With the aging of the global population, accumulating interest is focused on manipulating the fundamental aging‐related signaling pathways to delay the physiological aging process and eventually slow or prevent the appearance or severity of multiple aging‐related diseases. Recently, emerging evidence has shown that RNA modifications, which were historically considered infrastructural features of cellular RNAs, are dynamically regulated across most of the RNA species in cells and thereby critically involved in major biological processes, including cellular senescence and aging. In this review, we summarize the current knowledge about RNA modifications and provide a catalog of RNA modifications on different RNA species, including mRNAs, miRNAs, lncRNA, tRNAs, and rRNAs. Most importantly, we focus on the regulation and roles of these RNA modifications in aging‐related diseases, including neurodegenerative diseases, cardiovascular diseases, cataracts, osteoporosis, and fertility decline. This would be an important step toward a better understanding of fundamental aging mechanisms and thereby facilitating the development of novel diagnostics and therapeutics for aging‐related diseases. RNA modificaitons in aging‐related diseases Molecular hallmarks of aging include genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, deregulated nutrient sensing, mitochondrial dysfunction, cellular senescence, stem cell exhaustion, and altered intercellular communication. In humans, aging can be correlated to increased risks of multiple diseases such as neurodegenerative diseases, cardiovascular diseases, osteoporosis, metabolic dysfunction, defective tissue repair and regeneration, decreased regulation of gut microbes, and cataracts  tail in mature eukaryotic mRNAs, more than 100 distinct chemical modifications have been found to actively regulate mRNA behaviors, including differentially processing, splicing, translation, and decay  groups have been added to the ribose. The 2’‐OMe modification was first demonstrated in bacterial mRNA to affect translation efficiency  conserved across eukaryotes  family of enzymes on both intermolecular and intramolecular double‐stranded RNAs longer than 20 bp. Mammals have 3 ADAR enzymes, ADAR1 and ADAR2 being catalytically active while ADAR3 lacks catalytic activity. Considering both coding and non‐coding transcripts, tens of thousands of A‐to‐I editing sites have been identified in mice and millions have been identified in humans. A‐to‐I editing levels vary across transcripts, tissues, and throughout development ranging from 1 to 100 percent at any given site  that suppress gene expression post‐transcriptionally. Emerging evidence shows that they harbor multiple RNA modifications, which are catalyzed by the same enzymes as other RNA species. These modifications regulate their biogenesis, stability, and base‐pairing with targets. After transcription, the biogenesis of microRNAs starts with the processing of primary microRNAs (pri‐miRNAs) by the microprocessor complex formed by RNA‐binding protein DGCR8 and the type III RNase DROSHA. In 2015, Alarcon et al. discovered that in mammalian cells, METTL3 methylates pri‐miRNAs and marks them for recognition and processing by DGCR8, promoting miRNA maturation. Another methylation modification, “internal m7G,” catalyzed by METTL1, has been found to occur on pri‐miRNA and affects precursor‐miRNA processing. Notably, precursor‐miRNA is a double‐stranded RNA specie, making them potential substrates for adenosine deaminases acting on RNA (ADAR) enzymes. As miRNA's function is dependent on their base pairing with the target mRNA, A‐to‐I editing in miRNAs may modulate their target specificity, resulting in decreased suppressing efficiency of one or more downstream target genes. Other modifications such as m5C and pseudouridine can also affect the binding of miRNAs to targets  through base pairing and it guides the modifying enzymes to the modification sites; Box C/D snoRNAs guide 2’‐OMe and Box H/ACA snoRNAs guide pseudouridylation , Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), stroke, and frontotemporal degeneration. Accumulating evidence has indicated that age‐related neurodegenerative diseases result from various reasons. Among them, epigenetic changes especially RNA modification that could have serious implications in this aspect should be further explored.3.1AD was first described by Alois Alzheimer in 1906. It is one of the most damaging aging‐related neurodegenerative diseases, the prevalence of which increases as the global population ages , it is found that the expression level of METTL3 was significantly reduced along with m6A levels in 5xFAD mice when compared with control mice , m7G, and 2’‐O‐methylguanosine, were increased while N2, N2,7‐trimethylguanosine , and N2,N2‐dimethylguanosine were dramatically decreased in the 15‐25 nt RNA fraction from the cortex of AD brains. RNA‐seq analysis revealed that most of these fractions are miRNAs. Interestingly, in the 30–40‐nt small RNA fraction, Cm, 2’‐O‐methyluridine (Um), and m7G modifications showed higher levels compared with controls whereas m1G, m2,2,7G, and pseudouridine modifications were reduced Figure . By usin3.2ALS is one of the most common age‐related neurodegenerative diseases characterized by progressive weakness and muscle atrophy, causing damage to upper and lower motor neurons  site of GluA2, a subunit in the majority of AMPA receptors in the adult brain, and changes the glutamine  to an arginine (edited to CIG and translated as CGG) within the ion pore of GluA2, which is indispensable for normal AMPA receptor function  and stroke  . These findings underscore the significance of this novel mechanism of cardiac hypertrophy. It is well known that leptin directly induces cardiomyocyte hypertrophy. Recently, Gan et al. demonstrated that the expression levels of FTO were upregulated after leptin treatment in cultured myocytes, suggesting its potential role in mediating the hypertrophic response to leptin , was significantly increased in TAC‐induced hypertrophic mouse hearts. CHAPIR directly interacted with METTL3 which suppresses the m6A modification of Parp10 mRNAs thereby increasing Parp10 protein levels and promoting NFATC4‐induced pathological cardiac hypertrophy , NOP10, NHP2, and GAR1. Histological analysis has revealed that in Dkc16Aging‐related osteoporosis is characterized by low bone mass and over‐accumulation of fatty tissue in the bone marrow environment that increases the risk of fracture  were generated. These mice showed a significant decrease in bone volume and trabecular number at 30 weeks of age. Furthermore, the results of static and dynamic histomorphometric analyses showed that the bone formation rate in mutant mice was decreased by 66% with bone marrow adipocyte number per bone marrow area being increased when compared with controls. FTO functioned through demethylating and then enhancing the stability of the mRNAs of Hspa1a and other genes that can protect cells from genotoxic damage (Zhang et al., Oc KO mice manifest the phenotype consistent with age‐related bone loss. miR‐149‐3p was found to directly target FTO mRNA and modulate the adipogenic differentiation of bone marrow‐derived mesenchymal stem cells (Li et al., FTO is a key regulator associated with adipogenesis, and the complete depletion of FTO in mice results in postnatal growth retardation. FTO knockout mice have not only a significantly shorter body length over the lifetime but also a much lower bone mineral density (Gao et al., 7Age‐related fertility decline is inevitable and irreversible, especially for female reproductive potential. Several demographic and epidemiological studies have long recognized that female fertility declines with age, most notably the decline in ovarian function Leridon, . The mecAll m6A modifications of the granulosa cells of aged human ovaries were measured in order to investigate the relationship between ovarian aging and m6A modification. It was found the level of m6A modifications was significantly increased and the expression level of FTO was downregulated. m6A sequencing showed that increased m6A in the 3′UTR of FOS mRNAs resulted in reinforcing the stability of FOS mRNAs (Jiang et al., 8In conclusion, the field of epitranscriptomics has emerged rapidly in recent years and RNA modifications have been emerging as a new focus and novel therapeutical targets against aging‐related diseases. In this review, we have summarized RNA epitranscriptomic regulation and the mechanisms involved in aging‐related diseases. We found that RNA modification is involved in many diseases that are aging‐related, and plays an essential role in impacting mRNA stability, translation, and control of protein levels of key genes that are involved in pertinent disease‐associated pathways Table . CollectZ.J., D.S., L.H., Y.G., and G.L. conceived the manuscript. Z.J., D.S., L.H., S.S., and L.Z. drafted the manuscript, made the figures, and summarized the Tables. HI.L., P.G., G.V., Y.G., and G.L. finalized the manuscript, figures, and Tables. L.H. and Y.G. significantly contributed to the manuscript revision.The authors declare that they have no conflict of interest.


