Gross efficiency (GE) and the index of pedal force effectiveness (IFE) are important factors that enhance cyclists’ performance; however, the effects of changing pedal force (gear ratio) and cadence on these indices while riding on a road racing bicycle are poorly investigated. This study aimed to examine the effect of changing gear ratio or cadence on GE and IFE using a road racing bicycle. Nine male cyclists completed graded submaximal cycling tests . The work rate of each stage was determined using two principles: changing gear ratio at a fixed cadence and changing cadence at a fixed gear ratio. We determined GE and IFE using respiratory variables and pedal reaction forces, respectively. Increasing the gear ratio improved GE, and was associated with the IFE. Although increasing the cadence slightly improved GE from the initial level, the increased values then mostly maintained. IFE was almost stable even when cadence increased. Moreover, no significant correlation was observed between the changes in GE and IFE accompanied by increasing cadence. Our data indicate that an increasing gear ratio, but not cadence, may affect GE and IFE while riding on a road racing bicycle. During cycling competitions, it is important to maximize gross efficiency (GE), defined as the mechanical work rate relative to the total energy expenditure of a cyclist, to ensure the highest endurance . PreviouCompetitive road cyclists must control their cycling speed according to the road situation using not only the pedaling gear ratio and but also the cadence to maintain safe and efficient cycling. Moreover, as the bike used in track cycling has a fixed pedaling gear ratio, cyclists can control the cycling speed by only changing their cadence. Although many previous studies have focused on the effect of altering the gear ratio on GE and IFE, these practical views suggest the importance of evaluating the effect of changing cadence on GE and IFE. Several previous studies have confirmed that GE decreases as cadence increases at a fixed work rate; thus, it is thought that increasing cadence impairs GE . HoweverAnother concern is that most previous studies used GE and IFE with an electrobrake ergometer, which does not reflect real road racing bicycles ,6,7,11. Therefore, this study aimed to investigate the effects of pedal force and cadence on GE and IFE using a road-racing bicycle. To test this, we examined two types of multistage graded submaximal cycling tests: changing pedal force (gear ratio) at a fixed cadence and changing cadence at a fixed gear ratio in well-trained college male cyclists.VO2max), 64.9 ± 5.9 mL·kg−1·min−1. They had at least 3 years training experience (5.4 ± 1.5 years) and were all well-trained cyclists who had been training at least six days per week. All participants were informed of the possible risks associated with the experimental procedures prior to providing written consent. The study protocol was approved by the Research Ethics Review Board of Juntendo University (28-106), and the study was performed in accordance with the Declaration of Helsinki. Nine male national- and regional-level collegiate endurance cyclists volunteered to participate in this study. Their physical characteristics (means ± standard deviation) were age, 20.4 ± 1.7 years; height, 170.8 ± 4.7 cm; body weight, 64.1 ± 5.4 kg; and maximal oxygen uptake  and a 3-roller bicycle trainer with a constant friction-loaded apparatus . All participants rode the same experimental bike, which had 170 mm crank, wheels , and tire , in order to ensure uniform aerodynamic drag of the wheel and rolling resistance of the tire. The seat position was adjusted according to participants’ preferences. The free roller was specified for bicycle training, and the apparatus consisted of three rollers . Although the magnetic resistance was adjustable, with six levels of strength, we attached it to the rear roller position of the free roller at the first level. The resistance level was determined to adjust the target work rates in subsequent multistage cycling tests.VO2), carbon dioxide production (VCO2), and respiratory exchange ratio (RER)) were continuously measured throughout the experiment.All participants visited the laboratory three times throughout the experiment. During the initial visit, a familiarization session consisting of adjusting the saddle position of the aforementioned competition racer bike to match the participants’ preferences and practicing riding on a free bicycle roller (AC-Magroller) while wearing a face mask to collect respiratory gas for the exercise tests was performed. At least 24 h following the familiarization session, the participants visited the laboratory twice (main experiments) on different days. In the main experiments, participants performed two types of graded submaximal cycling tests while wearing a continuous heart rate (HR) monitor and face mask: increasing the work rate by changing the gear ratio at a fixed cadence  and changing cadence at fixed gear ratio . The racer bike used gear ratio levels between 1.9 and 4.8 for this study; gear levels were changed by alternating the front and rear gears, which in turn altered the pedal force. In the GEAR trial, participants initially cycled with a gear ratio of 1.6 and a cadence of 90 rpm for 4 min on a free bicycle roller. Following a 1 min passive rest on the bicycle, cycling was restarted with a gear ratio of 0.5 higher than the initial gear ratio, but the cadence was maintained at 90 rpm. This 4 min cycling and 1 min rest session was repeated five times until the gear ratio was 3.5 . In the The HR was continuously measured during the test using a wireless HR sensor . The cadence and pedal force  were evaluated from the data obtained using a pedaling power meter . The pedaling power meter system has two streamlined strain gauge monitors that can detect the slightest tangent and radial force direction and force to the crank arms. In addition, the magnetic sensor detects the angle of rotation. This proprietary system detects the amount and direction of force at 12 different points during each pedaling stroke for each leg . The valVO2, VCO2, and RER were continuously monitored by collecting respiratory gas using an automatic gas analyzer . Prior to each experiment, the flow sensor was calibrated using a 2 L syringe, and the O2 and CO2 sensors were calibrated using calibration gas . The respiratory data averaged every 30 s were recorded, and the averaged values for the last 1 min during each stage of cycling were used for subsequent data analysis.VO2) + (1.195 × VCO2)] × (4.186/60) [We calculated the energy expenditure (EE) as follows: EE (J) = [(3.869 × .186/60) . GE was .186/60) .The Spearman’s correlation coefficient was used to examine the relationship between GE and IFE in each trial when the data were pooled. Briefly, in this study, these indices were assessed at five stages  for nine participants; thus, the total number of plotted data would be 45. This analysis was based on previous studies that demonstrated that improvements in GE were closely associated with IFE during incremental cycling tests .VO2, VCO2, RER, and GE. For a graphical representation of these data, we represented the mean and individual values as solid and dotted lines, respectively. Microsoft Office Excel 2016 and Prism v. 9.0  were used for data processing and statistical analyses, respectively. Statistical significance was set at p < 0.05.Normality of the datasets obtained from each experiment was tested using the Shapiro–Wilk test; however, if equal variance failed, Friedman nonparametric and pairwise post hoc tests (Bonferroni) were used. Indeed, as equal variance failed in IFE, we analyzed it as a nonparametric dataset. One-way analysis of variance with Bonferroni’s post hoc test was used to compare the time course changes in VO2, VCO2, and RER), GE and IFE, during the five-stage graded cycling test  and cadence using a road racing bicycle. As shown in Several previous studies have reported that the increase in work rate with increasing pedal force at fixed cadence improved GE and IFE during incremental cycling tests ,7,11,18.VO2, VCO2, and RER) were increased, as in the GEAR trial. Based on physics theory, the energy demand of the primary work muscle relative to the same absolute pedaling load per revolution is steady when the cadence is increased. In contrast, an increasing work rate (exercise intensity) might be accompanied by increasing external energy costs that do not involve effective pedal force, such as stabilization in riding posture (recruiting previously inactive muscles) and changing muscle activation [Many previous studies have reported that increasing cadence decreases GE at a fixed work rate and concluded that increasing cadence has a deleterious effect on GE . Howevertivation . Moreovetivation ,19. BaseFurthermore, a significant correlation between GE and IFE, which was observed in the GEAR trial, disappeared in the CADENCE trial. Cycling athletes and their coaches have often sought training strategies for enhancing IFE based on the concept that increasing IFE can also improve GE . In contThis study had some limitations. First, we set the cycling duration at each stage to 4 min. However, prolonged cycling, such as a competitive race , may induce central and peripheral fatigue, thus, GE and IFE would be impaired in this situation. Second, we used the IFE that is calculated from the radial force and the tangential force in the direction of pedal axle. We did not consider the lateral-medial force. The lateral-medial force may be lower than the other two forces, although we cannot deny that it influences the variables in this study. Third, we did not consider the involvement of anaerobic components in the calculation of GE. The exercise intensity at Stage 5 (RER > 1.0) might reach over the anaerobic threshold, over-estimating the GE. Fourth, because the experimental bike does not allow for a strictly variable work rate owing to the methodological limitations of the experimental bike, we set gear ratio and in each stage at absolutely same among participants in this study. Thus, the validity of the hypotheses derived from the results of this study should be confirmed by further studies that consider the relative exercise intensity of the participants. Moreover, we could not compare GE and IFE between the GEAR and CADENCE trials because the same work rate could not be set. Therefore, future studies should examine the various combination patterns with gear ratio and cadence at a fixed work rate to determine their optimal combination to maximize the cyclists’ performance. At last, since we recruited only well-trained competitive young male cyclists, it would be important, in future studies, to examine other cyclist populations, including world-class, junior, older adult, and female cyclists.Our results demonstrated that increasing the pedal force at a fixed cadence linearly improved GE and IFE, even on a road racing bicycle. In contrast, GE and IFE remained almost stable despite an increase in cadence at a fixed pedal force. Although further studies are needed, our data raise the possibility that an increasing pedal force, but not cadence, influences GE and IFE while riding on a road racing bicycle. Moreover, not all training methods to improve IFE may reflect an enhanced GE. We expect these results to provide novel insights for reconsidering pedaling strategies in cycling and training methods to enhance cyclists’ performance.


