Elizabeth Nicholls
email: e.nicholls@sussex.ac.uk

Cristina Botías

Ellen L Rotheray

Penelope Whitehorn

Arthur David

Robert Fowler

Thomas David

Hannah Feltham

Jennifer L Swain

Patricia Wells

Elizabeth M Hill

Juliet L Osborne

Dave Goulson

Monitoring neonicotinoid exposure for bees in rural and peri-urban areas of the UK during the transition from pre-to post-moratorium

Concerns regarding the impact of neonicotinoid exposure on bee populations recently led to an EU-wide moratorium on the use of certain neonicotinoids on flowering crops. Currently evidence regarding the impact, if any, the moratorium has had on bees' exposure is limited. We sampled pollen and nectar from bumblebee colonies in rural and peri-urban habitats in three UK regions; Stirlingshire, Hertfordshire and Sussex. Colonies were sampled over three years; prior to the ban (2013), during the initial implementation when some seed-treated wintersown oilseed rape was still grown (2014), and following the ban (2015). To compare species-level differences, in 2014 only, honeybee colonies in rural habitats were also sampled. Over half of all samples were found to be contaminated (n=408), with thiamethoxam being the compound detected at the highest concentrations in honeybee-(up to 2.29 ng/g in nectar in 2014, median≤0.1 ng/g, n=79)

and bumblebee-collected pollen and nectar (up to 38.77 ng/g in pollen in 2013, median ≤0.12 ng/g, n=76). Honeybees were exposed to higher concentrations of neonicotinoids than bumblebees in 2014.

While neonicotinoid exposure for rural bumblebees declined post-ban (2015), suggesting a positive impact of the moratorium, the risk of neonicotinoid exposure for bumblebees in peri-urban habitats remained largely the same between 2013 and 2015.

INTRODUCTION

Neonicotinoids are the most commonly used insecticides worldwide [START_REF] Simon-Delso | Systemic Insecticides (Neonicotinoids and Fipronil): Trends, Uses, Mode of Action and Metabolites[END_REF] . Their systemic nature means that, following seed-application to crops such as oilseeds or cereals, neonicotinoids become incorporated into the tissues of a plant as it grows, including pollen and nectar, the main source of food for economically important pollinators, such as honeybees and bumblebees [START_REF] Ollerton | How Many Flowering Plants Are Pollinated by Animals?[END_REF] . Multiple studies have raised concerns regarding the negative impacts of neonicotinoid exposure on bees [START_REF] Alkassab | Sublethal Exposure to Neonicotinoids and Related Side Effects on Insect Pollinators: Honeybees, Bumblebees, and Solitary Bees[END_REF] . Whitehorn et al. (2012) [START_REF] Whitehorn | Neonicotinoid Pesticide Reduces Bumble Bee Colony Growth and Queen Production[END_REF] found that exposure of bumblebees to pollen and nectar containing 6 ng/g and 0.7 ng/g of imidacloprid respectively, resulted in slower colony growth and the production of fewer new queens, relative to unexposed colonies. Other studies have observed detrimental impacts on foraging and navigation [START_REF] Gill | Combined Pesticide Exposure Severely Affects Individual-and Colony-Level Traits in Bees[END_REF][START_REF] Henry | A Common Pesticide Decreases Foraging Success and Survival in Honey Bees[END_REF] , immunity [START_REF] Di Prisco | Neonicotinoid Clothianidin Adversely Affects Insect Immunity and Promotes Replication of a Viral Pathogen in Honey Bees[END_REF] and worker mortality [START_REF] Laycock | Repression and Recuperation of Brood Production in Bombus Terrestris Bumble Bees Exposed to a Pulse of the Neonicotinoid Pesticide Imidacloprid[END_REF] . Based on these findings, in 2013 the European Commission instated a EU-wide moratorium on the use of three types of neonicotinoid, thiamethoxam, clothianidin and imidacloprid on bee-attractive flowering crops such as oilseed rape 9 .

In 2018, this ban was subsequently expanded to include all field crops [START_REF][END_REF][START_REF][END_REF][12] .

Criticism has been levied against studies cited in support of the moratorium, mainly for using neonicotinoid concentrations purported to exceed those routinely experienced by foraging bees [START_REF] Carreck | The Dose Makes the Poison: Have "field Realistic" Rates of Exposure of Bees to Neonicotinoid Insecticides Been Overestimated in Laboratory Studies?[END_REF] , sparking demand for further evidence as to what constitutes a 'field-realistic' dose. Several studies have screened bee-collected pollen and nectar [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF][START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF][START_REF] Rundlöf | Seed Coating with a Neonicotinoid Insecticide Negatively Affects Wild Bees[END_REF][START_REF] Tosi | A 3-Year Survey of Italian Honey Bee-Collected Pollen Reveals Widespread Contamination by Agricultural Pesticides[END_REF][START_REF] Colwell | Honey Bee-Collected Pollen in Agro-Ecosystems Reveals Diet Diversity, Diet Quality, and Pesticide Exposure[END_REF][START_REF] Balfour | Landscape Scale Study of the Net Effect of Proximity to a Neonicotinoid-Treated Crop on Bee Colony Health[END_REF] for neonicotinoid residues, quantifying the 'exposure landscape' by incorporating multiple chemicals from several forage sources.