Several genetic models that recapitulate neurodegenerative features of Parkinson's disease (PD) exist, which have been largely based on genes discovered in monogenic PD families. However, spontaneous genetic mutations have not been linked to the pathological hallmarks of PD in non‐human vertebrates.Amazona ochrocepahala) siblings with a severe and rapidly progressive neurological phenotype.To describe the genetic and pathological findings of three Yellow‐crowned parrot  was performed on the three affected parrots and their parents.P < 0.05). WGS identified a single homozygous missense mutation (p.V559L) in a highly conserved amino acid within the pleckstrin homology (PH) domain of the calcium‐dependent secretion activator 2 (CADPS2) gene.The brains of affected parrots exhibited neuronal loss, spongiosis, and widespread Lewy body‐like inclusions in many regions including the midbrain, basal ganglia, and neocortex. Proteasome activity was significantly reduced in these animals compared to a control (CADPS2 gene causes a severe neurodegenerative phenotype with Lewy body‐like pathology in parrots. Although CADPS2 variants have not been reported to cause PD, further investigation of the gene might provide important insights into the pathophysiology of Lewy body disorders. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.Our data suggest that a homozygous mutation in the  Descriptions of neurodegenerative pathologies in parrots and other birds are uncommon, and in most instances the pathology is related to environmental exposures to toxins such as organophosphates and heavy metals that induce axonopathies.The Yellow‐crowned parrot  were brought to the School of Biosciences and Veterinary Medicine of the University of Camerino in Italy, showing severe neurological symptoms. Subsequently, DNA samples from their parents, two uncles, and their grandparents were acquired using blood collected during routine annual veterinary visits.Three 3‐month‐old Yellow‐crowned Amazons  testing for chlamydophila, avian polyomavirus, avian bornavirus, paramyxovirus, and beak and feather disease virus was conducted.2.33) were fixed in 2.5% glutaraldehyde for 24 hours and then in Millonig buffer for electron microscopy. After dehydration, the sections were embedded in epoxy resin. Semi‐thin toluidine blue 1% stained sections were produced to assess target areas for ultrastructural analysis. Ultra‐thin sections (75 nm) were then mounted on copper grids and examined under a Philips EM208S  transmission electron microscope.Given that the parrots were unable to feed themselves and the neurological symptoms worsened, with eventual death imminent, they were humanitarianly euthanized at the age of 3 months. A complete necropsy was carried out and all organs were fixed in 10% buffered formalin for histological examination. Brain tissue was stained with hematoxylin and eosin, Congo red dye, Luxol fast blue, PAS, and immunohistochemistry (IHC) was performed using an anti‐synaptophysin antibody , anti‐neurofilament antibody , and an α‐synuclein antibody . Small portions of brain  followed by a TUNEL assay. The expression of CADPS2 was analyzed and quantified using ImageJ/Fiji 1.52p software  as previously reported.To explore the involvement of a 2.4g for 20 minutes at 4°C and the supernatant was collected for enzyme activity assays and western blotting. Protein content was determined by the Bradford methodBrains were homogenized in 50 mM Tris buffer, 150 mM KCl, 2 mM EDTA, pH 7.5 (1:5 weight/volume of buffer). Homogenates were immediately centrifuged at 13,0002, 1 mM dithiothreitol, and 2 mM ATP. Brain homogenates were also analyzed through western blotting assays using anti α‐synuclein (C‐20) primary antibody . The bands were quantified by using a densitometric algorithm. Each western blot was scanned  and the obtained digital data were processed through Image J (NIH)Proteasome peptidase activities in brain homogenates were determined using synthetic fluorogenic peptides: Suc‐Leu‐Leu‐Val‐Tyr‐AMC was used for chymotrypsin‐like (ChT‐L) activity, Z‐Leu‐Ser‐Thr‐Arg‐AMC for trypsin‐like (T‐L) activity, and Z‐Leu‐Leu‐Glu‐AMC for peptidyl‐glutamyl‐peptide hydrolyzing (PGPH) activity.2.5Melopsittacus undulatus) reference genome (Budgerigar v6.3)Amazona ochrocephala genome. Annotation was performed with ANNOVAR softwarehttps://bioinfo.ut.ee/primer3-0.4.0/) and are available on request.DNA was extracted from liver of the three affected parrots and from blood of their family members. Whole genome sequencing (WGS) of the three offspring and their parents was performed at the University of Washington with 1 μg of DNA on a HiSeq 2000 Sequencing System . The WGS data were aligned using a standard BWA pipeline to the budgerigar , round to elongate, well‐defined eosinophilic structures that occasionally contained fine boat‐shaped clefts, imparting a crinkled appearance in both gray and white matter. These abundant neuronal and axonal eosinophilic inclusions lacked a distinctive core and halo and resembled Lewy bodies (LBs) and Lewy neurites,In the affected parrots, a conspicuous feature of neurons harboring α‐synuclein‐positive LBLIs was somal chromatolytic changes, defined by distension of the cell body, displacement of the nucleus toward the periphery of the soma, and dissolution of the Nissl substance. These neurons also showed nuclear condensation. Subsets of large neurons had this chromatolytic signature, showing TUNEL‐positive staining Fig. . SubsetsFinally, electron microscopy imaging identified the eosinophilic material constituting LBLIs as accumulations of short granular electrondense material on a translucid background and organized in short filaments at the periphery Fig. . The pre3.3P < 0.05; Fig. P < 0.05; Fig. Significantly reduced ChT‐L and T‐L proteasomal activities were observed in affected parrots compared to the control  gene. The ortholog position and mutation in the human CADPS2 gene located on chromosome 7 is: g.122130303C > G; c.1684G > C; p.V562L. This amino acid is highly conserved across species, including invertebrates  in the The onset of disease in these parrots was very early, even in comparison to early‐onset monogenic forms of PD in humans. This species of parrot has a long lifespan and has been reported to live up to 56 years in captivity.CADPS2 gene codes for a member of the CADPS protein family.CADPS1 and CADPS2, which are expressed at the highest levels in brain.The CADPS2 might induce neurodegeneration is by altering the appropriate release of BDNF.Taken together, these data suggest that CADPS2 regulates presynaptic BDNF release in dopaminergic neurons, which is particularly relevant to PD and other neurodegenerative diseases. In animal models of PD, BDNF enhances the survival of dopaminergic neurons, improves dopaminergic neurotransmission, and accelerates recovery of motor function.CADPS2 is located in the “autism susceptibility locus 1” on chromosome 7q31.32−/− knockout mouse was generated and assessed for behaviors that model facets of autistism.−/− knockout mice and the homozygous p.V559L parrots including large differences in the genetic background of the organisms and the nature of the specific mutations. While no CADPS2 protein was detected in the Cadps2−/− mouse brains, 2.5‐fold elevated levels of the mutant protein were found in the brains of the parrots, and how p.V559L alters CADPS2 function is not yet known. An example of such phenotypic heterogeneity has been reported for RAB39B in humans. Null mutations in the gene often result in mental retardation associated with autism and/or epilepsy,LRRK2 and SNCA.SNCA A30P mutation. In contrast to LRRK2‐overexpressing cells, CADPS activity was reduced ~20% and ~ 35% in WT‐ and A30P cells, respectively, when compared to control cells. The effect was again independent from disease‐causing mutations.Cadps2,Two of the most important familial PD‐related genes are A recent study that analyzed the contribution of all midbrain cell types to PD pathology using single‐cell sequencing of human mesencephalon tissue identified a neuronal cell cluster characterized by CADPS2 overexpression and low TH levels that was almost exclusively present in idiopathic PD but not controls.CADPS2 gene cause a severe neurodegenerative phenotype associated with LBLIs in parrots. Although CADPS2 variants have not been reported to cause neurodegenerative diseases in humans, further investigation of the gene in animal models might provide important insights into the pathophysiology of LB disorders.Our results suggest that mutations in the O.L.‐B.: conceptualization, methodology, data curation, formal analysis, visualization, writing – original draft preparation, review and editing. L.G.: resources, methodology, investigation, writing – original draft preparation. L.B., A.M.E., V.C., R.V., F.D., A.‐R.A., S.B., Lu.B., P.R., and M.C.S.: methodology, investigation, writing – review & editing. D.Y.: project administration, validation, writing – review and editing. M.O.D.: investigation, data curation, writing – review and editing. D.W.T.: conceptualization, writing – review and editing and supervision. G.R.: resources, methodology, investigation, writing – original draft preparation and supervision. C.P.Z.: conceptualization, funding acquisition, writing – original draft preparation, and supervision.None of the authors report any competing financial interests for this work.Table S1. Mutations identified in the Amazon parrots that met selection criteria.Click here for additional data file.Video S1. Supplementary Video.Click here for additional data file.


To evaluate and compare the effects of three courses of different structural patterns of electroencephalography neurofeedback on predominantly inattentive attention deficit hyperactivity disorder (ADHD‐PI) and combined ADHD (ADHD‐CT).θ, β, α, SMR and their ratios , and eighteen integrated visual and auditory continuous performance test (IVA/CPT) quotients were obtained and compared. Data were analyzed by SPSS software, and p < .05 was considered statistically significant.Thirty‐eight ADHD‐PI and ADHD‐CT children were selected and completed three courses of different structural patterns of electroencephalography neurofeedback according to their ADHD type. Before and after each course, relative power value of electroencephalography, including θ, three IVA/CPT quotients in both types and two comprehensive quotients in ADHD‐CT changed significantly . After two courses, θ/α, θ/β and five IVA/CPT quotients in both types, θ and α in ADHD‐PI, four comprehensive quotients, and four respond control quotients in ADHD‐CT varied significantly compared to before treatment and after one course . After three courses, α, β, θ, θ/α, θ/β and ten IVA/CPT quotients in both types changed significantly compared to before treatment and after one course . In addition, six IVA/CPT quotients in both types after three courses were significantly higher than those after two courses .After one course, Different structural patterns of electroencephalography neurofeedback targeted for ADHD‐CT and ADHD‐PI were both effective and feasible. Three courses of EEG neurofeedback were most effective.   