Acropora cervicornis genotypes that were originally collected from environmentally distinct regions of Florida’s Coral Reef and sampled after residing within Mote Marine Laboratory’s in situ nursery near Looe Key, FL (USA) for multiple years. By using 16S rRNA high-throughput sequencing, we described the microbial communities of 74 A. cervicornis genotypes originating from the Lower Florida Keys (n = 40 genotypes), the Middle Florida Keys (n = 15 genotypes), and the Upper Florida Keys (n = 19 genotypes). Our findings demonstrated that the bacterial communities of A. cervicornis originating from the Lower Keys were significantly different from the bacterial communities of those originating from the Upper and Middle Keys even after these corals were held within the same common garden nursery for an average of 3.4 years. However, the bacterial communities of corals originating in the Upper Keys were not significantly different from those in the Middle Keys. The majority of the genotypes, regardless of collection region, were dominated by Alphaproteobacteria, namely an obligate intracellular parasite of the genus Ca. Aquarickettsia. Genotypes from the Upper and Middle Keys also had high relative abundances of Spirochaeta bacteria. Several genotypes originating from both the Lower and Upper Keys had lower abundances of Aquarickettsia, resulting in significantly higher species richness and diversity. Low abundance of Aquarickettsia has been previously identified as a signature of disease resistance. While the low-Aquarickettsia corals from both the Upper and Lower Keys had high abundances of an unclassified Proteobacteria, the genotypes in the Upper Keys were also dominated by Spirochaeta. The results of this study suggest that the abundance of Aquarickettsia and Spirochaeta may play an important role in distinguishing bacterial communities among A. cervicornis populations and compositional differences of these bacterial communities may be driven by regional processes that are influenced by both the environmental history and genetic relatedness of the host. Additionally, the high microbial diversity of low-Aquarickettsia genotypes may provide resilience to their hosts, and these genotypes may be a potential resource for restoration practices and management.Effective coral restoration must include comprehensive investigations of the targeted coral community that consider all aspects of the coral holobiont—the coral host, symbiotic algae, and microbiome. For example, the richness and composition of microorganisms associated with corals may be indicative of the corals’ health status and thus help guide restoration activities. Potential differences in microbiomes of restoration corals due to differences in host genetics, environmental condition, or geographic location, may then influence outplant success. The objective of the present study was to characterize and compare the microbiomes of apparently healthy  Acropora cervicornis have exhibited marked declines since 1980 due to a multitude of stressors including infectious disease, poor water quality, high sea surface temperatures, and overfishing , with high variation in genetic diversity among regions  of  regions . Signifi species . Specifi species  and dise species . The com species , disease species , and gro species  in Acropent cues . Certainent cues . The divent cues .Coral-associated microbes tend to vary by host species , and witFlorida’s Coral Reef has distinct spatial regions because of varying seawater circulation patterns due to the underlying geology that contribute to regional differences in environmental parameters and hardbottom communities . While tA. cervicornis identified as susceptible or resistant to diseases  for multiple years. Microbiomes of genotypes collected from the Upper, Middle, and Lower Keys were described using 16S rRNA high-throughput sequencing to characterize and compare the microbiomes of apparently healthy A. cervicornis residing within the coral nursery. We found that while the majority of A. cervicornis genotypes from all collection regions harbored microbiomes dominated by putative parasites of the genus Aquarickettsia, a subset of genotypes possessed a significantly different microbiome characterized by the dominance of either an unclassified sequence variant or the genus Spirochaeta. Additionally, genotypes from the Upper and Lower Keys retained a microbiome signature that was specific to the geographic region of origination, while genotypes from the Middle Keys were distinct from those originating from the Lower Keys, but not those from the Upper Keys. Our results provide key information on the diversity and potential geographical influences on the microbiomes of an important restoration species.Here, we examined differences in the microbial community composition of Acropora cervicornis genotypes (n = 74) were collected from Mote Marine Laboratory’s in-situ nursery near Looe Key, FL (USA) in the Lower Keys and sampled for microbiome analysis. The Florida Keys National Marine Sanctuary authorized the use of nursery-grown corals under permit FKNMS-2015-163. These corals were curated for Mote’s coral restoration efforts throughout the Florida Keys and previously genotyped using both microsatellites  to Mote’s Elizabeth Moore International Center for Coral Reef Research and Restoration . Once back at the lab, the seawater in each Whirl-Pak was discarded and the fragments were flash-frozen in a dewar containing liquid nitrogen for one minute. The frozen samples were then immediately stored at −80 °C until further processing.In November-December 2019, tellites  and the tellites . Of the tellites . Genotyptellites , whereasDNA was isolated from the frozen coral fragments using DNeasy PowerSoil Kits  with modifications to the manufacturer’s protocol . Using shttp://www.mrdnalab.com, Shallowater, TX, USA) for barcoding, amplification, and sequencing. Amplification of the 16S rRNA gene variable region (V4) was conducted using primers 515F  for congruence with current identifications in the literature (phyloseq (v1.30.0)  . A totalphyloseq , samplesNon-metric Multidimensional Scaling analyses (nMDS) using the Bray-Curtis dissimilarity distances were performed to determine similarities in bacterial communities among samples due to collection region. Upon conducting the nMDS analysis, we visually identified 10 outliers. We then used the ‘OutlierDetection’ function  dominated the microbiomes of most genotypes, accounting for approximately 90% or more of the relative abundances . According to NCBI’s basic local alignment tool (BLAST), this ASV is most similar to an uncultured bacterium clone plBB03 sampled from A. palmata in Puerto Rico . The outlier genotypes in the Upper Keys were dominated by bacteria in the genera of both the unclassified proteobacteria and Spirochaeta. The relative abundance of this unclassified ASV was 0.78 ± 0.96% in non-outlier samples, and 29.3 ± 20.5% in outlier samples.Post-filtration, as described in the methods above, the dataset consisted of 1,195 ASVs, with a mean read depth per ASV of 1,643. The 16S rRNA sequences are available under BioProject ID is taxon . The aveundances  and S3. enotypes . The aveAquarickettsia sp., while the microbiomes of the outlier genotypes were composed of many highly relatively abundant taxa  demonstrated similarities among fragments by collection region  and Lower (P = 0.003) Keys; however, the microbial communities of the Upper and Middle Keys were not significantly different  were significantly different as shown by the 95% confidence ellipses in the nMDS analysis . Alpha diversity did not significantly differ between the Upper and Lower Keys low-Aquarickettsia genotypes . ASV 1, an Aquarickettsia sp., was highly relatively abundant in genotypes from all initial collection regions; however, it was more relatively abundant and less variable in genotypes from the Lower Keys (Aquarickettsia genotypes from the Lower Keys include an ASV from the Helicobacteraceae family (ASV 10), an unclassified Proteobacteria ASV (ASV 12), and a Cetobacterium sp. (ASV 17). According to NCBI BLAST, Unclassified ASV 12 was most similar to an uncultured bacterium clone p1BB03 found in A. palmata from Puerto Rico , and also closely matched to an uncultured bacterium clone Acer_J17 in A. cervicornis from Bocas del Toro, Panama , an Enterococcus sp. (ASV 9), unclassified ASV 21, a Cloacibacterium sp. (ASV 33), and ASV 35 in the family Microbacteriaceae were relatively positively enriched in the Upper Keys low-Aquarickettsia genotypes. Unclassified ASV 21 was most similar to an uncultured marine bacterium clone S82_61c03 found in the skeleton of Cladocora caespitosa in the Mediterranean Sea . We found that a single ASV from this species was dominant in our samples; there was a 100% match to the published 16S rRNA sequence for the type species, strain acerv44, for which the complete genome is available  scale.Coral host-microbe interactions are influenced by environmental factors that vary over both space and time , even ovferences  that likdistinct , thus it factors , and mic factors  have conin situ nursery in the Lower Keys for an average of 3.4 years, their microbiomes still held geographic markers, suggesting the potential for retaining microbiome signatures over time. Host identity is known to have greater control on microbiome composition than temporal differences  for multiple years. We found significant differences in bacterial diversity of the genotypes driven by the initial collection region, even after the corals had been kept under common garden conditions for multiple years. These differences in microbial communities due to geographical origin most likely reflect the genetic relatedness and environmental history of the genotypes sampled. Our results suggest that the abundance of two key bacterial taxa, Ca. Aquarickettsia and Spirochaeta, may play an important role in distinguishing bacterial communities among A. cervicornis populations. Most genotypes were dominated by Aquarickettsia sp., a presumptive bacterial parasite of A. cervicornis that is associated with disease susceptibility (Aquarickettsia sp. These low-Aquarickettsia genotypes may be a potential resource for restoration initiatives since populations that are more likely to succeed and survive in stressful environments should be thoughtfully targeted for restoration efforts.The present study characterized the microbiomes of apparently healthy tibility . Further10.7717/peerj.13574/supp-1Supplemental Information 1Click here for additional data file.