Concentrations have been shown to vary considerably across studies, depending on location, time of year and species. Pollen sampled from rural bumblebee colonies in Sussex, England, prior to the implementation of the moratorium in 2013, was found to contain 18 ng/g of thiamethoxam on average, with pollen collected from nests in nearby peri-urban areas containing up to 20 ng/g imidacloprid [START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF] (mean=6.5 ng/g), well above the 6 ng/g used by Whitehorn et al. 9 . A large scale Swedish field study found clothianidin concentrations averaging 5.4 ng/g in nectar sampled from bumblebees foraging in fields of seed-treated oilseed rape (range 1.4-14 ng/g) [START_REF] Rundlöf | Seed Coating with a Neonicotinoid Insecticide Negatively Affects Wild Bees[END_REF] . In contrast, a study conducted in Germany found considerably lower average concentrations (0.88 ng/g) in pollen collected from bumblebee nests adjacent to neonicotinoids treated winter-sown oilseed rape [START_REF] Rolke | Large-Scale Monitoring of Effects of Clothianidin-Dressed Oilseed Rape Seeds on Pollinating Insects in Northern Germany: Residues of Clothianidin in Pollen, Nectar and Honey[END_REF] , and a more recent study conducted across the UK, Hungary and Germany reported that concentrations detected in pollen and nectar collected by honeybees, bumblebees and the solitary bee Osmia bicornis rarely exceeded 1.5 ng/g [START_REF] Woodcock | Country-Specific Effects of Neonicotinoid Pesticides on Honey Bees and Wild Bees[END_REF] . The wide ranging values reported by these studies highlights the need for further data to determine the actual exposure risk, particularly for wild bees.

Here we monitored bees' risk of neonicotinoid exposure during the period from pre-to postmoratorium, by screening pollen and nectar collected from bumblebee colonies located in several regions; Sussex (2013-2015) and Hertfordshire (2014 only) in the south of England and Stirling, Scotland (2013 only) in the north of the UK. Given the total weight of neonicotinoids applied in Scotland is much lower compared to the south of England (FERA PUS STATS database [START_REF]FERA PUS STATS for insecticides[END_REF] ), we expected the exposure risk to be lowest for bees in this region. There is currently limited data on the exposure risk for wild bees from foraging on ornamental plants grown using neonicotinoids [START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF][START_REF] Botías | Quantifying Exposure of Wild Bumblebees to Mixtures of Agrochemicals in Agricultural and Urban Landscapes[END_REF][START_REF] Lentola | Ornamental Plants on Sale to the Public Are a Significant Source of Pesticide Residues with Implications for the Health of Pollinating Insects[END_REF] and the use of neonicotinoid-based garden sprays, therefore we monitored bumblebees in both rural and peri-urban habitats (Sussex and Stirling only), the latter consisting of domestic gardens located on the outskirts of urban areas. For bees in rural areas, we expected neonicotinoid concentrations in pollen and nectar collected in 2015 to be lower than those collected in 2013, before the implementation of the moratorium. In 2014, the impact of the ban may not have fully come into effect, as any wintersown oilseed crops would have been drilled prior to the implementation of the ban in December 2013 and therefore may still have been seed-treated with neonicotinoids. To compare species-level differences in exposure risk during this transitional year (2014), we also screened pollen and nectar from rural honeybee colonies located in Sussex and Hertfordshire.

MATERIALS AND METHODS

Site Information

Bumblebee colonies (B.terrestris audax) were obtained from Agralan Ltd., Swindon, UK (originating from Biobest, Belgium) and in late spring (late May to early June, see Table 1 for exact dates) were placed into the field: i) to monitor exposure risk over the course of the implementation of the ban for both rural and peri-urban habitats, bumblebee colonies were placed in rural (n=135, n=32-47/year) and peri-urban (n=42, 12-15/year) locations across Sussex each year between 2013 and 2015. While the UK granted a derogation to use neonicotinoids on oilseed rape in 2015, this was limited to a portion of East England and did not affect the study area;

ii) to assess regional differences in neonicotinoid exposure between the north and south of the UK, prior to the implementation of the ban (2013), bumblebee colonies were also placed in rural (n=10) and peri-urban (n=20) locations in Stirling. In 2014 only, bumblebees were also placed in rural locations across Hertfordshire (n=30) for comparison with Sussex colonies;

iii) to compare species-level differences in exposure risk, 15 rural bumblebee colonies were each paired with a honeybee colony (located within 10m distance and placed into the field at the beginning of April) in both Sussex and Hertfordshire in 2014 only. Queenright honeybee colonies were obtained from experimental stocks at the University of Sussex and Rothamsted Research, which at the beginning of the experiment consisted of a single brood box and a super containing frames of fresh foundation wax, with additional space for bees to store pollen and nectar added as necessary.