The structural patterns of EEG neurofeedback for ADHD‐PI and ADHD‐CT patients were 1st–5th of “2.3According to the literature . Numerical variables were expressed as mean ± standard deviation. Age, IQ, and the sex ratio of children between ADHD‐PI and ADHD‐CT groups were compared by the independent sample t test and Chi‐square test, respectively. The changes or differences in IVA/CPT quotients and brain waves among each treatment course or between two groups were analyzed by repeated measures analysis . The Greenhouse–Geisser corrected F test  was used when the data did not conform to Mauchly's test of sphericity. The Bonferroni test was used for comparisons of the confidence interval of main effect, and the F test was used for interactions between two groups and indexes among each course of treatment. All tests were two‐tailed, and 33.1t = .664, p = .511). There were 32 boys and six girls, the sex ratio was 16:3, and there was no significant difference in the sex ratio between ADHD‐PI (15:4) and ADHD‐CT  groups. The mean IQ of all subjects was 101.81 ± 12.79, and there was no significant difference in the IQ of subjects between ADHD‐PI (101.74 ± 14.76) and ADHD‐CT  groups.Before treatment, age, gender and IQ of children between two groups were balanced and comparable. All subjects were aged from 6.02 to 11.78 years, with a mean of 8.29 ± 1.46 years, and there was no significant difference between ADHD‐PI (8.06 ± 1.28) and ADHD‐CT .After one treatment course of EEG neurofeedback, θ/α and θ/β in both types, θ in ADHD‐CT decreased significantly, whereas α in ADHD‐PI increased after two treatment courses of EEG neurofeedback . Similarly, after two treatment courses of EEG neurofeedback, θ in ADHD‐PI decreased significantly and α in ADHD‐CT increased significantly compared to before treatment . The data are presented in Table Compared to before treatment and after one treatment course, θ/α, θ/β, and θ in both types decreased significantly, whereas α and β in both types increased . In addition, SMR in ADHD‐PI was significantly higher than that before treatment . The data are presented in Table After one treatment course of EEG neurofeedback, VAQ, VIGA, FOCA, and FOCV in both types, as well as FAQ and AAQ in ADHD‐CT increased significantly . PRUA and PRUV in ADHD‐CT, as well as ARCQ, VRCQ, FAQ, AAQ, STAA, and STAV in ADHD‐PI after two treatment courses of EEG neurofeedback were significantly higher than those before treatment . The data are presented in Table VAQ, VIGA, VIGV, FOCA, FOCV, SPDA, and SPDV in both types, as well as ARCQ, VRCQ, FAQ, AAQ, CONA, CONV, STAA, and STAV in ADHD‐CT after two treatment courses of EEG neurofeedback were significantly higher than those before treatment and after one treatment course . FAQ, AAQ, VAQ, STAV, VIGA, VIGV, FOCA, FOCV, SPDA, and SPDV in both types, as well as ARCQ, VRCQ, PRUV, CONA, CONV, and STAA in ADHD‐CT after three treatment courses of EEG neurofeedback were significantly higher than those before treatment and after one treatment course . FRCQ and PRUA in ADHD‐CT, as well as FRCQ, ARCQ, VRCQ, CONA, and STAA in ADHD‐PI after three treatment courses of EEG neurofeedback were significantly higher than those before treatment .As shown in Table 4α wave increased. In our study, during whole three‐course structural paradigm treatment of EEG neurofeedback, θ in both types decreased gradually with the extension of treatment time. In addition, θ/β and θ/α in both types showed a downward trend after two and three treatment courses. By contrast, α and β in both types increased significantly after two and three treatment courses. These results indicated that both structural patterns of EEG neurofeedback for ADHD‐PI and ADHD‐CT patients were effective. In addition, changes in EEG were observed after two treatment courses, and the curative effect showed an obviously increasing trend with the prolongation of treatment course, suggesting that at least two consecutive treatment courses are required so as to achieve relatively satisfactory therapeutic effects.In the connective process of brain function, neural resource allocation is different when attention is used for task control  in both types increased gradually. Namely, subjects’ attention improved significantly after two treatment courses, and further improved and stabilized after three treatment courses. The results also indicated that different structural patterns of the EEG neurofeedback targeted for different ADHD types could improve the attention retention ability and concentration of patients with different ADHD types, and the therapeutic effects are basically same.Six control quotients, namely prudence, consistency and stamina quotients of visual and auditory signals, were used to measure the ability to stop, think, recognize, respond correctly to disturbances, maintain the response consistency over time, and remain stable throughout the test, respectively. They jointly reflect the overall coordination and volitional control abilities of the subjects. In ADHD‐CT, prudence, consistency, and the stamina quotients of children increased significantly after two treatment courses, revealing that the ability to deal with disturbance and attention control of these children improved dramatically. And further improvement and stabilization of these two abilities were achieved after three treatment courses. In ADHD‐PI, only the stamina quotients of children increased after two treatment courses. This was reasonable because the ability to deal with disturbance of external stimuli or information and attention control of ADHD‐PI patients was still within normal range. And after each treatment course, the corresponding quotients reflecting these two abilities did not change significantly. Above results together suggested that different structural patterns of EEG neurofeedback targeted for characteristics of brain function of different ADHD types was necessary and feasible. In the other hand, three comprehensive quotients reflecting the respond control ability  in both types improved, except these changes were more obvious in ADHD‐CT. Taken collectively, these results indicate that different structural patterns of EEG neurofeedback targeted for different ADHD types adopted in this study can effectively improve the cognitive and executive functions of different types of ADHD patients. And the more treatment courses, the better treatment effect.This study had some limitations. The sample size of this study was comparatively small and only met the basic statistical requirements. The statistical efficiency would have improved if blind grouping was used and/or the sample size was increased appropriately. Furthermore, all subjects were from Jiaxing City. As a medium‐sized city, although Jiaxing City has a good representative, there might have regional limitations. Therefore, further studies are needed to verify the results of this study.5In summary, the results of this study indicated that different structural patterns of EEG neurofeedback targeted for ADHD‐CT and ADHD‐PI were both effective and feasible, especially in cognitive and executive functions, including attention and respond control abilities. Two treatment courses of EEG neurofeedback had a good therapeutic effect in ADHD patients, but three treatment courses were most effective. In the future, this targeted and different structural pattern multi‐course treatment of EEG neurofeedback deserves to be applied in the treatment of ADHD to achieve a better therapeutic effect.No financial or non‐financial benefits have been received or will be received from any party related directly or indirectly to the subject of this article.Concept and design: Tong‐kun Shi. Methodology: Xiao‐mei Cui and He‐dan Zhao. Data curation: Ling‐fei Yang and Zheng Wang. Writing—original draft: Feng‐hua Wang and Li‐yan Sun. Writing‐review and editing: Feng‐hua Wang and Li‐yan Sun. Project Administration: Tong‐kun Shi. Funding acquisition: Tong‐kun Shi. All authors have read and agreed to the published version of the manuscript.https://publons.com/publon/10.1002/brb3.2572The peer review history for this article is available at


Dear Editor,Dyshidrosiform bullous pemphigoid (DBP) is a rare variant of bullous pemphigoid (BP), autoimmune blistering disease mostly affecting the elderly.DBP was described by Levine et al. in 1979, as pruritic blisters on hands and feet, resembling pompholyx, often generalizing to other body sites.In pediatric population, dyshidrosiform presentation of BP is a constant initial BP manifestation in infants, while in adults the incidence of DBP ranges from 3.5% to 28%.Steroids represent the primary treatment of BP, adding steroid‐sparing agents when needed. Almost all adults and 93% of infants achieve remission.We report a case of DBP, successfully treated with intravenous immunoglobulins (IVIGs) after ineffective classical steroid therapy.3), eosinophilia , and thrombocytosis . Skin biopsy was performed for histopathological examination and direct immunofluorescence (DIF), revealing respectively multiple eosinophils in the superficial dermis and in subepidermal clefts and linear staining of IgG and C3 along the dermoepidermal junction. Enzyme‐linked immunosorbent assay  evidenced IgG anti‐BP180‐antigen (234.5 U/ml) and IgG anti‐BP230‐antigen (22.8 U/ml) .A 1‐month‐old boy who developed pruritic blisters on his hands and feet, progressively generalizing  was diagnosed. Oral betamethasone was administered (0.2 mg/kg/day), obtaining clinical remission within 1 month; it was progressively tapered. A relapse after tapering was controlled with prednisone (25 mg/day) and dapsone (50 mg/day), but weight gain and insomnia forced to taper steroids, worsening BP. IVIGs were added  with immediate clinical improvement, allowing tapering of prednisone (5 mg/day) and dapsone (25 mg/day) within 4 months, as remission was obtained after the fourth IVIGs administration. Dapsone (25 mg/day) was still administered for 1 year. During 2‐year‐follow‐up no further relapses were reported; ELISA IgG autoantibodies were negative.BP represents the most common bullous autoimmune disease in adults, whereas pediatric BP is rarer.Though no official treatment guidelines exist for IBP, steroids are administered as first‐line therapy (97% of cases), adding steroid‐sparing agents in 60% of patients. Overall, 93% obtain remission.Occasionally, IBP may be refractory to systemic corticosteroids and dapsone, as in the present case: IVIGs (2 g/kg/4w) may be of aid in these circumstances, rapidly and safely leading to clinical improvement.In literature cases of IBP treated with IVIGs are anecdotal, reporting variable dosages, (200–1000 mg/kg/day), number (1–7), duration (1–5 days) of infusion‐courses and intervals between courses (2–5 weeks). Mostly, administered doses corresponded to the IVIG regimen of adults  for autoimmune bullous diseases comprising BP.Conclusively, IVIG are a well‐tolerated, effective, broad‐spectrum therapeutic alternative for severe, therapy‐refractory immune diseases as BP.Still, in difficult‐to‐treat, fragile BP patients, as the presented one‐month‐old infant, IVIGs' safety and therapeutic potency highly favor their administration, which is mostly resolutive.Gianmaria Viglizzo, Astrid Herzum, Emanuele Cozzani, Corrado Occella, and Aurora Parodi contributed equally to the manuscript and read and approved the final version of the manuscript.The authors certify that there is no conflict of interest with any financial organization regarding the material discussed in the manuscript.The present research study complies with the guidelines for human studies and includes evidence that the research study was conducted ethically in accordance with the World Medical Association Declaration of Helsinki. Written informed consent to publish the case (including publication of images) was obtained.