Dupilumab, an anti-IL-4α receptor antibody, is a new treatment for severe or refractory asthma. However, real-world evidence on the efficacy of dupilumab in patients with mild to moderate bronchial asthma is lacking.We retrospectively evaluated the effects of dupilumab in 62 patients who received dupilumab for eosinophilic sinusitis comorbid with asthma at a single centre in Japan. Type 2 inflammatory markers, ACT, respiratory function tests, and forced oscillation technique (FOT) were analysed before, three months after, and one year after dupilumab administration, mainly in patients with mild to moderate asthma.FEV1, %FEV1, %FVC, treatment steps for asthma and ACT improved significantly after three months of dupilumab treatment. FeNO was markedly decreased, whereas IgE and eosinophil counts showed no significant changes. Pre- and post-treatment respiratory resistance (Rrs) and respiratory reactance (Xrs) correlated significantly with FEV1. Improvement in %FEV1 was associated with higher FeNO and higher serum IgE before dupilumab treatment.Dupilumab treatment for sinusitis may improve respiratory functions, asthma symptoms, and asthma treatment reduction, even if the associated bronchial asthma is not severe. In the last few years, the treatment of refractory asthma has made dramatic progress, especially with the development of biological therapies targeting molecular effectors and signalling associated with type 2 airway inflammation –5. DupilThe upper airway, such as the nasal cavity and paranasal sinuses, and the lower airway, such as the bronchi and lungs, are anatomically connected and recognized as the "united airway" . The uppThe forced oscillation technique (FOT) is a non-invasive, effort-independent method used for respiratory function testing in recent years and is expected to be beneficial, especially in children and the elderly, as it can be performed during tidal breathing ,19. HoweThe present study aims to retrospectively evaluate the real-life effects of dupilumab on comorbid mild to moderate asthma in patients treated with dupilumab for CRSwNP in terms of the changes in type-2 inflammatory markers, ACT, respiratory function tests, and forced oscillation technique (FOT).This research is a retrospective observational study conducted at a single centre (Department of Otolaryngology and Respiratory Medicine in Matsuwaki Clinic Shinagawa). Sixty-two consecutive adult patients treated with dupilumab for eosinophilic sinusitis with nasal polyps complicated with asthma from May 2020 to July 2021 were retrospectively analysed. All asthma patients were diagnosed by respiratory physicians based on the Japanese guidelines . Blood eFractional exhaled nitric oxide concentration (FeNO) was measured using electrochemical sensor NO analysers  with a 50 mL/s flow rate, as recommended by the American Thoracic Society / European Respiratory Society, using a spirometer  that assessed forced expiratory volume in 1 s (FEV1), FEV1/FVC, and  ratio). Respiratory impedance was measured by a broadband FOT using MostGraph-02 . An impulse oscillatory signal generated by a loudspeaker was applied to the airway through a mouthpiece for about 30 s during tidal breathing.In this study, we measured and analysed respiratory resistance (Rrs) at 5 Hz (R5) and 20 Hz (R20), and reactance (Xrs) at 5 Hz (X5), resonant frequency (Fres).Data were analysed using Prism 8 . A p-value of < 0.05 was taken as the threshold for statistical significance. All values are expressed as the means ± standard deviation (SD). Data for individual variables before and three months after starting dupilumab were analysed using the Paired t-test. The factors associated with patient characteristics were analysed using the unpaired t-test and the chi-square test. The Pearson correlation coefficient was used to investigate the correlation between the FEV1 and FOT values.During the observation period, 62 patients underwent respiratory, otolaryngological, and blood examinations after three months of treatment with dupilumab, and 23 patients underwent tests after one year of treatment. During the observation period, all patients who received dupilumab had a past history of asthma or were currently receiving asthma treatment.All the patients had not experienced any asthma attacks during the past three months and were well controlled by the current asthma treatment (mean ACT score is 22–23) or treatment-free. Of the 62 patients who received dupilumab, 58 (93.5%) received it as their first antibody therapy for asthma, and the remaining 4 (6.5%) had received antibody therapy  immediately before dupilumab. Most patients in this study received around 5 mg of oral corticosteroids (prednisone) for poorly-controlled sinusitis. Serum IgE and FeNO were mildly elevated than normal in all patients, but there were no significant differences between severe and mild-moderate asthma. The mean serum eosinophil counts were within the normal range regardless of severity. Although there was a mild increase in each of the spirometry indices, as well as the respiratory resistance and reactance in FOT, there were no significant differences in these indices between mild to moderate and severe cases, indicating that asthma treatment was well controlled before the initiation of dupilumab in the present study. The number of patients with mild to moderate asthma was 50, accounting for 80% of all patients, and the efficacy of dupilumab for severe cases has already been established (ref). Accordingly, we performed a series of evaluations mainly focusing on mild to moderate cases . Nonetheless, three months of dupilumab treatment significantly improved ACT score, which was maintained after one year. In addition, the asthma treatment steps based on the GINA strategy were also significantly downgraded at three months and after one year Fig. B. In terWith regard to respiratory functions, forced expiratory volume in 1 s (FEV1), the percentage of predicted forced expiratory volume in 1 s (%FEV1), the percentage of predicted forced vital capacity (%FVC), and the FEV1/FVC ratio (FEV1%) were measured by spirometry. The pre-dupilumab values of FEV1, %FEV1, and %FVC were significantly improved at three months, and the improvement was maintained for one year, while FEV1% did not significantly change by dupilumab treatment Fig. A.Fig. 2AAs for the respiratory impedance, we measured R5 and R20 as respiratory resistance (Rrs) and X5 and Fres as respiratory reactance (Xrs). Respiratory resistance  tended to improve, but the change was not without significance at three months  before and three months after dupilumab treatment. FEV1 and respiratory impedance correlated significantly before and three months after dupilumab treatment for mild to moderate asthma, suggesting that FOT shows a similar trend to spirometry with respect to obstructive disorders Fig. A. HoweveIn this study, most patients had mild to moderate asthma, and the efficacy of dupilumab is mainly established in severe asthma. Although all participants, including those with severe symptoms, were clinically controlled at dupilumab administration, we compared the improvement in respiratory functions, ACT, treatment step, and FeNO in response to three months of dupilumab treatment between mild to moderate and severe asthma. The spirometry indices  showed a trend of improvement in both groups, and no significant difference was observed Fig. . With reTo determine the patient characteristics contributing to improved respiratory functions with dupilumab in mild to moderate asthma, we compared nine factors associated with asthma and CRSwNP between the group of patients with %FEV improvement below 3.8 L and that with %FEV improvement above 3.8 L. In this study, the reference value was set at 3.8 L, which is the mean improvement in %FEV1 with 3 months of dupilumab treatment for all patients. Among the nine factors, pre-dose FeNO and serum IgE were significantly different between the two groups, suggesting that the higher values of FeNO and IgE can be associated with more improved respiratory functions by dupilumab treatment Table . In addiDupilumab is approved for uncontrolled CRSwNP, severe or uncontrolled bronchial asthma, and atopic dermatitis and has the broadest range of use among antibody products against type 2 inflammation. Although unintentional administration of dupilumab to well-controlled asthma may happen during CRS treatment, respiratory function tests for asthma control are rarely monitored in the clinical practice of otolaryngology. The LIBERTY ASTHMA QUEST trial, a phase-3 trial that showed significant improvement in respiratory functions in bronchial asthma patients with dupilumab, included patients with moderate as well as severe asthma , 25. SimThe present study showed that dupilumab significantly improved respiratory functions  in patients with clinically-controlled mild to moderate asthma associated with CRSwNP at three months of the administration. In addition, asthma treatment steps were significantly reduced after the initiation of dupilumab  observed in both allergic and non-allergic asthma patients . HoweverThe forced oscillation technique (FOT) is a respiratory function test that measures respiratory system impedance (Zrs) by applying pressure oscillation of air through the mouth during resting ventilation. The resistance at 5 Hz (R5) reflects the total airway resistance, while that at 20 Hz (R20) reflects the central airway resistance. Respiratory reactance (Xrs) is considered a lung compliance index but can be explained as an index of dynamic obstruction of the peripheral airways, reflecting increased air trapping . Only a There are several limitations to this study. Because of the retrospective observational study, the present study could not be compared with the placebo control. In addition, although this study evaluated 50 patients primarily for the treatment effectiveness over three months, there were still 23 patients who were evaluated for testing after one year. The TRAVERSE study, which followed patients with asthma who had completed a clinical trial of dupilumab for an extended period, showed that the safety and efficacy of dupilumab in patients with severe asthma were maintained even after extending the treatment to 148 weeks . In the This study is unique in that it examined the response of patients with well-controlled asthma to treatment, unlike previous studies that examined the effects of treatment with dupilumab in severe asthma patients. This study is also unique in that all patients had sinusitis that was poorly controlled after sinusitis surgery alone, which is not possible in the usual practice of respiratory medicine alone. In summary, dupilumab therapy not only has a marked effect on CRSwNP but may also improve respiratory functions and symptoms and reduce the dose of asthma treatment, even if comorbid asthma is not severe and is clinically controlled.


