraging at the superior feeder directly switch the preference of opposing foragers to the better feeder; (b) indiscriminate stop signalling: when two foraging bees meet, one will stop foraging; (c) self-inhibition: when two bees foraging at the same feeder meet, one will stop foraging; and (d) discriminate stop signalling: when bees foraging at different feeders meet, one stops foraging. These interactions are visualized in figure 3a–d and their evolution equations are given in appendix B(a) .Figuree). The colony will thus mostly forage at the higher-yielding feeder. On the other hand, strong discriminant stop signalling can produce colonies with bistable consensus behaviours (figure 3f). Such hysteresis in stop-signalling populations was also identified in [We can divide these forms of social inhibition into two classes, based on the group foraging dynamics they produce: monostable or bistable consensus behaviours. The first three forms of social inhibition yield groups with monostable consensus behaviours (see appendix B(b) and electronic supplementary material of ), tendinfigure 3ified in . As a reified in ,35. Inhi2.2.2.T (min) and the better feeder quality A and B. As in the single-feeder case, we tune interactions of each strategy (a–d) to maximize RR over a discrete set of strategies (see appendix B(c) for details). Comparing each social inhibition strategy type’s RR in different environments (a), we find direct switching generally yields higher RRs than other strategies. Differences in the effectiveness of distinct strategies are most pronounced at intermediate environmental timescales T. As expected, RRs increase with the maximal feeder quality b,c).To determine the most robust forms of social inhibition for foraging in dynamic environments, we studied how the rate of reward, equation , dependestrategy a–d to maronments a, we finality α¯ b,c.Figua), as opposed to other strategies which interrupt foraging with an uncommitted stage (b–d) and rely on recruitment β to restart foraging. To study how interactions should be balanced to yield effective foraging, we examined how to optimally tune  across environments in the direct switching model  and abandonment γ (b), and strong direct switching (c) yield the highest RRs in slow (large T) and high-quality  and low quality . Changes in the optimal recruitment strength are less systematic, and there are stratified regions in which the best β can change significantly for small shifts in environmental parameters. Overall, a mixture of abandonment and direct switching is more effective in more difficult environments , since the colony can forage more efficiently by keeping some bees uncommitted, and not risking the cost of foraging at the lower-yielding feeder. Strong self-inhibition ρ keeps more bees from foraging. Overall, both direct switching and self-inhibition can perform similarly, as recruitment interactions can be strengthened in self-inhibiting colonies, so more bees return to foraging after such inhibitory encounters . Focusing specifically on these measures, we demonstrate both how they shape foraging efficiency and how they distinguish each social inhibition strategy.Each interaction mechanism differentially shapes both the fraction of bees that forage at the better feeder in the long-time limit (consensus T (see appendix B(d) and λ (rate this fraction is approached):J increases with consensus λ (a). Efficient colonies rapidly recruit a high fraction of the colony to the better feeder. Consensus and adaptivity are approximated in each model using linear stability (appendix B(b)). The impact of varying abandonment γ and social inhibition ρ on λ is consistent with our optimality analysis of the full nonlinear model : social inhibition generates more robust switching between feeders than abandonment. While strengthening abandonment adaptivity γ can increase λ, it decreases consensus b). Such consensus–adaptivity trade-offs do not occur in most models, as social inhibition ρ is strengthened , but the other three models  do not. Rather, consensus We leverage our approach developed for the single-feeder model, and consider linear approximations of equation  in the ltivity λ a. Efficiar model b,c: sociommitted b. Such cst model .Figure λ  or indiscriminate stop signalling (figure 6c) increase adaptivity but limit consensus β leads to stronger consensus λ to environmental changes. Bee colonies probably use a combination of social inhibition and abandonment mechanisms [Direct switching sustains both high levels of consensus chanisms , which w3.Foraging animals constantly encounter temporal and spatial changes in their food supply . The sucEfficient group decision-making combines individual private evidence accumulation and information sharing across the cohort . HoweverWe find that social recruitment can speed along initial foraging decisions, but it can limit adaptation to change. This is consistent with experimental studies that show a reduction in positive feedback can help collectives steer away from lower value decisions. For example, challenging environmental conditions  are best managed by honeybee colonies whose individuals do not wait for recruitment but rely on their own individual scouting . Ants enIn concert with a reduction in recruitment, we predict that honeybee colonies foraging in volatile environments will benefit from strengthening inhibitory mechanisms at the individual and group level. Bees enacting social inhibition dissuade their nest-mates from foraging at opposing feeders. We found the most efficient form of social inhibition is direct switching whereby bees flip the opinion of committed bees to their own opinion. So do honeybees use this mechanism in dynamic environments? Observations of bee colonies making nest site decisions show scouts directly switching their dance allegiance ,51, but Most work studying the effects of social inhibition on honeybee colony decisions has focused on swarms choosing a place to build a nest from sites whose qualities are fixed in time ,28. SociOur simple parametrized model, developed from previously validated house-hunting models ,28,29,32Previous computational modelling studies of honeybee collective decisions primarily focused on groups solving house-hunting problems in static environments ,29, emphThere are a number of possible extensions of our work here. For instance, one could consider separate populations of scouts and foraging recruits as in some previous modelling studies ,53. Our d,f), so the stochasticity in the finite-sized model could allow colonies to break free from less-profitable feeders, similar to noise-driven escapes of particles in double potential well models [Collective decision strategies and outcomes can depend on group size ,62, thoufigure 3l models . Fluctual models ,66, and l models ,68.