Testosterone has an impact on metabolic disorders and men with type 2 diabetes mellitus (T2DM) are predisposed to hypogonadism; meanwhile, patients with T2DM have higher risk of NAFLD. Therefore, we speculate that testosterone may affect the progression of NAFLD in T2DM patients and we aim to investigate whether total testosterone is associated with NAFLD progression in men with T2DM.A cross-sectional study. A total of 1782 male participants with T2DM were enrolled from seven communities in Shanghai. Probable nonalcoholic steatohepatitis (NASH) was defined by the concurrence of NAFLD and metabolic syndrome (MetS). NAFLD fibrosis score was used to identify patients with probable advanced fibrosis. Multinomial logistic regression and ordinal logistic regression was used to measure the association of total testosterone (independent variable) and the progression category of NAFLD (dependent variable).P for trend = 0.001) and inflammatory progression of NAFLD with OR of 1 SD increment of ln (TT) 0.81 , but positively with fibrotic progression  with OR of 1 SD increment of ln (TT) 1.24 (95% CI 1.07–1.45). According to stratified analyses, for inflammatory progression, the interactions of age strata, duration of diabetes strata, and dyslipidemia status with 1 SD increment of ln (TT) were significant ; as for fibrotic progression, we found no interactions .In male, TT quartiles were negatively associated with probable NASH (Q1 vs. Q4 OR 2.07 95% CI 1.31–3.28, Different associations between TT and inflammatory and fibrotic progression of NAFLD in male were observed, suggesting different roles of TT in inflammatory and fibrotic stages of NAFLD. Nonalcoholic fatty liver disease (NAFLD) has become the most prevalent chronic liver disease not only in western countries but also in China , 2. PatiSex steroids have direct impact on both hepatic and systematic metabolism, thereby involving NAFLD pathobiology. Meanwhile, as an important organ which regulates glucolipid and sex steroid metabolism, liver has interaction effects with reproductive system, thus taking on sexual dimorphism . It has Thus, we assume that total testosterone may be associated with inflammatory and fibrotic progression of NAFLD in men with type 2 diabetes. Liver biopsies are unavailable in large epidemiology studies. Since MetS is one of the strong noninvasive NASH predictors and NASH risk increases with the increasing number of MetS components, we used MetS to define probable NASH . In ordeHence, in this large community-based study, we investigated the relationship between total testosterone and inflammatory and fibrotic progression of NAFLD assessed by aforementioned noninvasive methods in Chinese men with T2DM.www.chictr.org.cn, ChiCTR1800017573). Participants from seven communities in Huangpu and Pudong District, Shanghai, China were enrolled in this study. In each community healthcare center, we used simple random sampling to select half of the diabetic patients from the registration platform. Chinese citizens ≥18 years old who had lived in their current area for ≥6 months were included. A total of 4937 subjects with diabetes received the examination. Participants who were missing laboratory results (n = 8) or questionnaire data (n = 116) were excluded. Further, female participants (n = 2590), who had a history of excessive consumption  of pure alcohol , had self-reported viral hepatitis , were using medications associated with secondary NAFLD , and did not have ultrasound results (n = 30), were excluded. Totally, 1782 male diabetic participants were involved in the final analyses  and the minimal detectable limit for it was 0.087 nmol/L. Glycated hemoglobin (HbA1c) was measured by HPLC . Fasting plasma glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and lipid profile were performed with a Beckman Coulter AU 680 . Samples with values below the minimal detectable limit were assigned a value midway between zero and the minimal detectable limit for the analyses . The intHypertension was defined as systolic blood pressure ≥ 140 mmHg, diastolic blood pressure ≥ 90 mmHg, or self-reported previous diagnosis of hypertension by physicians. According to the modified National Cholesterol Education Program-Adult Treatment Panel III, dyslipidemia was defined as total cholesterol ≥ 6.22 mmol/L (240 mg/dL), triglycerides ≥ 2.26 mmol/L (200 mg/dL), low-density lipoprotein ≥ 4.14 mmol/L (160 mg/dL), high-density lipoprotein <1.04 mmol/L (40 mg/dL), or self-reported previous diagnosis of hyperlipidemia by physicians.Liver fat accumulation (steatosis) was detected by ultrasound  and the same sonologist , 27. In 2) + 1.13*impaired fasting glucose/diabetes  + 0.99*ST/ALT ratio − 0.013*platelet (*109/L) − 0.66*albumin (g/dL) [The NFS was used to classified NAFLD patients into different risk groups of having advanced fibrosis . The forn (g/dL) . A scoren (g/dL) .p value <0.05 indicated significance (two-sided). The concentration of total testosterone was logarithmically transformed to achieve a normal distribution if necessary.Data analyses were performed using IBM SPSS Statistics, Version 26 . Continuous variables are shown as mean ± SD while categorical variables as percentages (%). A Total testosterone was divided into quartiles. The first quartile of total testosterone (Q1) represents the lowest one and the fourth quartile (Q4) represents the highest. Multinomial logistic regression was used to measure the association of testosterone (independent variable) and the progression category of NAFLD (dependent variable). Odds ratios (ORs) with 95% confidence interval (CI) were calculated by ordinal logistic regression to measure the association between the quartiles of testosterone, NAFLD inflammatory progression, and fibrotic progression. The model was adjusted for age, duration of diabetes, current smoking, waist circumference, dyslipidemia, and hypertension.In subgroup analyses, logistic regression was used to explore the relationship between 1-SD increment of ln (TT) and inflammatory and fibrotic progression of NAFLD stratified by age (≥60 years or <60 years), waist circumference (≥90 cm or <90 cm), duration of diabetes (≥10 years or <10 years), hypertension (yes or no), and dyslipidemia (yes or no) and to examine the potential interaction association.General clinical and demographic characteristics of the study population by quartiles of TT are shown in Table P for trend <0.001) and probable NASH  after adjustment for age, duration of diabetes, waist circumference, current smoking, hypertension, and dyslipidemia, and in ordinal regression, TT was also negatively associated with the inflammatory progression of NAFLD, with OR of 1 SD increment of ln (TT) 0.81 .Table P for trend = 0.697); however, unlike their negative association with the inflammatory progression, TT quartiles were positively associated with the fibrotic progression  after adjustment for age, duration of diabetes, waist circumference, current smoking, hypertension, and dyslipidemia, with OR of 1 SD increment of ln (TT) 1.24 (95% CI 1.07–1.45).Table P < 0.05) and the interaction was significant in age strata, duration of diabetes strata, and dyslipidemia status . As for fibrotic progression, its associations with TT were not significant in age strata (< 60 years), waist circumference strata (< 90 cm), hypertension status (no) and dyslipidemia status , and the interaction was not significant .To investigate the association of inflammatory and fibrotic progression of NAFLD with a 1 SD increment of ln (TT) in subgroup of strata variables in Chinese diabetic men, we performed stratified analyses Figs. , 3 We foIn this population-based study including 1782 male patients with T2DM, we found that TT of male patients was negatively associated with probable NASH and NAFLD inflammatory progression but positively with fibrotic progression. Our results indicate that TT may play significant while opposite roles in inflammatory and fibrotic stages of NAFLD in Chinese type 2 diabetic men.The association between low level of testosterone and NASH has been found in several studies. Epidemiological studies have reported the negative association between testosterone and liver steatosis or NASH in general male , 31, 32;Men are predisposed to advanced hepatic fibrosis relative to women. Besides excessive alcohol consumption  and highP for interaction <0.05). Meanwhile, we found no interactions between fibrosis progression and testosterone in any strata variables  and the association of TT and fibrosis progression was not significant in NAFLD patient aged <60 years or with dyslipidemia (both P ≥ 0.05). Therefore, TRT may have greater advantage for men aged <60 years, with shorter duration of diabetes or with dyslipidemia.In subgroup analysis, we found that protective effects of TT on inflammatory progression of men with T2DM in age strata , duration of diabetes strata , and dyslipidemia status  were relatively more significant  of Chinese decreased slightly compared with other population. Also, diminished clearance of sex hormone binding globulin (SHBG) may affect the level of TT .Although the current study has some strengths, including the large sample of community-dwelling participants, novelty, and strong quality control, there are also some limitations. First, since it is a cross-sectional study, the causal relationship between TT and NAFLD progression cannot be clarified. Second, we used NFS to assess probable advanced fibrosis in patients with NAFLD and type 2 diabetes and defined probable NASH as concurrence of NAFLD and MetS. Liver biopsy, albeit the gold standard to assess liver inflammation and fibrosis, is an invasive examination, so it could not be used in large-scale epidemiological studies. NFS can predict presence of advanced liver fibrosis with high sensitivity and specificity. Lastly, we did not measure sex hormone binding protein and free testosterone, so we cannot explore the relationship between SHBG or free testosterone and NAFLD progression. However, TT was closely related to free testosterone in the elder  and the In conclusion, our results indicate that in male patients with T2DM, testosterone may play significant but opposite roles in inflammatory and fibrotic progression of NAFLD. Hence, understanding androgen hormone changes during T2DM and differentiating the stages of NAFLD may help clinicians personalize treatment strategy for male NAFLD patients with T2DM. Further studies are warranted to identify the causality and potential mechanisms of testosterone in NAFLD inflammatory and fibrotic progression.


The study aimed to examine parental, longitudinal and current associative factors for atopic dermatitis (AD) and to compare those to other atopic diseases in 46‐year‐old adults.n = 5373) were performed for birch, timothy, cat, and house dust mite at age 46.Questionnaire data from the Northern Finland Birth Cohort 1966 study were used. To analyze allergic sensitization, skin prick tests  and paternal allergy , sensitization to any of the four tested aeroallergens  as well as polysensitization  were associated with current AD. Living on a farm in infancy was negatively associated with allergic rhinitis, allergic conjunctivitis, and atopic multimorbidity. Current AD  and all atopic diseases associated with indoor air related symptoms. Current AD associated with other atopic diseases, most strongly with allergic rhinitis .Current AD in a 46‐year‐old general population occurred frequently with allergic rhinitis, allergic conjunctivitis, and asthma in the Northern Finland Birth Cohort study 1966. Parental allergy and sensitization to common aeroallergens were found as shared associative factors for AD, allergic rhinitis, allergic conjunctivitis, and asthma. AD and other atopic diseases associated with symptoms related to poor indoor air quality. In daily practice, it is important to take these comorbidities into consideration when treating patients with AD. Parental atopic diseases have shown to increase the risk of atopic morbidity in the offspring.A few studies have investigated the risk factors in AD alongside AR, AC and asthma in adults, and the findings vary between studies.22.1Our study used data from the Northern Finland Birth Cohort 1966 (NFBC 1966), which is a longitudinal research program in the two northernmost provinces in Finland. Initially, all 12,058 children whose expected date of birth fell in the year 1966 were included in the NFBC 1966. Health questionnaires and clinical examinations have been conducted regularly on the whole cohort since birth.2.2SPT were performed at age 46 and the procedure has been described in detail in a previous article.Data on factors  with 95% confidence intervals (CI) for atopic diseases were assessed using a binary logistic regression model. Adjusted ORs were calculated using a binary logistic regression model, and all above‐mentioned potential confounding factors were entered in the model as potential predictors for atopic diseases. All significance tests were two‐tailed, and values of p < 0.05 were considered statistically significant. The statistical analyses were conducted using the R software package version 4.1.2 (https://cran.rstudio.com).Means and standard deviations (SD) or proportions were calculated and presented for each variable. The statistical significance of the mean differences and proportions of categorical variables between the atopic disease groups were estimated by a Mann–Whitney U‐test and a 33.1n = 10,321); 5861 participants attended the clinical examination. SPTs were performed in 5714 (55.4% of invited) participants. Because of invalid data, 331 participants were excluded from the analysis, and 10 participants did not give their consent to data processing. Complete SPT data on all allergens were available for 5373 participants (2394 [44.6%] were men) who comprised the final study population.At 46 years, invitations and questionnaires were sent to every living member of the cohort whose address was known n = 0,321; 583.2Any current atopic disease was reported by 2101 participants (39.1%). Current AD, lifetime AD, AR, AC, asthma, and multimorbidity occurred in 7.3%, 25.4%, 32.8%, 28.1%, 9.2%, and 24.9%, respectively. Women reported significantly more atopic diseases than men (Table 3.33.3.1Allergy of either parent associated with AD (both lifetime and current) Figures  and 2, A3.3.2In the multivariate model, female gender was associated with current AD , lifetime AD  Figures .3.3.3Living on a farm in infancy was not associated with current AD, lifetime AD Figures  and 2, o3.3.4Smoking was not associated with atopic diseases. Having pets at age 46 negatively associated with current AD Figure , AC and 3.3.5Monosensitization at age 46 was associated with current AD  Figure . For cur3.4n = 355), another atopic disease was reported by 272 participants (76.6%). The most common combination was AD with AR and AC (n = 130/36.6%). Sixty‐four (18%) participants had all four atopic diseases , AC  and asthma .  found that any positive SPT to aeroallergens increased the risk of any eczema,n = 1172), atopic sensitization was not a risk factor for current eczema.n = 1805 adults) did not find an association between sensitization or polysensitization and AD as a single disease in their study in Poland.n = 2096), sensitization to any of the 9 pollen and HDM allergens was associated with both eczema and “eczema with asthma and/or hay fever.”n = 52,976), in which sensitization to common inhalant allergens was a risk factor for AR, AC, and asthma.Aeroallergen sensitization and polysensitization associated with AD in the current study. In previous studies, the association between sensitization and AD in adults has varied.n = 52,976), upbringing on a farm with livestock compared to other levels of urbanization had a protective effect on allergic rhino‐conjunctivitis,Living on a farm in infancy did not protect from current or lifetime AD or asthma. However, the protective effect on AR, AC, and multimorbidity was seen in our study. A study conducted in Sweden found a protective effect of farm upbringing on lifetime eczema, but not on current eczema.Current residence or current farming were not associated with atopic diseases in our study. Having pets at age 46 negatively associated with AD, AC, and atopic multimorbidity, but not with asthma or AR. This may come from the fact that those who already have atopic diseases, especially the multimorbid phenotype, might not acquire pets in adulthood as readily.Smoking was not associated with current or lifetime AD in our study. Previous studies on the relation between AD and smoking suggest a positive association: A systematic review and meta‐analysis of 86 studies with 680,176 patients  found that AD in both adults and children associated with active smoking and exposure to passive smoke, but not maternal smoking during pregnancy.In the present study, symptoms related to indoor air were significantly associated with AD, AR, AC, asthma, and atopic multimorbidity, as in previous studiesIn this cohort of middle‐aged adults, 7.3% reported current AD and 25.4% reported lifetime AD. According to a cross‐sectional multicenter study, approximately 4.4% of the adults in the EU and 4.9% in the United States have AD.AD associated with all the other atopic diseases in the current study, most strongly with AR. In previous studies, AD in adults has clearly associated with hay fever and asthma,The strengths of the study are the large and unselected study population which represents well the middle‐aged adult population, both men and women. Compared to the Danish study,In conclusion, AD had shared associative factors with AR, AC, and asthma, including parental allergy and sensitization to aeroallergens. In addition, a common association was found with indoor air related symptoms while current residence or farming were not associated with any of the atopic diseases. Women reported atopic morbidity more often than men. Differences between associative factors were also found as living on a farm in infancy did not protect from AD or asthma, but was negatively associated with AR, AC, and multimorbidity. Patients with AD commonly had atopic comorbidities and suffered from poor indoor air, which should be remembered as factors that could contribute to the exacerbation of AD when treating AD patients.Anna K. Haarala: Conceptualization; visualization; writing – original draft. Suvi‐Päivikki Sinikumpu: Conceptualization; investigation; project administration; supervision; writing – review & editing. Jari Jokelainen: Data curation; formal analysis; methodology; software; visualization. Juha Pekkanen: Conceptualization; resources; supervision. Laura Huilaja: Conceptualization; investigation; project administration; supervision; visualization; writing – review & editing. All authors have read and approved the final version of the manuscript.The authors declare no conflict of interest.The Ethical Committee of the Northern Ostrobothnia Hospital District approved the study (§94/2011), which was performed according to the Helsinki Declaration of 1983. Written informed consent for scientific purposes was received from all participants.The lead author Suvi‐Päivikki Sinikumpu affirms that this manuscript is an honest, accurate, and transparent account of the study being reported; that no important aspects of the study have been omitted; and that any discrepancies from the study as planned  have been explained.Supporting information.Click here for additional data file.