Breastfeeding is widely supported in clinical and home practices, and it is known that different forms of infant feeding differ in hormone content. Thyroid hormones have essential physiological roles. In our study, we examined thyroid-stimulating hormone (TSH), thyroxine, and albumin levels in breast milk produced for term (n = 16) or preterm (n = 15) infants throughout the first 6 months of lactation. Moreover, we analyzed these components in donor human milk and in three different infant formulas. Term and preterm breast milk samples were collected monthly. The two groups had similar levels of TSH (18.4 ± 1.4 vs. 24.7 ± 2.9 nU/L), but term milk contained higher amounts of thyroxine  during the examination period. The albumin level was significantly higher in preterm breast milk than in term breast milk (328.6 ± 17.1 vs. 264.2 ± 6.8 mg/L). In preterm breast milk we detected downward trends in the levels of TSH (−30.2%) and thyroxine (−29.2%) in the 3rd through 6th month compared to the first 2 months of lactation. Microbiological safety of donor milk was ensured by Holder pasteurization (HoP). From the Breast Milk Collection Center of Pécs, Hungary, we enrolled 44 donor mothers into the study. HoP decreased TSH (−73.8%), thyroxine (−22.4%), and albumin (−20.9%) concentrations. Infant formulas used by the Neonatal Intensive Care Unit of the University of Pécs were found to not contain the investigated hormones, but their albumin levels were similar to the breast milk samples. The present study shows the lack of thyroid hormones in infant formulas compared to human milk and raises the question of whether formula-fed infants should be supplemented with thyroid hormones. Transplacental hormonal exposure impacts intrauterine development. Premature birth disrupts the placentofetal connection. Therefore, for a preterm infant, after birth breast milk is the absolute best source of maternal hormones. Breast milk is the recommended form of infant feeding, supported by numerous organizations worldwide. Adequate nutrition of preterm infants remains a challenge with regard to achieving a sufficient amount of bioactive substances, assuring linear growth and neurocognitive development ,2. DurinDuring fetal life, the thyroid gland secretes thyroxine (T4) and triiodothyronine into the circulation from about 12 weeks’ gestation, and its levels increase until term. In limited amounts, the maternal T4 crosses the placenta throughout gestation, and this plays a critical role in the development of the central nervous system in the first trimester, as demonstrated by the fetal neurological impairment described in fetomaternal Pit-1 deficiency  and seriIn the present study, we collected breast milk samples from mothers who delivered preterm or term infants, and measured the thyroxine, TSH, and albumin levels in the samples. We measured these substances in samples of donor milk and also attempted to detect the presence of the hormones and albumin in infant formula. The purpose of our investigation was to provide more information about the hormonal content of own mother’s milk, donor milk, and commercial infant formula.The present study was conducted after the approval of the Regional and Local Research Ethics Committee of the University of Pécs, Pécs, Hungary (PTE KK 7072-2018). Waivers for participant consent were obtained.The subjects were mothers who had given birth to term (n = 16) or preterm (n = 15) infants at the Department of Obstetrics and Gynecology, University of Pécs. Breast milk sample collection started at the 4th postpartum week and continued monthly until six months after birth. Participants pumped their entire breast expression into sterile polypropylene bottles between 1 p.m. and 3 p.m., and 5 mL from the bottle was poured separately and then stored in a sterile polypropylene tube until analysis.g for 15 min. Then the skim milk was transferred to polypropylene tubes for measurements based on previous laboratory preparation methods [As a part of our study, 44 mothers—all of them registered and approved donors—were involved from the Breast Milk Collection Center of the Unified Health Institution at Pécs, Hungary. Based on the breast milk collection centers’ protocol, mothers donated freshly pumped milk. We collected breast milk samples at six random occasions in the milk bank. We took samples individually and then the milk samples were pooled and Holder pasteurized (30 min at 62.5 °C) at the laboratory of the Unified Health Institution . After the procedure, we took three samples from the pooled and Holder pasteurized donor milk and stored them at −80 °C until laboratory measurements. During the analysis, first, the samples were sonicated in order to disrupt the milk fat globule membranes and, as previously described , centrif methods .Three different infant formulas were tested in our analysis to obtain information about the hormone composition of these fluids. Infant formulas were prepared and applied regularly in the Department of Neonatology, University of Pécs and were tested at five different time points: Nutricia Milumil Pepti Pronutra , Beba Optipro Hypoallergenic (HA) Start , and Beba Optipro HA Pre . Samples were taken at the Neonatal Intensive Care Unit ten times in the morning between 8 and 9 a.m. and were stored similarly to the breast milk samples, in sterile polypropylene tubes at −80 °C. An infant has three feeding options. The first is own mother’s milk; when it is not available, donor milk is the recommended form of infant feeding; the third possibility is infant formula. The TSH assay employs monoclonal antibodies  specifically directed against human TSH. For TSH detection, first, a 50 μL breast milk sample was mixed with a biotinylated monoclonal TSH-specific antibody and a monoclonal TSH-specific antibody labeled with ruthenium complex. Streptavidin-coated microparticles were added. The TSH detection range was 0.0005–100 mU/L. No crossreactivities were found with LH, FSH, human growth hormone, or human chorionic gonadotropin.To measure thyroxine, a 15 μL breast milk sample and a thyroxine-specific antibody labeled with a ruthenium complex were mixed, and then biotinylated thyroxine and streptavidin-coated microparticles were added to the mixture. The thyroxine detection range was 0.3–100 pmol/L. The following cross-reactivities were found with L-triiodothyronine: 0.005%, D-triiodothyronine: 0.001%, reverse triiodothyronine: 0.003%, 3,3′,5-triiodothyroacetic acid: 0.0002%, and 3,3′,5,5′-tetraiodothyroacetic acid: 0.001%.ARCHITECT i1000 system was applied for the measurements, and we followed the manufacturer’s instructions throughout. The fully automatized Cobas e 411 analyzer system performed all the measurements . After the previously described automatized preparation steps, the induced chemiluminescent emission was detected by a photomultiplier. Results were given after a two-point calibration curve was generated. Parallel with the breast milk samples, quality controls were tested.Albumin was detected with an immunoturbidimetric assay . In this method, antialbumin antibodies react with the sample’s antigen and form complexes. After agglutination, the samples were measured turbidimetrically in a fully automatized way, and quality controls were also applied.t-test or ANOVA was used for analysis. With post-hoc Dunnett test, repeated measures one-way ANOVA test was used to compare the effect of Holder pasteurization on the concentration of the chosen hormones. Differences were determined statistically significant when p values were <0.05. The study was powered to detect moderate effect sizes (Cohen’s d = 0.6). Results are presented as mean ± SEM or median values with interquartile ranges.GraphPad  was used for statistical analysis, and Shapiro-Wilks tests were performed to test the data normality. Paired All mothers involved in our study conceived spontaneously. Differences between preterm and term groups based on maternal age, body mass index (BMI), or infant sex were not found . MothersThroughout the first 2 months of lactation, breast milk contained significantly higher amounts of thyroxine and TSH compared to the period between the third and sixth months. The albumin level of breast milk produced for preterm infants was similar across the first six months of lactation .The concentrations of TSH, thyroxine, and albumin in breast milk produced for term infants did not change with time since birth. Therefore, we presented the average concentration values pooled from the first six month of lactation. While comparing the average thyroxine and TSH levels of preterm and term breast milk, thyroxine was present at significantly higher levels in term milk. In contrast, albumin was present at significantly higher levels in preterm milk, whereas the TSH level was the same in preterm and term breast milk .TSH, thyroxine, and albumin levels were significantly decreased by Holder pasteurization. The TSH level was decreased with 73.8%, the thyroxine concentration with 22.4%, and the albumin level with an average 20.9% concentration .Three of the infant formulas in use at the Department of Neonatology, University of Pécs are the Milumil Pepti Pronutra, the Beba Optipro HA Start, and the Beba Optipro HA Pre. Thyroxine and TSH were not found above the lower limits of detection in any of these formulas. Analyzing pooled results of the breast milk samples collected during the first 6 months of lactation, formulas had an average 315.58 ± 19.11 mg/L albumin level, which was similar to the levels measured in term and preterm breast milk.In the present study, we detected the presence of TSH, thyroxine, and albumin during the first six months of lactation in breast milk produced for term or preterm infants. The same components were analyzed in donor breast milk, and in commercial infant formulas as well, to provide previously unavailable information about the TSH, thyroxine, and albumin concentrations of three different feeding options of an infant.Hormones and nutrients, including growth factors, immunoglobulins, chemokines, cytokines, oligosaccharides, vitamins, and immune cells have been demonstrated to transfer from the mother to the infant via breast milk ,17. The Regarding our knowledge about the presence of TSH in infant feeding sources, we concluded that TSH is measurable during the first 6 months of lactation in both preterm and term breast milk, but it is unavailable in infant formulas. The oral bioavailability of thyroxine is known, but it is not known whether TSH is bioavailable if provided orally. The presence of TSH receptors in the gastrointestinal tract raises the possibility of local effects .In maternal milk produced for preterm infants, composition changes were detected. In the first two months of lactation, mother’s milk contains higher thyroxine and TSH compared to the time period between the 3rd and 6th month of breastfeeding. We hypothesize that after the suddenly interrupted pregnancy, maternal thyroid hormones may decrease slowly, with a drop present in the composition of breast milk. Sack et al. described a thyroxine peak in the 5th postpartum week at 4.3 μg/100 mL and concluded that the mother secretes 40–50 μg of thyroxine per day in her milk, covering the thyroxine need for an infant to avoid hypothyroidism .Severe hypothyroidism that is often diagnosed in neonates proposes the possibility of the transplacental supply of thyroid hormones. Biological half-life of serum T4 was 3.6 days (between 2.7 and 5.3 days). In infants with thyroid agenesis and in neonates with a total organification defect, the serum T4 concentration and its disappearance kinetics were the same, suggesting that thyroxin also had a maternal origin in these infants. The authors concluded that in infants diagnosed with severe congenital hypothyroidism, a substantial amount of thyroxine is transferred from the mother to the fetus .Gastric and intestinal digestion of proteins and other components are attenuated during the first few postnatal months. The infant’s gastrointestinal tract has immature barrier functions and the cell–cell connections. The amount of iodine through deiodination of T4 is insignificant in comparison with the inorganic iodine concentration of breast milk produced for preterm infants; the median concentration value was 105 μg/L. Iodine concentration showed a relatively large variation (38–293 μg/L) in breast milk, which might be caused by the fluctuating maternal iodine intake . The gasIn extremely preterm infants, transient low levels of plasma T4 is often detected and observed, described as the transient hypothyroxinaemia of prematurity. Van Wassenaer et al. investigated whether maternal milk is a relevant resource of thyroid hormones for very preterm infants and can relieve transient hypothyroxinemia . Both thThe reason for postnatal hypothyroxinemia is multifactorial, including the loss of the transplacental maternal T4 contribution, immaturity of the hypothalamic–pituitary axis, the responsiveness of the thyroid gland to TSH, and immaturity of peripheral tissue deiodination. In the first few weeks after birth, iodine balance is negative in very low birthweight babies, suggesting an inability to augment thyroidal iodine uptake and increase T4 secretion . PreviouThe effect of thyroxine supplementation was investigated previously in the studies of Amato et al.  and van When own mother’s milk is not available in neonatal care, the recommended form of infant feeding is donor milk. Before feeding it to neonates, donor milk is pasteurized to ensure microbiological safety in human milk banks. The impact of this procedure on the concentration of various hormones is different. Pasteurization is necessary to assure the safety of donor milk, although it may affect milk quality by the reduction of bioactive components ,6,15,34.Albumin is a circulatory protein, with numerous vital physiological functions, such as regulating metabolic and vascular functions, maintaining microvascular integrity and oncotic pressure, providing antioxidant activities, binding ligands for different substances, and having anticoagulant effects. Its presence in human milk was known , but no Our study had limitations. Sonication disrupts milk fat globules and allows proteins to enter the aqueous phase of the milk samples, but the removal of this fat layer during analysis may have led to an underestimation of hormone levels. Hormones were not measured in the blood of the infants, thus the impact of human milk content remains unknown. We investigated one method, the HoP; however, other techniques used to ensure microbiological safety of donor milk may differently influence the concentration of these breast milk components.Our study highlights that different forms of infant feeding provide different amounts of thyroxine and TSH to the infant. Intrauterine development is influenced by numerous unreplaceable factors. After birth, bioactive compounds, such as hormones, impact postnatal development via absorption from the gastrointestinal tract. Preterm infants are born deficient in these transplacental hormones because their intrauterine development is interrupted early. Our results showed for the first time the long-term presence of TSH, thyroxine, and albumin in preterm and term breast milk during the first 6 months of lactation. The present study improves our knowledge of the composition of infant formulas by demonstrating the absence of measurable amounts of thyroxine and TSH in the formulas. Term infants receive higher thyroxine concentrations in their mothers’ milk than do preterm infants. It is known that HoP influences the composition of human milk. Our study showed that the HoP decreased the thyroxine content of breast milk. Oral thyroxine replacement is an accepted medical treatment. Its use should be investigated for exclusively formula-fed infants.