We also mapped which crops were grown in ten, 5 km 2 surrounding the experimental colonies in Sussex and Hertfordshire in 2014 (Fig. S4) and, where possible, asked farmers growing winter-sown oilseed rape which seed treatments they had used (Table S4).

Sampling Pollen and nectar was collected from bumblebee colonies following four, eight and ten weeks of foraging in the field. Pollen was scraped out of the colony using a stainless steel microspoon, which was cleaned using methanol to avoid cross-contamination. From each colony, we aimed to collect enough pollen to fill a 1.5 ml micro-centrifuge tube, to ensure enough material for chemical analysis. Concurrently, 1.5 ml of nectar was obtained from nectar pots using disposable glass pipettes. However, care was taken not to completely deplete bumblebee colony stores. Where stores were low, no sample was collected (Table 2).

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For honeybees, samples were collected once per month in April, May and June 2014, with the last two sampling dates coinciding with sample collection from adjacent bumblebee colonies. Samples were obtained from freshly drawn comb, where possible, to minimise contamination from previous years. Enough pollen to fill a 1.5 ml micro-centrifuge tube was scraped out of ~10 cells using a stainless steel micro-spoon as described above, and 1.5 ml of recently stored nectar was obtained from uncapped and newly drawn comb using disposable glass pipettes. Freshness was determined by first shaking the frame to ensure nectar dripped easily out of the comb. All pollen and nectar samples were stored in individually labelled tubes and put on ice during transport back to the lab, and were then frozen at -20°C until residue analysis was performed.

Chemical analyses: Pollen and nectar samples were extracted using the QuEChERS method [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF] and screened for five neonicotinoids: thiamethoxam (TMX), clothianidin (CLO), imidacloprid (IMC), acetamiprid (ACT) and thiacloprid (THC), using ultra high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). Pollen samples collected in Sussex in 2013 were not screened for acetamiprid.

Sample preparation:

Pollen samples were extracted as described by Botias et al. (2015) [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF] .

Briefly, 100 mg of pollen was weighed into an Eppendorf tube and 400 pg of deuterated pesticides in ACN were added. The extraction was performed by the addition of 400 µl of water, 500 µl of ACN, Nectar samples were centrifuged at 13,000 relative centrifugal force (RCF) for 10 min to remove plant debris and the supernatant transferred into a clean eppendorf tube. Nectar samples were very viscous and were therefore weighed for more accuracy (175 ± 50 mg depending on availability)

125
and the volume then increased to 400 µl with water. Four hundred pg of deuterated pesticide standard

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UHPLC-MS/MS analyses.

The ultra high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method described by Botias et al. (2015) [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF] was used for the analysis of samples. UHPLC-MS/MS analyses were carried out using a Waters Acquity UHPLC system coupled to a Quattro Premier triple quadrupole mass spectrometer from Micromass (Waters, Manchester, UK). Data were acquired using MassLynx 4.1 and the quantification was carried out by calculating the response factor of neonicotinoid compounds to their respective internal standards.

Concentrations were determined using a least-square linear regression analysis of the peak area ratio versus the concentration ratio (native to deuterated). Method detection and quantification limits (MDL and MQL, respectively) as well as recoveries were determined as described by Botias et al. (2015) [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF] (Table S1-3).

Quality control. One blank workup sample (i.e. solvent without matrix) per batch of eleven samples was included and injected on the UHPLC-MS/MS to ensure that no contamination occurred during the sample preparation. Solvent samples were also injected between sample batches to ensure that there was no carryover in the UHPLC system that might affect adjacent results in analytical runs.

Samples were analysed in a random order and quality control samples (i.e. standards) were injected during runs every ten samples to check the sensitivity of the machine. Identities of detected neonicotinoids were confirmed by comparing ratio of MRM transitions in samples and pure standards.

Statistical Analysis. All analyses were performed using R-3.3.3. Residue concentrations that were above the MDL but below the MQL were assigned the MDL (Tables 23, range 0.03-0.10 ng/g).