Viral load (VL) testing is still challenging to monitor treatment responses of antiretroviral therapy (ART) for HIV treatment programme in Asia. We assessed the association between routine VL testing and virological failure (VF) and determine factors associated with switching to second‐line regimen.Among 21 sites from the TREAT Asia HIV Observational Database (TAHOD), people living with HIV (PLHIV) aged ≥18 years initiating ART from 2003 to 2021 were included. We calculated the average number of VL tests per patient per year between the date of ART initiation and the most recent visit. If the median average number of VL tests was ≥ 0.80 per patient per year, the site was classified as a routine VL site. A site with a median < 0.80 was classified into the non‐routine VL sites. VF was defined as VL ≥1000 copies/ml during first‐line therapy. Factors associated with VF were analysed using generalized estimating equations with Poisson distribution.3, p <0.001). Overall, 1021 subsequent VF at a rate of 2.15 (95% CI 2.02–2.29) per 100 person‐years (PY). VF was more frequent at non‐routine VL sites (adjusted incidence rate ratio 2.85 [95% CI 2.27–3.59]) compared to routine VL sites. Other factors associated with an increased rate of VF were age <50 years and CD4 count <350 cells/mm3. A total of 817 (13%) patients switched to second‐line regimen at a rate of 1.44 (95% CI 1.35–1.54) per 100 PY. PLHIV at routine VL monitoring sites were at higher risk of switching than those at non‐routine VL sites (adjusted sub‐hazard ratio 1.78 95% CI [1.17–2.71]).Of 6277 PLHIV starting ART after 2003, 3030 (48%) were from 11 routine VL testing sites and 3247 (52%) were from 10 non‐routine VL testing sites. The median follow‐up was 9 years (IQR 5–13). The median age was 35 (30–42) years; 68% were male and 5729 (91%) started non‐nucleoside reverse‐transcriptase inhibitor‐based regimen. The median pre‐ART CD4 count in PLHIV from routine VL sites was lower compared to non‐routine VL sites (144 vs. 156 cells/mmPLHIV from non‐routine VL sites had a higher incidence of persistent VF and a low switching regimen rate, reflecting possible under‐utilized VL testing. There were almost 38 million PLHIV globally at the end of 2020, and 74% of adults living with HIV had access to ART . IncreasThe TREAT Asia HIV Observational Database (TAHOD) of IeDEA Asia‐Pacific  is a longitudinal observational cohort study, which monitors long‐term treatment outcomes in PLHIV in Asia. VL monitoring varies across TAHOD sites. In this study, we aimed to assess the association between routine VL testing (routine VL) and VF during first‐line therapy and investigate factors associated with switching to a second‐line regimen.22.1As TAHOD is an adult HIV observational cohort, our analysis included PLHIV aged ≥18 years at ART initiation, from January 2003 to September 2021. There were 21 sites from Cambodia, China, Hong Kong, India, Indonesia, Japan, Malaysia, Philippines, Singapore, South Korea, Taiwan, Thailand and Vietnam. The TAHOD cohort contributes to the IeDEA global consortium and has been described previously , 13, 14.2.2We classified sites according to whether they performed routine VL testing, that is routine VL versus non‐routine VL. We defined routine VL by calculating the average number of VL tests for each patient between ART initiation and the last visit date. If the median of the average number of VL tests was above 0.80 per patient per year, the site was classified as routine VL site. If the median was less than 0.80, the site was classified as a non‐routine VL site. We utilized 0.80 as the cut of as the WHO's recommendation for VL testing was done once a year. As there would have been some variability in appointment scheduling and attendance, we utilized 0.80 as the cut‐off to allow some flexibility associated with the WHO's guidelines , 14, 15.2.3Baseline characteristics, including demographics, age, income country level, HIV exposure, hepatitis C co‐infection, prior AIDS‐defining illness, year of ART initiation, first‐line ART regimen and pre‐ART CD4 count level, were summarized using descriptive statistics for routine VL and non‐routine VL sites. The comparison of characteristics between routine VL and non‐routine VL sites was performed using Pearson's Chi‐square or Fisher's exact test, as appropriate, for categorical data and Wilcoxon rank sum test for continuous data. The outcomes rate was calculated by dividing the total number of outcomes by the total number of person‐years (PY) of follow‐up and expressed as a rate per 100 PY. The associations between routine VL status including with other predictors and VF were presented as incidence rate ratios (IRR) by using generalized estimating equation (GEE) with log link function (Poisson distribution) and exchangeable correlation structure. We used Fine and Gray's competing risk regression  to assesp <0.10 in the univariate analysis were considered for inclusion in multivariate models. Statistical significance was identified using a two‐sided p value less than 0.05. Statistical analysis was performed with SAS version 9.4  and with Stata version 16 .Variables with 33.1n = 464, 4%), (2) starting ART with only mono/dual or combination of NNRTI and PI drugs , (3) initiating ART before 2003 , (4) age <18 years at ART initiation , (5) no sex information , (6) assumed to be treatment experienced due to VL <50 copies/ml at baseline  and (7) no VL measurement after ART initiation   not having yet initiated ART (p <0.001), and the median age at ART initiation was 35  years. We found that 91% started with NNRTI‐based ART  in lower middle‐income; 95%  in upper middle‐income; and 60%  in high‐income countries) and 9% started with PI‐based ART  in lower middle‐income; 5%  in upper middle‐income; and 40%  in high‐income countries). Most patients had a heterosexual mode of HIV exposure. Thirty‐six percent had a prior AIDS diagnosis. The median pre‐ART CD4 count in patients from routine VL sites was higher compared to non‐routine VL sites . Forty percent had pre‐ART CD4 < 200 cells/mm3 (42% in non‐routine VL vs. 38% in routine VL). We found that 42% had pre‐ART CD4 < 200 cells/mm3 with advanced HIV disease stage. There was no significant difference in the proportion of pre‐ART VL ≥1000 copies/ml between non‐routine VL and routine VL sites  patients from 11 routine VL testing sites which consisted of five sites from high‐income countries and six sites from upper‐middle‐income countries, and 3247 (52%) patients from 10 non‐routine VL sites (eight in lower‐middle‐income countries and two in upper‐middle‐income countries). The majority were male  time/year. For routine VL sites, it was 1.31 (1.17–2.00) time/year, and for non‐routine VL sites, it was 0.15 (0.10–0.66) time/year throughout the study period. The median duration on ART during the first‐line regimen for non‐routine VL sites was 9 (IQR 6–13) years with a total of follow‐up 29,671 PYs compared to 9 (IQR 5–13) years with a total of follow‐up 27,015 PYs for routine VL sites. For non‐routine VL sites, 849 (14%) patients were LTFU  and 76 (1%) died . The number of LTFU in routine VL was 524 (8%) with 1.94 per 100 PYs; (95% CI 1.78–2.11) and 136 (2%) died .3.2Overall, 1021 subsequent VF at a rate of 2.15 (95% CI 2.02–2.29) per 100 PYs. We found that the proportion of VF on first‐line ART was 16% for non‐routine VL and routine VL sites. The crude VF rate of non‐routine VL (2.24 [95% CI 2.06–2.44] per 100 PYs) was higher than the crude rate of routine VL sites (2.06 [95% CI 1.89–2.26] per 100 PYs). Figure 3.33  and unknown CD4 count  were associated with a higher rate of VF when compared with those who had CD4 count ≥350 cells/mm3. Other factors associated with an increased rate of VF were HIV exposure and income country level.Table 3.4p‐value <0.001). The cumulative incidence of switching regimens in routine VL sites at 1, 2 and 3 years were 1.20% (95% CI 0.9–1.7%), 4.0% (95% CI 3.3–4.7%) and 6.2% (95% CI 5.4–7.1%), respectively. For non‐routine VL sites, the cumulative incidence of switching regimens at 1, 2 and 3 years were 0.2% (95% CI 0.1–0.5%), 1.3% (95% CI 1.0–1.8%) and 2.7% (95% CI 2.2–3.3%), respectively.A total of 817 (13%) patients switched to a second‐line ART regimen according to our definition. Of these, 481 (8%) patients switched due to VF, and 336 (5%) patients switched due to other reasons without evidence of VF. The median duration from VF to switching second‐line ART for PLHIV having VL results was 6 (2–23) months (7 [2–25] months in non‐routine VL sites and 5 [1–21] months in routine VL sites). In addition, the