TFF1) is considered to be able to inhibit the formation of kidney stone. However, genetic variants in TFF1 and corresponding function in kidney stone development are still not well studied. In this study, the discovery set including 230 cases and 250 controls was used to analyze the association between seven tagSNPs of TFF1 gene and the nephrolithiasis risk. Further evaluation was confirmed by the validation set comprising 307 cases and 461 controls. The consequences of the two-stage case–control study indicated that individuals with the rs3761376 A allele have significantly increased nephrolithiasis risk than those with the GG genotypes . Moreover, we also carried out a stratified analysis and found the increased nephrolithiasis risks at A allele among males, overweight individuals, no hypertensive individuals, nondiabetic individuals, smokers, and drinkers. In the following functional experiments, the notably lower expression of TFF1 was exhibited by the vectors carrying A allele compared with those carrying G allele in both luciferase (P = 0.022) and expression vectors (P = 0.041). In addition to tissue detection, we confirmed a significant inverse association of rs3761376 G > A and TFF1 gene expression (P < 0.001). These results suggest that TFF1 rs3761376 may serve as a potential biomarker to predict the risk of nephrolithiasis.Trefoil Factor 1 (The online version contains supplementary material available at 10.1186/s12894-022-01081-w. Kidney stone disease  remains a public health problem around the world, of which the lifetime prevalence is approximately 10% in men and 5% in women . Of all So far, exact pathogenesis of kidney stone formation is still poorly understood. A long-standing hypothesis suggested that stones originally formed in the lumen of the renal tubules . The calTFF1, TFF2, and TFF3  were under-represented in the nephrolithiasis patients compared with the controls. Hence, individuals with the rs3761376 G allele genotypes [GG and AG] had a significantly lower nephrolithiasis risk compared to those with the AA genotype . Finally, the results of the combined set also explicitly manifested the increased nephrolithiasis risk effected by A allele of rs3761376 in additive, dominant and recessive genetic model.Genotype frequencies of P = 0.013), overweight individuals , not hypertensive individuals , nondiabetic individuals , smokers  and drinkers . For the test of heterogeneity, most factors had low heterogeneity, instead the heterogeneity of drinking status was high (Pheterogeneity = 0.005). An obvious increase in nephrolithiasis risk existed with the interaction between polymorphisms of TFF1 and drinking status (Pinteraction = 0.037).We conducted the stratified analyses by age, gender, BMI, hypertension, diabetes, smoking and drinking status, basing on the dominant genetic model of the rs3761376 in the combined set. As showed in Table TFF1 gene expression, we first constructed the sequence including the promoter of TFF1 with either rs3761376 allele (G or A) into different luciferase reporter vectors, and then transfected the vectors into HEK-293 cells. As presented in Fig. P = 0.022). Next, the synthesized fragment of TFF1 with either G or A rs3761376 allele was inserted into expression plasmid. In Fig. P = 0.041).To evaluate the genetic effect on TFF1 with known genotypes in 52 kidney tissues of nephrolithiasis patients. In Fig. TFF1 expression (P < 0.001), which was in accordance with the results of the cell experiments.For further study of the rs3761376 effects, we evaluated the expression levels of TFF1 may play a potential pathophysiological role in nephrolithiasis  were observed among males, overweight individuals, not hypertensive individuals, nondiabetic individuals, smokers and drinkers.As a novel potent CaOx crystal growth inhibitor, ithiasis , 26. Numithiasis –35. To dTFF1 on the risk of nephrolithiasis may be regulated by specific demographic factors and environmental exposures. A possible explanation is that some environmental hazards, such as smoking and drinking, increase the risk of kidney stones, thereby enhancing the effect of rs3761376 variants. This also provides evidence that the pathogenesis is a complex process involving both genetic and environmental factors [In China, it is a fact that the proportion of drinking and smoking is higher in male than in female; in addition, the prevalence of kidney stones in male is higher than in female, which may lead to different risks among the gender . Other rTFF1 and polymorphisms in promoter region were reported possessing potential functions to affect the transcription activity by altering the bindings of transcription factors [TFF1 by changing the bindings of transcription factors. Moreover, by evaluating the expression levels of TFF1 with known genotypes in 52 kidney tissues of nephrolithiasis patients, a significantly lower TFF1 expression was found along with the increase of the rs3761376 A allele. Combined the results of cell and tissue experiments, it is plausible that TFF1 rs3761376 may have the ability to alter the bindings of transcription factors and modulate the TFF1 expression. Studies have shown that the C-terminus of TFF1 binds to Ca2+, which facilitates the entrapment of Ca2+ ions, resulting in the CaOx crystal growth inhibition [TFF1 at the kidney may lead to the increase of CaOx crystals, contributing to the increased risk of nephrolithiasis.For further genetic function, we carried out two vector assays and discovered that the vectors carrying rs3761376 A allele both had a significantly reduced luciferase and transcriptional activities compared to those carrying G allele. Rs3761376 was located in the promoter of  factors –40. Therhibition , 41. It TFF1. Many previous researches indicated that TFF1 was an estrogen-induced protein and rs3761376 was capable of changing the motifs of specified transcription factors, especially estrogen receptor 1 (ESR1) [TFF1 genetics and nephrolithiasis risk in terms of mechanism, we need do more well-designed functional study. Moreover, some studies have pointed out that a multitude of comorbidities  are associated with an increased risk of kidney stone disease [TFF1 tagSNPs with nephrolithiasis is independent of these factors. In addition, we could not detect the TFF1 levels in the kidney samples of the controls, so it is impossible to evaluate the difference in the expression levels of TFF1 between the patients and the controls, and it’s difficult to explore the effects of tagSNPs specifically through TFF1 causing kidney stones.Nonetheless, considered the complex process for kidney stone formation, it is impossible that any single SNP or gene would exert a clear influence on nephrolithiasis risk. Although a strong correlation between rs3761376 and nephrolithiasis risk was observed, we didn’t explore the potential effect of rs3761376 on regulation of 1 (ESR1) , 42, 43. disease –46, but  disease  reportedTFF1 promoter activity reduced by rs3761376 A allele could at least partially lead to the decreased TFF1 expression and ultimately increase the risk of kidney stones. The results show that SNP rs3761376 can be used as a potential biomarker for predicting the risk of kidney stones in Chinese populations.In summary, our study provided evidence that the Additional file 1: Table S1. The distribution of the demographic characteristics of discovery set.Additional file 2: Table S2. The distribution of the demographic characteristics of validation set.Additional file 3: Table S3. The primers applied for reverse transcription


Black cumin meal (BCSM) and copra or coconut meal (CM) are by-products resulting from the extraction of black cumin and coconut oil, respectively. They contain valuable functional nutrients such as proteins, dietary fibers and phytochemicals. The objective of this study was to investigate the effects of different levels of BCSM and CM in broiler diets on the growth performance and cecal microbiota of birds. It was found that the addition of coconut meal to broiler diet significantly improved body weight and feed conversion ratio (FCR). In addition, the hot carcass dressing percentage was increased due to the addition of BCSM and CM either separately or in combination compared to the control group. In conclusion, the use of BCSM and CM in broiler diets had positive effects on growth performance and gut health (inhibitory effect against pathogenic microbes and improvement of gut microbiota diversity).Clostridioides difficile, Roseburia and Streptococcus were not significantly changed in all treatments (p > 0.05). Treatment T1 resulted in a significant decrease in gene expression of the entire microbiota, while treatment T2 resulted in a significant increase in gene expression of all microbes, leading to an enriched and diverse microbial community. It can be concluded that supplementation with 10% BCSM is beneficial in inhibiting pathogenic microbes during early post-hatch days. In contrast, CM may promote and enhance the diversity of microbial communities during broiler growth. The inclusion of non-conventional feed ingredients in poultry diets may improve growth performance and may reduce the cost of broiler feed.The objective of this study was to investigate the effect of dietary supplementation with black cumin seed meal (BCSM) and coconut meal (CM) on the performance and cecal microbiota of Cobb 500 hybrid broilers. The study was conducted on 600 chicks on the first day of hatching; the chicks were randomly distributed equally into 12 equal-sized floor pens. Four dietary treatments  were replicated three times (50 chicks/replicate): C was the control group; T1 was supplemented with 10% BCSM; T2 was supplemented with 10% CM; T3 was supplemented with 5% BCSM and 5% CM. At slaughter age on day 35, our findings showed that treatment T2 increased significantly body weight and feed conversion ratio (FCR) compared to C, T1 and T3. In addition, the hot carcass dressing percentages in treatments T1, T2, and T3 were significantly higher than that of the C group. The results of relative normalized comparative gene expression of  Nigella Sativa) meal (BCSM) is a by-product of black cumin oil production. This meal represents a valuable source of plant protein in many countries. It was found that the inclusion of BCSM and cumin seed by-products in broiler diets improves the growth, health and meat characteristics of broilers [Black cumin seed  meal (CM) is a by-product obtained from the mechanical extraction of oil from coconuts [Copra meal or coconut . These chicks were randomly housed in an open-sided broiler house that was divided into 12 equal-sized floor pens. Four dietary treatments were replicated three times (50 chicks/replicate). Dietary treatments were as follows: birds in the first treatment (control group) received conventional broiler diets (C); birds in the second treatment (T1) received diets containing 10% BCSM; birds in the third treatment (T2) received diets containing 10% CM; birds in the fourth treatment (T3) received diets containing 5% BCSM and 5% CM. Birds were housed according to the Cobb broiler management guide.Chicks were raised on wood shavings 10 cm deep. Moisture in the litter was managed daily; wet spots were removed and new shavings were added whenever necessary. The house temperature was 32 °C for the first seven days and was lowered by 2.5 °C weekly thereafter. Chicks were exposed to a conventional light regimen; birds were exposed to 24 h of light for the first 4 days and 23 h of light and 1 h of darkness thereafter.Iso-energetic and iso-protein diets were formulated in mash form in accordance with the recommended  nutrientBody weight gain was calculated by subtracting the live weight at the beginning of the week from the live body weight at the end of the