proportion of those who switched from routine VL sites was higher than non‐routine VL sites (18% vs. 9%). The overall rate of switching to second‐line ART was 1.44 (95% CI 1.35–1.54) per 100 PYs. The rate for routine VL sites was 1.97 (95% CI 1.81–2.15) per 100 PYs, which was higher than the rate for non‐routine VL sites  was associated with a higher rate of switching regimen when compared with those who had CD4 count ≥500 cells/mm3. Other factors associated with increased risk of switching were HIV exposure, year of ART initiation and PI‐based regimen initiation.In the adjusted competing risk regression Table , patient4VL testing can be used as a tool to monitor HIV treatment responses and inform decisions for treatment switching. In our study, the overall crude rate of the first VF after ART initiation at non‐routine VL sites was slightly higher than at the routine sites. The rate of subsequent VF during first‐line ART was higher in PLHIV at non‐routine VL sites compared with PLHIV at routine VL sites. As expected, the rate of switching to second‐line in PLHIV at non‐routine VL sites was lower than PLHIV at routine VL sites due to our definition of treatment failure, including virological, immunological and clinical failure. The median overall VL testing frequency for the cohort was 0.75 (0.15–1.24) time/year. The VL testing frequency at routine VL sites was higher than at non‐routine VL sites. We did not utilize a second confirmatory VL testing to define VF in order to capture all VF that occurred in our cohort. This finding indicated immediately switching to second‐line ART after detecting VF for routine VL sites were better clinical management and HIV treatment. Compared to non‐routine VL sites, the rate of switching regimens at routine VL was about two times by 3 years of first‐line therapy higher in PLHIV at routine VL sites. Moreover, younger aged < 25 years were at higher risk of treatment failure and switching to second‐line ART compared to those aged ≥ 25 years old in our study who need to concern for long‐term treatment outcome in this age group.The implementation of routine VL testing is still a major challenge for the Asia‐Pacific region since the WHO's recommendations in 2017 , Uganda We also found that PLHIV from routine VL sites had a higher chance of switching to second‐line ART compared to non‐routine VL sites. This suggests that routine VL sites were able to identify treatment failure and switch patients promptly to second‐line ART.n = 84) switched to DTG as a second‐line regimen during the early period of WHO's recommendation for DTG use. Most of these PLHIV were from upper middle‐ and high‐income countries. The majority of PLHIV in our study have been using NNRTI‐based ART. There is a recommendation to switch to DTG in combination with an optimized NRTIs backbone as the preferred second‐line regimen [Prior studies have indicated that scaling up routine VL testing can lead to earlier detection of treatment failure, which allows for immediate switches of ART regimens . In reso regimen . This isOur study has strengths and limitations. One of the strengths is to indicate the IRR of subsequent VF between non‐routine VL and routine VL testing sites after adjusting with other factors during first‐line therapy. There are several limitations to our study. First, we defined VF according to a single VL ≥1000 copies/ml without secondary confirmatory testing. This may overestimate our VF proportions. However, to accommodate sites that do not perform routine VL testing or sites that do not perform confirmatory testing, we believe our definition of VF allowed us to identify VF in our cohort settings appropriately. Second, we did not assess treatment failures associated with routine and non‐routine CD4 testing. This would provide an alternative tool to identify risks in treatment outcomes for sites without routine VL testing. Third, as TAHOD patients were selected to be enrolled based on the likelihood of remaining in care, our results are not generalizable to the wider PLHIV population. Lastly, only a small proportion of PLHIV in our study switched to DTG. We believe that there could be a delay in the application of WHO's recommended HIV treatment policies in lower/upper middle‐income countries due to the varying costs and standard of care , 30. The5In conclusion, PLHIV from non‐routine VL sites had a higher incidence of persistent VF and low switching regimen rate with delay detecting VF, reflecting possible under‐utilized VL testing within these sites. However, the expanding access to routine VL should be continued for HIV treatment monitoring and more common to consider switching ART earlier. Findings suggest that different VL monitoring strategies may have an impact on the time of treatment failure and switch to second‐line, as well as the development of drug resistance in long‐term treatment.The authors declare they have no competing interests.ST designed the study, drafted the initial manuscript, reviewed and critically revised and approved the final manuscript as submitted. AJ was responsible for data aggregation and data management. ST conducted the analysis. AJ and ML advised on the analysis, revised and approved the final manuscript. All authors critically reviewed the manuscript and approved the manuscript for submission.PS Ly*, V Khol, National Center for HIV/AIDS, Dermatology & STDs, Phnom Penh, Cambodia; FJ Zhang*, HX Zhao, N Han, Beijing Ditan Hospital, Capital Medical University, Beijing, China; MP Lee*, PCK Li, TS Kwong, TH Li, Queen Elizabeth Hospital, Hong Kong SAR; N Kumarasamy*, C Ezhilarasi, Chennai Antiviral Research and Treatment Clinical Research Site (CART CRS), VHS‐Infectious Diseases Medical Centre, VHS, Chennai, India; S Pujari*, K Joshi, S Gaikwad, A Chitalikar, Institute of Infectious Diseases, Pune, India; RT Borse*, V Mave, I Marbaniang, S Nimkar, BJ Government Medical College and Sassoon General Hospital, Pune, India; TP Merati*, IKA Somia, AAS Sawitri, F Yuliana, Faculty of Medicine Udayana University & Sanglah Hospital, Bali, Indonesia; E Yunihastuti*, A Widhani, S Maria, TH Karjadi, Faculty of Medicine Universitas Indonesia ‐ Dr. Cipto Mangunkusumo General Hospital, Jakarta, Indonesia; J Tanuma*, S Oka, T Nishijima, National Center for Global Health and Medicine, Tokyo, Japan; JY Choi*, Na S, JM Kim, Division of Infectious Diseases, Department of Internal Medicine, Yonsei University College of Medicine, Seoul, South Korea; YM Gani*, NB Rudi, Hospital Sungai Buloh, Sungai Buloh, Malaysia; I Azwa*, A Kamarulzaman, SF Syed Omar, S Ponnampalavanar, University Malaya Medical Centre, Kuala Lumpur, Malaysia; R Ditangco*, MK Pasayan, ML Mationg, Research Institute for Tropical Medicine, Muntinlupa City, Philippines; YJ Chan*, WW Ku, PC Wu, E Ke, Taipei Veterans General Hospital, Taipei, Taiwan; OT Ng*, PL Lim, LS Lee, T Yap, Tan Tock Seng Hospital, National Centre for Infectious Diseases, Singapore  National Medical Research Council (NMRC) Clinician Scientist Award (MOH‐000276). Any opinions, findings and conclusions or recommendations expressed in this material are those of the author(s) and do not reflect the views of MOH/NMRC; A Avihingsanon*, S Gatechompol, P Phanuphak, C Phadungphon, HIV‐NAT/Thai Red Cross AIDS Research Centre, Bangkok, Thailand; S Kiertiburanakul*, A Phuphuakrat, L Chumla, N Sanmeema, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Bangkok, Thailand; R Chaiwarith*, T Sirisanthana, J Praparattanapan, K Nuket, Chiang Mai University ‐ Research Institute for Health Sciences, Chiang Mai, Thailand; S Khusuwan*, P Payoong, P Kantipong, P Kambua, Chiangrai Prachanukroh Hospital, Chiang Rai, Thailand; TN Pham*, KV Nguyen, DTH Nguyen, DT Nguyen, National Hospital for Tropical Diseases, Hanoi, Vietnam; CD Do*, AV Ngo, LT Nguyen, Bach Mai Hospital, Hanoi, Vietnam; AH Sohn*, JL Ross*, B Petersen, TREAT Asia, amfAR ‐ The Foundation for AIDS Research, Bangkok, Thailand; MG Law*, A Jiamsakul*, D Rupasinghe, The Kirby Institute, UNSW Sydney, NSW, Australia. * TAHOD Steering Committee member.The Kirby Institute is funded by the Australian Government Department of Health and Ageing, and is affiliated with the Faculty of Medicine, UNSW Sydney. International SciKU Branding (ISB) is funded by the Faculty of Science, Kasetsart University.Data were presented in part at the 4th Asia Pacific AIDS & Co‐infections Conference, June 27–29, 2019, HongKong. The content of this publication is solely the responsibility of the authors and does not necessarily represent the official views of any of the governments or institutions mentioned above.


We investigated the associations between fasting glucose level ranges with lifestyle factors and metabolic profiles among adults without previous diagnosis of diabetes.We analyzed 13 625 adults without previous diagnosis of diabetes from the Korea National Health and Nutrition Examination Survey during 2016 to 2018. We categorized fasting glucose levels (mg/dl) as follows: <90, 90 to 99, 100 to 109, 110 to 124, and ≥125. We evaluated trends in the proportions of individuals with obesity, abdominal, obesity, current smoking, heavy drinking, and low physical activity according to these categories, and the odds for uncontrolled blood pressure (BP), low‐density lipoprotein cholesterol (LDL‐C), triglycerides (TG), and high‐density lipoprotein cholesterol (HDL‐C) for each fasting glucose level compared to a fasting glucose level of <90 mg/dl.P for trend <.05). The odds for BP ≥140/90 mm Hg, TG ≥150 mg/dl, HDL‐C < 40 mg/dl in men, and HDL‐C < 50 mg/dl in women increased with increasing fasting glucose levels; however, the odds for LDL‐C ≥ 130 mg/dl increased with increasing fasting glucose levels only in women. The increases in odds for uncontrolled BP and lipid profiles were mostly observed for fasting glucose levels ≥90 mg/dl.The proportions of individuals with obesity, abdominal obesity, and heavy drinking increased according to fasting glucose level (Efforts are needed to prevent increased fasting glucose levels, as higher levels, even within normal range, were associated with poor metabolic profiles. HighlightsThe proportions of individuals with obesity, abdominal obesity, and heavy drinking increased according to fasting glucose level.The risk for uncontrolled blood pressure and lipid profiles increased with increasing fasting glucose levels.The increases in risks for uncontrolled blood pressure and lipid profiles were mostly observed for fasting glucose levels ≥90 mg/dl.Efforts are needed to prevent increased fasting glucose levels, as higher levels, even within normal range, were associated with poor metabolic profiles. Previous studies have shown that poor lifestyle behaviors such as heavy drinking, cigarette smoking, and low physical activity increase the risk for IFG, which, in turn, increases the risks for metabolic risk factors such as high blood pressure and dyslipidemia.Therefore, this study investigated the association between multiple lifestyle risk factors, fasting glucose levels, and metabolic parameters in a nationwide representative sample of adults without previous diagnosis of diabetes. We evaluated the trends of lifestyle factors and metabolic parameters according to fasting glucose levels to determine whether higher fasting glucose levels were associated with poor lifestyle behaviors and result in worse metabolic profiles in individuals who are not treated for diabetes.22.1n = 1805), taking oral hypoglycemic agents (n = 1666), taking insulin (n = 135), and with missing values for the fasting glucose levels (n = 1304). We additionally excluded pregnant women (n = 66) and those with chronic debilitating diseases such as cancer (n = 394), liver cirrhosis (n = 49), and chronic kidney disease (n = 51). Finally, the analysis in this study included 13 625 participants.We used data from the seventh Korea National Health and Nutrition Examination Survey (KNHANES) (2016‐2018). The KNHANES is an annual nationwide representative cross‐sectional survey designed by the Korea Centers for Disease Control and Prevention (KCDC). The survey applies complex, stratified, clustered, and multistage probability sampling based on age, sex, and geographic areas. Further information about the study design and methodology has been reported previously.2.2Demographic characteristics including age, sex, household income, and educational level were collected. Household income was divided into quartiles, with the lowest quartile defined as low income. Educational level was categorized as <12 or ≥12 years. Lifestyle factors including smoking status, alcohol consumption, and physical activity were also collected. Smoking status was categorized into either current smoking or nonsmoking. Alcohol consumption was categorized according to the definitions of the National Institute on Alcohol Abuse and Alcoholism,2). Waist circumference was measured to the nearest 0.1 cm at the midpoint between the lowest rib and the top of the iliac crest. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured according to standardized methods using a sphygmomanometer while the participants were sitting. Blood pressure was measured three times at 5‐minutes intervals, with the average of the second and third measurements used in the analysis. Blood samples were collected after fasting for at least 8 hours, and biochemical values, including levels of fasting glucose, HbA1c, total cholesterol (TC), low‐density lipoprotein cholesterol (LDL‐C), triglycerides (TG), and high‐density lipoprotein cholesterol (HDL‐C), were evaluated in a certified laboratory .Participant height and weight were measured using standardized techniques and equipment. Height was measured to the nearest 0.1 cm in the erect position using a portable anthropometry device. Weight was measured to the nearest 0.1 kg on a balanced scale with the participants wearing light clothing. Body mass index (BMI) was calculated by dividing the body weight by the square of height  and 95% confidence intervals (CIs) for obesity, abdominal obesity, and each lifestyle factor were calculated after adjusting for age and potential confounders. The associations between fasting glucose levels and the control of blood pressure and cholesterol levels were evaluated by multivariate logistic regression analysis. We presented age‐adjusted ORs with 95% CIs and multivariable adjusted ORs and 95% CIs for uncontrolled blood pressure and uncontrolled LDL‐C, TG, and HDL‐C after adjusting for age, income, education, obesity, abdominal obesity, current smoking, heavy drinking, and low physical activity. Furthermore, we plotted a restricted cubic spline of ORs for uncontrolled blood pressure and uncontrolled LDL‐C, TG, and HDL‐C using fasting glucose as continuous variables to examine the association between fasting glucose levels and poor metabolic parameters. Analyses were separately performed for men and women. Analyses were conducted with IBM SPSS Statistics for Windows version 23.0 , and restricted cubic splines were plotted using rms package in R software version 3.6.2  with two‐tailed P values <.05 considered statistically significant.All analyses were performed after accounting for the complex sample design, stratification, clustering, and sample weights. We presented demographic characteristics, anthropometric variables, and laboratory variables of the study participants according to their fasting glucose levels. Mean and SE were presented for continuous variables, and unweighted numbers and weighted percentages were presented for categorical variables. To compare the prevalence over fasting glucose level groups, we calculated age‐adjusted prevalence of obesity, abdominal obesity, lifestyle factors, and metabolic parameters using direct standardization by applying standardized prevalence for the Korean population in 2005 as reference. 33.1P for trend <.001). The proportion of participants with low income and participants with ≤12 years of education increased according to fasting glucose levels in both men and women (P for trend <.001). As fasting glucose levels increased, height and HDL‐C decreased and weight, BMI, waist circumference, SBP, DBP, HbA1c, TC, and TG increased in both and women (P for trend <.05). The LDL‐C levels increased according to fasting glucose levels in women (P for trend <.001); however, we observed no significant association between LDL‐C and fasting glucose levels in men.Table 3.2P for trend <.05). We observed no significant association between smoking status and fasting glucose levels. The age‐standardized proportions of participants with low physical activity increased according to fasting glucose levels in men (P for trend <.001). In the multivariate model, the ORs and 95% CIs for obesity, abdominal obesity, and heavy drinking increased according to fasting glucose levels in both men and women (P for trend <.05). Compared to fasting glucose levels of <90 mg/dl, the odds for obesity and abdominal obesity increased from fasting glucose levels of 90 to 99 mg/dl. The odds for heavy drinking increased from fasting glucose levels of 90 to 99 mg/dl in men and 110 to 124 mg/dl in women. We observed no significant association between current smoking, low physical activity, and fasting glucose levels.Table 3.3P for trend <.001). As for the control of cholesterol levels, the age‐standardized proportions of participants with TG ≥ 150 mg/dl and HDL‐C < 40 mg/dl increased according to the fasting glucose levels in men, and the age‐standardized proportions of participants with LDL‐C ≥ 130 mg/dl, TG ≥ 150 mg/dl, and HDL‐C <50 mg/dl increased according to the fasting glucose levels in women (P for trend <.001). In the multivariate model, the ORs and 95% CIs for BP ≥140/90 mm Hg increased as fasting glucose levels increased in both men and women (P for trend <.001). Compared to fasting glucose levels of <90 mg/dl, the increase was significant from fasting glucose levels of 90 to 99 mg/dl. Furthermore, as for control of cholesterol levels among men, the ORs and 95% CIs for TG ≥150 mg/dl and HDL < 40 mg/dl increased according to fasting glucose levels (P for trend <.05). Compared to fasting glucose levels of <90 mg/dl, the odds for TG ≥ 150 mg/dl increased from fasting glucose levels of 100 to 109 mg/dl whereas that for HDL < 40 mg/dl increased only at fasting glucose levels of ≥125 mg/dl. In women, the ORs and 95% CIs for LDL‐C ≥ 130 mg/dl, TG ≥150 mg/dl and HDL‐C < 50 mg/dl all increased according to fasting glucose levels (P for trend <.05). Compared to fasting glucose levels of <90 mg/dl, the odds for LDL‐C ≥ 130 mg/dl increased from fasting glucose levels of 100 to 109 mg/dl whereas those for TG ≥ 150 mg/dl and HDL‐C < 50 mg/dl increased from fasting glucose levels of 90 to 99 mg/dl.Table Figure 4In this nationwide cross‐sectional study, obesity, abdominal obesity, and heavy drinking were modifiable lifestyle risk factors that were associated with higher fasting glucose levels in individuals without previous diagnosis of diabetes. Higher fasting glucose levels were associated with poor control of blood pressure, TGs, and HDL‐C in both men and women. Furthermore, higher fasting glucose levels were associated with poor LDL‐C in women. Notably, the odds for poor control of these metabolic profiles mostly increased from the fasting glucose ranges of 90 to 99 mg/dl and above.Higher fasting glucose levels in individuals without previous diagnosis of diabetes were associated with poor blood pressure control in our study. The association between diabetes or IFG and hypertension has been well established; however, to our knowledge, only two studies have shown a linear association between different ranges of fasting glucose and blood pressure.Higher fasting glucose levels were associated with uncontrolled lipid profiles in our analysis; specifically, uncontrolled TG and HDL‐C in men and uncontrolled LDL‐C, TG, and HDL‐C in women. Previous studies also demonstrated the associations between IFG and poor lipid profiles.Among modifiable risk factors for diabetes, obesity, abdominal obesity, and heavy drinking were positively correlated with fasting glucose levels. Obesity or abdominal obesity often coexists with diabetes as they are linked with defective insulin secretion and insulin resistance.The relationship between alcohol consumption and type 2 diabetes can be illustrated by a J‐shaped curve. A recent meta‐analysis reported that, compared to abstainers, the risk for type 2 diabetes decreased for <63 g/day and increased above this threshold.This study has several limitations. The major limitation was its cross‐sectional design. Although our results demonstrated the positive associations between fasting glucose level and several lifestyle factors and metabolic parameters among adults without diagnosis of diabetes, causal interpretation of these findings is difficult. Furthermore, recall bias may have influenced the classifications of lifestyle factors in this study. Despite these limitations, our study included a large number and nationally representative sample of Korean adults, which enabled us to convey the public health implications regarding the relationship between fasting glucose levels with multiple lifestyle factors and metabolic parameters among individuals who are not treated for diabetes.In conclusion, higher fasting glucose levels in individuals without diagnosis of diabetes were associated with poor blood pressure control and lipid profiles even in the pre‐IFG range of fasting glucose levels. Obesity, abdominal obesity, and heavy drinking were associated with higher fasting glucose levels in individuals without diagnosis of diabetes. Efforts should be undertaken to prevent increased fasting glucose levels, as higher levels, even within the normal range, were associated with poor metabolic profiles. Furthermore, individuals with fasting glucose levels of 90 to 99 mg/dl should also be screened for hypertension and dyslipidemia as they are more likely to have worse metabolic profiles than those with levels of <90 mg/dl. Lifestyle interventions, especially the prevention of obesity, abdominal obesity, and heaving drinking, should be emphasized to prevent increased fasting glucose levels.The authors declare that there is no competing interest.