week. Average feed consumed per bird was calculated by dividing the amount of consumed feed by the number of chicks of every dietary treatment. FCR was calculated weekly as the amount of feed consumption per average of body weight gain (average weekly feed consumption (g)/average weekly gain (g)).At the end of the experimental period, nine chicks were randomly selected from each replicate. The birds were slaughtered to determine meat yield and characteristics of cut-up parts. The selected birds were slaughtered and processed in a small-scale facility. After slaughter, birds were bled, scalded (60 °C for one minute), plucked, eviscerated, dressed and finally dissected according to the commercial protocols of the facility. The initial body weight of each bird was recorded. The carcasses were firstly cooled with tap water at ambient temperature, followed by cooling in a refrigerator at 4 °C for the next day. Carcass, plucked and dressed weights (Dressing % = Carcass weight/live weight * 100) were then measured. Following cutting, the breasts, thighs, drumsticks, wings, neck, legs, head and giblets  of each slaughtered bird were weighed.At 35 days of age, euthanasia was performed humanely by manual cervical dislocation (CD) and confirmed by loss of all reflexes and musculoskeletal movements; luminal contents of the cecal samples were collected by squeezing the ruptured ceca and extracted from five chicks from each group at the end of the experiment at 35 days of age. Total DNA was extracted from 220 mg of cecal luminal contents according to the manufacturer’s protocol of the QIAamp DNA Stool Kit . Gene expression levels of cecal microbiota were examined and evaluated by relative quantitative real-time RT-qPCR analysis. Primers targeting 11 cecal microbiota were used, as shown in Relative quantitative CFX96 Touch Real Time qPCR analysis  was performed using the GoTaq qPCR Master Mix kit . The 20 µL reaction mix was prepared from 10 µL of the GoTaq qPCR Master Mix (2X), 2 µL of the forward primer pm/μL, 2 µL of the reverse primer pm/μL , 2 µL of−ΔΔCt) method according to Livak and Schmittgen [p < 0.05.The original data were arranged using Excel 2007 software . Gene expression levels were expressed as means ± SE. The relative quantitative expression results were calculated using the comparative ct- of the experimental birds revealed a significant difference. Birds fed the diet containing 10% CM had the highest body weight compared to body weight in treatments C, T1 and T3 at different ages . There were no significant differences in body weights between C, T1 and T3. Differences in FCR between treatments were more significant at ages 22–28 days and 29–35 days than at ages 1–7 days and 8–14 days. Birds in treatment T2 had significantly lower FCR than treatments C, T1 and T3 at 22–28 days and 29–35 days of age. At ages 1–7 days and 8–14 days, treatment T3 exhibited significantly higher FCR than treatment T2 while the remaining treatments had moderate differences.After completion of the experiment, nine birds from each replicate in each feeding treatment were randomly selected and sacrificed to determine meat yield and percentage of cuts. The effects of BCSM, CM and a combination of both meals on meat yield and cut percentages (expressed as g/whole carcass weight) are shown in p < 0.05), carcass weight  and dressing percentage  than those in control treatment C. There were no differences in live weight, carcass weight and dressing percentage between treatments T1 and T3. Treatment T2 had significantly higher breast and drumstick weights than other treatments. Treatment T3 had higher intestine, liver, gizzard, crop and proventriculus weights than control treatment C, while there were no significant differences between treatments T1 and T2. The inclusion of black cumin seed meal and coconut meal either alone or in combination had no effect on the weights of the thigh, wings, neck, back, legs, head and heart.Birds in treatment T2 had significantly higher live weight  did not reach a significant level in any of the treatments or the control. T1 decreased all other microbiota significantly, causing lower expression of all microbial communities, while T2 increased all the microbial expression levels, causing enriched and diverse microbial communities. The mixture of BCSM and CM in T3 caused a variation in gene expression; Bifidobacterium and Lactobacillus were downregulated while Akkermansia muciniphila and Butyricicoccus increased when compared to the control. Escherichia coli and Enterobacteriaceae were less expressed in treatments than in the control .No significant differences in feed intake were observed between dietary treatments throughout the experiment. These results were in agreement with the results of Lee et al. . Our resThe body weights of broilers in the current study were significantly different between dietary treatments, with those fed CM having the highest body weight throughout the experiment. There were no differences in body weight and FCR of broilers fed the control diet or the diets containing BCSM or a combination of both BCSM and CM. The results of the current study were in agreement with those reported by EL-Deek et al.  and SundEnterobacteriaceae and E. coli than those found in the group that received the control diet.Coconut is mainly found in tropical and subtropical areas and has several health-promoting effects such as antiseptic, antitumor, bactericidal, antihelminthic, astringent, diuretic, refrigerant, stomachic, vermifuge, antioxidant and vasorelaxant effects . CM has The current study highlights the role of microbiota in broiler production and the use of feed ingredients that enhance microbial balance in the gut. CM showed superiority over BCSM. However, sequential feeding may be suggested to emphasize the role of microbiota, e.g., feeding with BCSM may be employed for the first week of life to create a low-pathogen environment, and then chicks may be fed with CM to enhance microbial diversity.


Information collected: age, sex, medical history, additional anomalies, anomalous vessel course, and respiratory symptoms. Data were analyzed with independent samples t-test and Fisher’s exact test. PALPAs were more commonly anterior than posterior. Mean age: 5.3 years (SD = 12.4) for anterior and 6.8 years (SD = 18.5) for posterior (p = 0.77). Respiratory symptoms: 20% of anterior and 60% of posterior cases (p = 0.032). Tracheobronchial anomalies: 35% of anterior and 60% of posterior cases (p = 0.182). Non-cardiac and non-tracheobronchial anomalies: 30% of anterior and 47% of posterior cases (p = 0.511). Kabuki syndrome: 25% of anterior and 6.7% of posterior cases (p = 0.207). In conclusion, respiratory symptoms were the only significant difference between anterior and posterior PALPA types.The aim of this study was to investigate the features of partial anomalous left pulmonary artery (PALPA) and differences between cases with posterior versus anterior a nomalous vessels in relation to the tracheobronchial tree. We hypothesized that statistical significance was dependent on the course of the anomalous vessel due to airway compression in the posterior type. This study included cases obtained from the literature ( Partial anomalous left pulmonary artery (PALPA) or duplicated left pulmonary artery (LPA) is a very rare congenital vascular anomaly. It is characterized by an arterial branch arising from the right pulmonary artery (RPA) and feeding the left lung, in addition to the presence of proper LPA arising from the main pulmonary artery in a normal fashion resulting in a duplicated LPA . Most PASimilar to what is the case with pulmonary artery sling, different tracheobronchial malformations including right upper lobe bronchus, bridging bronchus and tracheobronchial stenosis, as well as congenital heart disease, skeletal, gastrointestinal, and genitourinary malformations have been reported in PALPA patients. Current literature on this rare pathology is limited to single case reports and small case series. The purpose of this study was to systematically collect the largest collection of PALPA cases from the literature and a single teaching institution to help better delineate the unique clinical and radiologic features of this rare entity and further assess differences between subtypes according to their anterior versus posterior relationship to the tracheobronchial tree. The literature was searched for cases of PALPA using the Scopus database and following the PRISMA criteria . This stData were analyzed using the IBM SPSS statistics (Version 27) predictive analytics software. In deciding between the Chi-squared test and the Fisher’s exact test for the analysis of categorical variables, our relatively small sample sizes (less than 30 for each cohort of anterior versus posterior PALPA) made the Fisher’s exact test the most accurate and appropriate test for the present study. To facilitate data analysis, PALPA with an anterior and anterior–inferior course were grouped together. Categorical variables analyzed included presence or absence of respiratory symptoms, tracheobronchial anomalies, left lower lung supply, cardiovascular anomalies other than PALPA, anomalies not of cardiovascular or tracheobronchial origin, and a reported diagnosis or suspicion of Kabuki syndrome. To allow for analysis, all categorical variables were codified with binary numerical values representing presence of absence of said variable  in the statistical software. Age represented the single continuous numerical variable analyzed via a two-sample t-test. Given that different publications reported age in different units , all ages were converted to days using the conversions 1 year = 365 days, 1 month = 4 weeks, and 1 week = 7 days to allow for the appropriate pooling of data.p = 0.77). Sex was reported in 16/20 of anterior PALPA patients, with 10/16 (62.5%) being male; similarly, 10/15 of posterior PALPA patients had the reported sex with 6/10 (60%) being male. Respiratory symptoms were reported in 4/20 (20%) of anterior PALPA and included “issues with airway”, “cough” × 2, and “shortness of breath” while 9/15 (60%) of those with posterior PALPA had respiratory symptoms which included “increased work of breathing”, “dyspnea/dyspnea on exertion” × 2, “stridor” × 2, “pulmonary infections and cough”, “pulmonary infections and pulmonary hypertension”, “cough and shortness of breath” and “severe respiratory distress” (p = 0.032). Tracheobronchial anomalies were reported in 7/20 (35%) of anterior and 9/15 (60%) of posterior PALPA patients (p = 0.182). The anomalous vessel was reported to feed exclusively the left lower lung in 15/20 (75%) cases of anterior and 10/15 (66.7%) of posterior PALPA (p = 0.712). Cardiovascular anomalies other than PALPA were reported in 18/20 (90%) of anterior and 9/15 (60%) of posterior cases (p = 0.051). Anomalies not of cardiovascular or tracheobronchial origin were reported in 6/20 (30%) of anterior and 7/15 (46.7%) of posterior cases (p = 0.481). Confirmed or suspected Kabuki syndrome was reported in five cases of anterior (25%) and in only one case of posterior (6.7%) PALPA (p = 0.207).The Scopus database search yielded 645 articles that resulted in 21 articles describing 33 individual cases of PALPA after application of the inclusion and exclusion criteria. These 33, in addition to two cases from a single participating teaching institution, resulted in a total of 35 cases . AnterioLPA abnormalities can be distinguished by their relation to the tracheobronchial tree, the presence of native LPA, an anomalous origin and suspected airway compression. In patients with PALPA, the accessory LPA may have a variable course, including between the tracheobronchial tree and the esophagus or may run either anterior or