Migratory species are protected under international legislation; their seasonal movements across international borders may therefore present opportunities for understanding how global conservation policies translate to local‐level actions across different socio‐ecological contexts. Moreover, local‐level management of migratory species can reveal how culture and governance affects progress towards achieving global targets. Here, we investigate potential misalignment in the two‐way relationship between global‐level conservation policies (i.e. hunting bans and quotas) and local‐level norms, values and actions  in the context of waterfowl hunting in northern Kazakhstan as a case‐study.Anser erythropus; IUCN Vulnerable) under different scenarios of survival rates and hunting offtake, to understand how goose population demographics interact with the local socio‐ecological context.Northern Kazakhstan is globally important for waterfowl and a key staging area for arctic‐breeding species. Hunting is managed through licences, quotas and seasonal bans under UN‐AEWA intergovernmental agreements. To better understand the local socio‐ecological context of waterfowl hunting, we take a mixed‐methods approach using socio‐ecological surveys, informal discussions and population modelling of a focal migratory goose species to: (a) investigate motivations for hunting in relation to socio‐economic factors; (b) assess knowledge of species' protection status; and (c) predict the population size of Lesser White‐fronted Geese . Here, we investigate how global conservation policy and local norms interact to affect the management of a threatened migratory species, which is particularly important for the protection and sustainable management of wildlife that crosses international borders where local contexts may differ. Our study highlights that to be effective and sustainable in the long‐term, global conservation policies must fully integrate local socio‐economic, cultural, governance and environmental contexts, to ensure interventions are equitable across entire species' ranges. This approach is relevant and adaptable for different contexts involving the conservation of wide‐ranging and migratory species, including the 255 migratory waterfowl covered by UN‐AEWA (United Nations Agreement on the Conservation of African‐Eurasian Migratory Waterbirds). Intergovernmental agreements including the United Nations Convention on the Conservation of Migratory Species of Wild Animals CMS,  are in pTop‐down conservation decisions and interventions may be misaligned with local norms, values and actions  investigate motivations for waterfowl hunting in relation to socio‐economic factors; (b) assess knowledge of waterfowl species protection status and (c) predict population sizes of LWfG under different scenarios of survival rates and hunting offtake. This multidisciplinary approach allows greater understanding of how the local socio‐ecological context and uncertainty in population parameters  may affect LWfG conservation outcomes, and illuminates the complex two‐way relationship between local contexts and global policies.22.12) and 542,500 . Census figures include the regional capital cities, Kostanay and Petropavl.The northern Kazakhstan landscape comprises a mosaic of agriculture, forest‐steppe and steppe with numerous lakes and wetlands Figure , which a2.1.1Anser anser and Greater White‐fronted Geese Anser albifrons, of which the latter looks very similar to LWfG . Licences vary in cost depending on both the species listed and the hunting quota. Illegal hunting (e.g. over quota or targeting species not included on a licence) is punishable by fines or imprisonment if caught hunting a red listed species. Regional inspectors monitor compliance, but enforcement efficacy can be limited by the size of area covered by single inspectors , and limited legal powers to search hunting bags. Hunting inspectors can be hunters themselves, and thus enforcement of hunting regulations can be complex and challenging  via key staging areas in Central Asia and eastern Europe, are only partially known . Respondents did not need to be hunters to participate in the survey because we were interested to know how common species protection knowledge is. Discussions were solicited following questionnaire surveys: bilingual co‐authors facilitated communication between the lead author and respondents.Surveys were undertaken during the autumn waterfowl hunting season in September and October 2017 throughout the Kostaney region and North Kazakhstan  with a high risk of participants refusing to engage with the questionnaire. Thus, to ensure engagement was maximised, the UCT focused on hunting geese in general, without detailing a specific species. By doing so, we could use ‘hunting geese’ as the sensitive activity because hunting is illegal without a licence, and because a spring hunting ban had been newly implemented.Participants were randomly assigned to ‘Control’ or ‘Treatment’ groups and were asked a training question (how many activities have you undertaken in the past month?) to ensure the UCT was understood . Participants were then asked: (a) How many of these activities have you done in the past year?; (b) How many activities have you done in autumn/winter (September–February); (c) How many of these activities have you done in spring/summer (March–August)?; (d) How many of these activities have you done for cash? .2.2.2Information on participant demographics, hunting licence ownership and knowledge of waterfowl species protection status was obtained through a questionnaire . We asse2.3This study was reviewed by the General University Ethics Panel . Key ethical considerations included complete anonymisation of data, including survey locations . Predictor variables .Data were analysed using  linear mixed‐effects models ([G]LMMs) using the ‘2.4.1r package ‘list’; Blair & Imai, p values between 0.31–1 for each list item; Whytock et al., Using ‘ict.test’  with demographic variables and licence type Table . Knowled2.5t + 1, Nt+1, is the result of three processes: (a) the survival and return of individuals present in the study area at year t (hereafter referred to as the ‘return rate’); (b) the recruitment into the population of a fixed number of juveniles accompanying returning individuals; and (c) a fixed carrying capacity effect, which increases the probability of individual mortality linearly whenever population size exceeds carrying capacity. Illegal harvesting takes place once all individuals have returned to the study area. This assumes that mortality along the rest of the flyway, reflected in the return rate, and mortality due to illegal hunting are additive.Using demographic parameters sourced from empirical literature, we constructed a simple population model to numerically simulate the effect of varying levels of illegal hunting on LWfG population dynamics and extinction probability over a 20‐year period. In a single year of our model, the LWfG population size observed in the study area at time Nt independent Bernoulli trials, such that the total number of adults returning to the study area at t + 1  was modelled by sampling from a binomial distribution,ϕ is the mean individual return rate. Note that this assumes mortality along the flyway is independent across all individuals . The number of juveniles recruited into the population at t + 1 is modelled as,F=12pNa,t+1 and r is the mean brood size. Values of F were floored to model a natural number of breeding individuals. The effect of density dependence (ψ) on mortality for each individual in the population was then modelled as a function of Na,t+1, Nj,t+1 and a fixed carrying capacity K, such thatNa,t+1 + Nj,t+1) < K, then ψNa,t+1,Nj,t+1,K=0 , and the probability of mortality increases linearly when  > K. In addition, values of ψ > 1 were set to 1 . To model stochastic effects of density dependence applied to each individual, we then model the predicted size of the LWfG population by sampling from a binomial distribution,Nt+1 individuals in the population become adults at the beginning of the following time step.Adult return rate was modelled using ϕ between 0.6 and 0.84, reflecting a range of plausible survival values found in populations under reduced harvesting pressure , mean annual growth rate (i.e. mean{[Nt+1 − Nt]/Nt}), and extinction probability . Population size at t = 0 was set to 32,000 individuals as per the estimated number of individuals in northern Kazakhstan reported by Cuthbert & Aarvak . This was varied from 0 to 10,000 animals (in increments of 100) to represent an extreme scenario in which each hunter within the study area harvested one LWfG. Due to uncertainty related to LWfG return rate, we also varied the value of Lampila, . In all & Aarvak  and K tong Mooij . We ran & Aarvak . We test3n = 70) owned hunting licences, and of these licences, a combined ‘Goose and Duck’ licence was most common  73.91% did not know LWfG are protected, 8.7% misclassified LWfG as unprotected and 17.39% correctly stated that LWfG are protected  and our focal species, 6.6% misclassified LWfG as unprotected, while 28.3% stated that they did not know Figure . Althougd Figure .3.3ϕ = 0.6 to 5.3 for ϕ = 0.84  passing through staging grounds in northern Kazakhstan, an area popular for recreational hunting. We show that hunting waterfowl in northern Kazakhstan is more strongly associated with social and cultural factors, rather than being financially motivated. We found no evidence that hunters surveyed were non‐compliant with newly implemented seasonal hunting bans and licensing regulations. However, significant knowledge gaps existed surrounding the protection status of waterfowl, including LWfG. Accidental and illegal LWfG offtake occurs, although the exact extent remains unknown. To assess the potential consequences of this, using simulations of population dynamics under varying harvesting scenarios, we showed that the LWfG population is sensitive to changes in the level of potential offtake occurring in northern Kazakhstan, particularly so when survival along the rest of the flyway, here represented by the return rate, was low. More generally, for a given return rate, our predictions suggest the existence of a clear tipping point at which increases in harvest bring about a rapid shift from positive to negative population growth rate values. Taken together, our results highlight the importance of integrating local‐level context and management with conservation policy targeted at the broader migratory range ; although in one area offtake was as high as one LWfG per 20–30 Greater White‐fronted or Greylag Geese (Yerokhov, We found no evidence for non‐compliance with the new spring hunting ban, yet hunting was socially and culturally important (von Essen et al., The numerous wetlands and lakes and low human population density of northern Kazakhstan offers huge scope for illegal hunting to remain undetected. Hunting inspectors explained that illegal hunters can be wealthy individuals and often ‘outsiders’ from larger towns and cities, with powerful off‐road vehicles that enable easy evasion of inspectors. Despite considerable efforts, limited resources can preclude effective local‐level enforcement of global‐level hunting regulations. In order to bolster local stakeholder engagement, maximise on resources, and embed the social and cultural importance of hunting for local people within conservation policies, we suggest that greater consideration of the knowledge, views and perceptions of hunters can help inform the active management of waterfowl species, similar to goose hunting and adaptive management governance in Denmark (Williams et al., 4.2In order to connect local action and global policy, partnerships of governments, businesses, non‐governmental organisations and local people are crucial Berkes, . For insIn the case of northern Kazakhstan and LWfG, despite local implementation of hunting bans and restrictions the LWfG population is still declining. Our current knowledge of LWfG population movement, behaviour and demographics is incomplete: drivers of decline may lie in multiple locations along its flyway, despite being covered by international agreements provided by UN‐AEWA (Jones et al., 4.3Global conservation policies and targets may not fully address population declines in wide‐ranging or migratory species if interventions are not dynamic and locally appropriate for differing socio‐economic, environmental and governance contexts across range states. In order to address this issue, detailed and long‐term information on socio‐economic, environmental and governance contexts is needed. To gain such information, we recommend the following, complementary, approaches. First, socio‐ecological data, stakeholder decision‐making and species population modelling needs to be fully integrated (c.f. Duthie et al., I.L.J., R.C.W., N.B., A.T., I.Z. and K.Z. designed the socio‐ecological surveys; I.L.J., A.T., I.Z. and K.Z. collected the data; I.L.J., J.J.C. and J.M. led the data analysis. All authors contributed important intellectual content and to manuscript writing and editing; all authors gave final approval for publication.Appendix S1 Supporting InformationClick here for additional data file.