antero-inferior to the tracheobronchial tree . This anPALPA and other pulmonary artery abnormalities are thought to be related to developmental defects of the sixth aortic arch ,18,27,28In the present study, respiratory symptoms were reported in over half (60%) of patients with a posterior PALPA course which was a statistically significant difference when compared to the 20% of anterior PALPA patients reporting such symptoms, although no significant difference was found between presence of tracheobronchial anomalies. These findings suggest that symptoms may potentially be related to a dynamic narrowing of the tracheobronchial tree, which may not be seen without imaging. Dynamic evaluation of PALPA anomalies using ultrasound or CT with careful attention to the relationship between the anomaly and tracheobronchial tree during the respiratory cycle may be a worthwhile area of further research. To our knowledge, this is the first reported instance of a significant clinical difference between PALPA subtypes. Awareness of PALPA course may provide the treating team with an ability to anticipate and triage those patients at higher risk of developing respiratory symptoms prior to presentation or even antenatally.In an autopsy series of 68 LPAS patients, 40% were found to have tracheobronchial abnormalities including hypoplasia of the distal trachea (38%) with complete cartilaginous rings, stenosis of the left main bronchus (5%) and direct origin of the right upper lobe bronchus from the trachea  . Major cBridging bronchus represents a rare congenital anomaly in which the right lower lobe and occasionally the right middle lobe ventilation is supplied by an aberrant bronchus arising from the left main bronchus crossing the mediastinum while the right upper lobe is supplied by the right main bronchus. It is commonly accompanied by a long or short segment of tracheal and/or bronchial stenosis . AffecteIn this series, there was a high prevalence of congenital heart disease with a broad spectrum of conditions ranging from no additional cardiovascular anomalies to complex multiple additional cardiovascular anomalies. Similarly, in a series of patients with LPAS, Xie et al. reported 85% of patients were associated with congenital heart disease, with ventricular septal defects 47%), atrial septal defects (43%), patent ductus arteriosus (34%) and persistent left superior vena cava (30%) being the most common [7%, atriaPatients with a diagnosis of Kabuki syndrome have been reported to have some type of congenital heart disease in 60 to 90% of cases, in particular coarctation of the aorta (23%-46%), atrial septal defect (20%) and ventricular septal defect (17%) ,32. ThisThis study is limited by its retrospective nature and the limited number of published literature on the topic related to the exceedingly rare nature of the condition and relatively new technology of cross sectional imaging. It is important to highlight that this work is limited by the exclusive evaluation of symptoms related to the respiratory system and did not assess for non-respiratory symptomatology. Additionally, not all published cases had all the information that was being investigated as some cases were published singularly as rare, interesting cases, while others were presented as single-center series. Lastly, the grouping together of anterior and anterior–inferior coursing PALPA did not allow for the statistical discrimination between these two PALPA subtypes, although their close morphology makes for any clinical differences to be probably unlikely. Notwithstanding, the results presently discussed are derived from what is to our knowledge the largest collection of patients with PALPA to date and helps gain new insights into this rare but clinically relevant entity. PALPA is a complex condition characterized by its anterior or posterior course about the tracheobronchial tree. This anomaly is most often diagnosed in childhood, is commonly associated with congenital heart disease, tracheobronchial anomalies and extra-thoracic abnormalities, most often feeds the left lower lung and may be associated with complex genetic syndromes. Of the analyzed variables, the only statistically significant finding was that respiratory symptoms were more common in posterior PALPA, possibly due to a dynamic narrowing of the tracheobronchial tree by a sling morphology. Additionally, our study demonstrates that posterior PALPA is more common than previously thought.


One concern caused by the changes in the ocean due to climate change is the potential increase of neurotoxic methylmercury content in seafood. This work quantifies the impact of global change factors on marine mercury cycles. The air–sea exchange is influenced by wind speed weakening and solubility drop of mercury due to seawater warming. The decreased biological pump shrinks the methylation substrate and causes weaker methylation. The advantageous light environment resulting from less attenuation by sea ice and phytoplankton increases the photodegradation potential for seawater methylmercury. Responses of seawater methylmercury can propagate to biota, which is also modulated by the changes in anthropogenic emissions and ocean ecology. Our results offer insight into interactions among different climate change stressors. 3Hg), a potent neurotoxin, is principally through the consumption of seafood. The formation of CH3Hg and its bioaccumulation in marine food webs experience ongoing impacts of global climate warming and ocean biogeochemistry alterations. Employing a series of sensitivity experiments, here we explicitly consider the effects of climate change on marine mercury (Hg) cycling within a global ocean model in the hypothesized twenty-first century under the business-as-usual scenario. Even though the overall prediction is subjected to significant uncertainty, we identify several important climate change impact pathways. Elevated seawater temperature exacerbates elemental Hg (Hg0) evasion, while decreased surface wind speed reduces air–sea exchange rates. The reduced export of particulate organic carbon shrinks the pool of potentially bioavailable divalent Hg (HgII) that can be methylated in the subsurface ocean, where shallower remineralization depth associated with lower productivity causes impairment of methylation activity. We also simulate an increase in CH3Hg photodemethylation potential caused by increased incident shortwave radiation and less attenuation by decreased sea ice and chlorophyll. The model suggests that these impacts can also be propagated to the CH3Hg concentration in the base of the marine food web. Our results offer insight into synergisms/antagonisms in the marine Hg cycling among different climate change stressors.Human exposure to monomethylmercury (CH However, its effectiveness is compounded by the changing climate and marine environment  by employing a global ocean model for Hg . We consider a simulation running from 1991 to 2000 as the present-day or base scenario. To disentangle the impact of different influencing factors, we conduct a set of sensitivity experiments by alternatively changing one factor to the condition in 2091 to 2100 with other factors the same as the present-day scenario. These factors include ocean physics such as ocean current velocity and mixing, seawater temperature, shortwave radiation, near-surface wind speed, and sea ice coverages, ocean biogeochemistry such as seawater chlorophyll (Chl), dissolved organic carbon (DOC), particulate organic carbon (POC), and organic carbon remineralization rates (OCRR); and ocean ecology such as plankton community structure. We also differentiate the ocean biogeochemistry changes caused by ocean physics and acidification  . The input of Hg from the atmosphere is held constant for all the present-day and future scenarios, so the difference between them could represent the impacts of individual changing ocean environmental factors.Predicting the responses of marine Hg cycles to climate change remains challenging, requiring an understanding of multiple influencing mechanisms and a comprehensive process-based model . Zhang el for Hg  coupled l for Hg  that is −12 mol/L) at present-day, consistent with the observed values in both magnitudes and spatial distributions . There are three tracers of inorganic Hg in the model, which are dissolved phase elemental Hg (Hg0) and divalent Hg (HgII), and particulate phase Hg (HgP) .We calculate a modeled global average total inorganic Hg concentration in the surface layer  of 0.51 ± 0.16 picomolar (pM = 10Hg (HgP) . The bidSI Appendix, Fig. S2A) reduces the solubility of gaseous elemental Hg [(Hg0(g)]  , resulting in a rise in the surface total inorganic Hg concentration. Indeed, intensified wind in certain areas could cause a faster air–sea exchange velocity and subsequent Hg0 evasion. Yet on a global basis, the net evasion flux of Hg0 is predicted to decrease by 2.9% due to wind speed changes, and this leads to a global increase of 6.3% in the modeled surface total inorganic Hg concentrations  , facilittrations C. On thetrations D. To sumP concentration is calculated from dissolved HgII concentration using a partition coefficient and local POC level. Then with the export of organic particles in the water column, HgP sinking flux is calculated. In the subsurface layer , the simulated global average total inorganic Hg concentration is 0.98 ± 0.38 pM, consistent with observations. The spatial distribution of total inorganic Hg in the subsurface ocean from our model is close to available observations as well, indicating that our model also captures the vertical distribution of observations .A key process in transporting inorganic Hg to subsurface waters is particle sinking , which largely shapes the vertical profile of seawater Hg concentrations. The HgP sinking flux and a 2.0% increase in Hg0 evasion flux are modeled, resulting in a decrease in the subsurface inorganic Hg of 4.2% 2Hg] in the surface ocean, especially the latitudinal pattern . The formation rates of methylated Hg [CH3Hg and (CH3)2Hg] are parameterized proportionally to microbial respiration/remineralization of organic carbon in the model, which is influenced by ocean warming .The seawater CHts CH3Hg . The mod warming , 9, 25. 3Hg changes is chlorophyll concentrations . Earlier studies of in situ measurements have reported a global-scale decline in surface chlorophyll over the past century , however, have a modest effect on photodemethylation processes due to a lower specific attenuation coefficient of light than chlorophyll in our model. There is also a positive feedback loop as the increased light decomposes DOC and further increases the incident light, which is, however, not included in the ecosystem model : at high latitudes, surface solar radiation increases, enhancing the photodecomposition of CH3Hg, while surface solar radiation decreases in the tropical regions , and the projected ocean warming results in a 5.9% decrease in CH3Hg 2Hg] (3)2Hg and also promote the loss of seawater CH3Hg. Recent measurements show lower Arctic seawater (CH3)2Hg concentrations, suggesting likely less importance for seawater (CH3)2Hg evasion (3)2Hg from sea ice is close to that from seawater, which also confirms the influence of Arctic sea ice regime shift on the long-term trends of methylmercury. The influence of other factors on future seawater (CH3)2Hg concentration is in line with those of seawater CH3Hg.The dark demethylation via biotic/abiotic processes is presumed to be temperature-dependent in the model , 33 2Hg] , which w evasion . Schaefe evasion  also sug3Hg concentrations in the surface ocean . This denotes decreased microbial methylation activity and will lead to a decrease in the surface CH3Hg concentration. This possible variation is consistent with what previous studies have pointed out . As elevated seawater pCO2 promotes diffusive uptake of CO2 by phytoplankton, this saves the energy consumption of CO2 uptake and diverts the energy to other activities such as photosynthesis, which potentially increase the OCRR .The results of sensitivity experiments suggest that future changes in OCRR will also have some impact on CHce ocean F. Futurented out . Althougthe OCRR –40. This3Hg is influenced by a variety of factors such as the vertical and horizontal transport of CH3Hg and in situ methylation/demethylation activities. The degradation of CH3Hg below euphotic layers involves both biotic and abiotic pathways .In the subsurface ocean, the distribution of seawater CHpathways , 42. Thipathways . Future decrease J, similadecrease , and an decrease  is simul3Hg below the top 100 m, resulting in a substantial decrease of 18% in the global mean. This can be attributed to the significant decline in CH3Hg production in regions that exhibit the highest subsurface CH3Hg concentration, i.e., eastern tropical Pacific and Benguela upwelling system as noted by Zhang et al.  indicate the reduction in microbial methylation, which causes the decrease of subsurface CH3Hg. OCRR changes induced by ocean acidification have a relatively smaller impact on subsurface CH3Hg (~1%), consistent with previous studies  is simulated to reduce the seawater CH3Hg concentration in the subsurface ocean by 4.7%. As was pointed out in the previous discussion of subsurface inorganic Hg, the substrate HgII required for methylation would be reduced, thereby inhibiting the formation of CH3Hg  with seawater CH3Hg concentration. VCF varies as a function of phytoplankton cell diameter (d) and seawater DOC concentration . The phytoplankton CH3Hg concentration is thus directly limited by the availability of CH3Hg in seawater. Hence, simulated plankton CH3Hg concentrations adjust similarly to seawater CH3Hg concentrations in response to climate change-driven alterations . The chemical composition and origin of DOC are known to impact the bioavailability of CH3Hg . Our simulation reveals that this will increase the global average concentration of CH3Hg in all phytoplankton by 7.0%, including a 9.0% increase attributable to ocean acidification and a 2.0% decrease to physical factors, consistent with earlier findings in Zhang et al.  and other large (10 μm) are modeled to migrate to the polar regions due to ocean warming, which could cause up to a factor of two changes in polar regions (Synechococcus (d = 1.8 μm) under the impact of ocean acidification also increases its CH3Hg quota in most regions   increases, this favorable transition to more Synechococcus in the community structure would promote phytoplankton to uptake more CH3Hg, especially in high latitudes (Prochlorococcus (d = 0.6 μm) to Synechococcus causes a decrease in plankton uptake of CH3Hg in the tropical regions as the former is even smaller in cell size. Subsequently, the CH3Hg concentration in the small zooplankton (d = 30 μm), which prefers to feed on smaller phytoplankton , is projected to increase substantially by 65.7% under the influence of planktonic community changes. However, for the large zooplankton (d = 300 μm), which has higher CH3Hg concentrations due to slower elimination, the loss of diatoms in the equatorial Pacific and the Southern Ocean disrupts the CH3Hg transfer to the large zooplankton. The changes within the food web dynamics bring about a great 71.0% decline in the CH3Hg concentration of large zooplankton. Compared with the changes in the availability of seawater CH3Hg, plankton community changes may have an overall smaller global impact but could affect the biogeographic distribution and play a much more important role at basin scales.Anticipated changes in plankton community dynamics are also a key determining factor of future CHortality   . Accordiatitudes . On the 3Hg exposure is tightly associated with dietary seafood, the response of coastal area Hg contamination to future climate change is of great intrigue. Indeed, coastal fishing and aquaculture are becoming a more dominant source of human seafood consumption worldwide, especially the Asian countries . We use results from different scenario simulations to obtain changes in mean Hg concentrations at coastal grid points and find that the responses are mostly consistent with those of the global mean . Under such circumstances, the retention of inorganic Hg in the surface layer allows the facilitation of sea–air exchange processes and the loss of inorganic Hg from seawater. Besides, compared to a 7% increase in the global mean, an 11% increase in surface phytoplankton CH3Hg at the coastal scale indicates that future changes in plankton community structure are more favorable for CH3Hg bio-uptake near shore.Since CHbal mean . However3Hg subjected to photodegradation by forming complexes with CH3Hg or inhibit photodemethylation by attenuating solar radiation (3Hg concentration (Methods). Ocean physics and other parameters are kept the same to highlight the effects of eutrophication.The effects of excessive nutrient discharge to estuaries and coastal waters, for example, include direct DOC enrichment, which may either increase the activity of CHadiation . Besidesntration . We perfntration , 50. TheSI Appendix, Table S4 and Fig. S7). For example, the nearshore regions of East Asia, the Bay of Bengal, and the Arctic are subject to a more prominent increase in surface seawater CH3Hg concentrations due to the faster increase in riverine nutrient load, while decreasing trends are modeled for the Arabian Sea and the North Atlantic . Marine productivity changes in these regions alter microbial Hg methylation . The changes in pigments, such as chlorophyll and DOC, also influence the photodegradation of CH3Hg. Furthermore, the Hg-related risks for coastal eutrophication are possible to be of greater concern when combined with future physical changes in the coastal areas and the change of riverine Hg discharges, which could be a considerable source of the Hg burden in coastal oceans   are needed to better predict the impact of future light environment on the marine Hg cycle.Our projection highlights the potential spur of demethylation potential of CH% day−1) . Experimaterials . Lehnheraterials  observed3Hg bioconcentration through the global ocean also requires more measurements of VCF in environment-relevant concentrations with the consideration of the influence of seawater chemistry . Thus, the simulated changes in seawater or plankton CH3Hg cannot be directly extrapolated to other trophic levels. Indeed, Schartup et al. : Seawater Hg0 evasion is enhanced by ocean warming but suppressed by decreased wind speed, considerable variation in light environment boosts the photodemethylation of CH3Hg, reduced vertical mixing decreases subsurface CH3Hg fluxes into the surface ocean, and the loss of sea ice cover facilitates air–sea exchange and also increases photodemethylation. These impacts could also be propagated to the CH3Hg concentration in plankton and its potential human exposure.This study sets out to investigate the impact of climate-driven changes on global marine Hg cycling in the hypothesized twenty-first century. Even though the overall prediction is subjected to significant uncertainty, especially regarding the formation of seawater CHClimate changes could also exacerbate marine Hg contamination. Previous studies also indicated the possibility of Hg release as a consequence of glaciers melting and permafrost thawing , and the3Hg and improving relevant regulatory strategies in the context of global climate change.Taken together, there is still a great technical challenge to predict the future marine Hg cycles, and the uncertainty arises from our mechanical understanding of the Hg cycle itself, the direction and magnitude of climate change and incurred environmental changes, as well as the future path of anthropogenic emissions. Our results are thus diagnostic, yet offer insight into synergisms/antagonisms on marine Hg cycling among ocean warming, light environment, circulation change, ocean acidification, and other ecological and biogeochemical stressors. This may present a perceptible way of identifying future human exposure to CH0) and inorganic divalent Hg (HgII), methylation and demethylation of monomethylmercury (CH3Hg) and dimethylmercury [(CH3)2Hg], air–sea exchange of Hg0 and (CH3)2Hg, partitioning between dissolved and particulate phases, particle-bound Hg sinking, and CH3Hg bioaccumulation within marine plankton food webs. Further details can be found in SI Appendix, Methods and Table S5.We apply a global three-dimensional ocean model for mercury (Hg) to calculate the impacts of climate change on the marine Hg cycle. We use the MITgcm-Hg model, which comprehensively accounts for the physical, chemical, and biological processes that control marine Hg biogeochemistry. MITgcm-Hg was described and tested against measurement data by Zhang et al. . The modhttps://esgf-node.llnl.gov/search/cmip5) multi-model mean for the future scenario. Details of the modeling system are described in SI Appendix, Methods and Fig. S5 shows the model coupling configuration for this study. The model is run for 10 y for each scenario simulation (1991 to 2000 for the present-day or 2091 to 2100 for the future). This setup allows Hg species to achieve a steady state for responses to changes in ocean physics and biogeochemistry  framework  that prohemistry . The upphemistry  and are circulation, temperature, radiation, wind speed, sea ice, community structure, Chl, DOC, POC, and OCRR, in which the above variables are altered, respectively. The experiment community structure allows variables of phytoplankton biomass, zooplankton grazing fluxes for each phytoplankton category, and mortality fluxes of zooplankton, which reflect the structure of planktonic food web, to change to future conditions. Experiment all represents that all these factors above are altered to the future conditions.To unravel the effects of individual factors on the marine Hg cycle, a suite of simulations is performed in the hypothesized twenty-first century under the business-as-usual scenario. First, we simulate the marine Hg cycling from 1991 to 2000 using the ocean physical state results and Darwin model results from the same time period and other physical variables  that are representative of the present-day conditions to serve as the present-day baseline condition. Each experiment in the future climate change scenario then simulates the year 2091 to 2100. Separating the effects of different factors is achieved by alternatively changing merely one factor to the future condition with other variables repeating present-day conditions. These factors specifically include ocean velocity and mixing, seawater temperature, surface downward shortwave radiation, near-surface wind speed, sea ice coverage, marine plankton community structure, chlorophyll concentration, DOC concentration, POC concentration, and OCRR. A total of 10 sensitivity experiments are performed and denoted as community_physics, Chl_physics, DOC_physics, POC_physics, and OCRR_physics, in which “_physics” indicates that pCO2 was held at present-day conditions while all other climate fields are allowed to change. Similarly, the scenario with a subscript “_acid” is ascribed to ocean acidification .To further distinguish the effects of ocean acidification and physical factors  on ocean biogeochemistry , we supphttps://doi.org/10.17632/br2psms8b9.1. Other resources of public accessible data are given in the article and/or SI Appendix.Modeling results and analysis scripts in this study are deposited in Mendeley Data: Appendix 01 (PDF)Click here for additional data file.