Concentrations below the MDL were assumed to be zero [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF] . Shapiro-Wilk tests, combined with inspection of q-q plots, confirmed that residue data were not normally distributed. Therefore we compared the frequency of neonicotinoid contamination using contingency tables and either χ 2 or Fisher's exact tests (where expected frequencies were <5). To compare total neonicotinoid concentrations between regions (Sussex vs. Stirling; Sussex vs. Herts), habitats (Rural vs. Peri-Urban)

and years of the study (2013 vs. 2015) we used non-parametric Mann-Whitney tests. For honeybee data, where frequencies of contamination and residue concentrations were compared between samples from the same hive over several months, we used Cochran's Q test (with McNemar's test for post-hoc comparisons) and the Wilcoxon Signed-Rank test, with Bonferroni corrections to account for multiple comparisons. Given the relatively small number of pollen and nectar samples collected from each bumblebee colony, for analyses involving bumblebees we pooled samples collected after four and eight weeks in the field.

RESULTS

Bumblebees:

In total, 233 pollen and nectar samples were collected from bumblebee colonies placed in rural and peri-urban habitats in the regions of Stirling, Sussex and Hertfordshire between 2013 and 2015. Forty percent of all samples screened were found to be contaminated with neonicotinoids, predominantly thiamethoxam (23%), thiacloprid (15%) and imidacloprid (10%). Pollen samples were more often contaminated (62% samples) than nectar (25% samples) and the mean combined total residues detected in pollen (Pollen N=132, 62% samples, mean± standard deviation (SD) =1.44±5.44 ng/g, median ˂MDL, max= 38.77 ng/g) were more than ten times higher (Nectar N=101, mean± SD= 0.12±0.44 ng/g, median ˂MDL, max=3.58 ng/g).

Differences in exposure by habitat and year:

In 2013, the frequency of neonicotinoid contamination was similar for pollen (Table 1, χ 2 1 =0, p=1.000, Rural =58%; Peri-urban= 59%) and nectar (χ 2 1 =0, p=1.000, Rural=14%, Peri-urban =14%) sampled from peri-urban (PU) and rural (R) bumblebee colonies across the regions of Sussex (SU) and Stirling (ST) (Table 1). Concentrations of neonicotinoids were very similar in nectar (Mann-Whitney U 21, 21 =225, p=0.867, mean PU ≤0.10, median PU ≤0.10, mean R ±SD=0.22±0.55 ng/g, median R <MDL), and though higher in pollen from rural colonies, this difference was not significant (U 36, 32 =603.5, p=0.73; mean R =3.37±9.36 ng/g, median R ≤0.12, mean PU = 1.28±3.62 ng/g, median PU ≤0.12). While nectar from both habitats contained only one type of neonicotinoid, predominantly thiamethoxam, over a quarter of pollen samples from bumblebee colonies in rural (28%) and peri-urban (26%) habitats contained more than one residue.

Thiamethoxam (up to 38.77 ng/g, median ˂0.12, mean±SD= 2.08±7.47 ng/g) and clothianidin (up to 2.08 ng/g, mean ≤0.12 ng/g, median ˂0.12 ng/g) were present at the highest concentrations in rural

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In 2014, less than 10% of pollen (n=13) and nectar (n=13) samples from rural bumblebee colonies in Sussex contained neonicotinoids, all thiamethoxam and below the method quantification limit, whereas a significantly higher proportion of both pollen (85%, χ Pollen sampled from Sussex colonies was more frequently contaminated with multiple residues (Peri-urban=37%, Rural=35%) compared to Stirling samples (Peri-urban=8%, Rural=15%), and the concentrations of thiamethoxam detected in pollen were considerably higher (mean SU =0.58±1.64 ng/g, median=0.12 ng/g vs. mean ST ≤0.12 ng/g, median ˂0.12 ng/g). Sussex periurban colonies in particular also contained higher concentrations of thiacloprid compared to Stirling (mean SU =1.47±4.41 ng/g median ˂0.03 ng/g vs. mean ST = 0.07±0.22 ng/g, median ˂0.03 ng/g).

Imidacloprid was also frequently detected in pollen from Sussex nests in 2013, but was not detected in any samples from Stirling. Clothianidin was not detected in any Sussex nests, but accounted for the highest residue concentrations detected in nests in Stirling (mean ST = 0.16±0.58 g/g, median ˂MDL, max ST = 2.08 ng/g).

In 2014, residues detected in pollen and nectar samples collected from bumblebee colonies placed in rural habitats in Hertfordshire (H) and Sussex (SU) were all below the limits of quantification (<0.04-0.1 ng/g). Though there was a higher frequency of contamination of both pollen (H=36%, SU=7%) and nectar (H=20%, SU= 8%) from Hertfordshire colonies, this difference was not significant (Nectar: Fisher's Exact Test p=0.560; N SU =13, N H =10; Pollen p=0.142, N SU =13, N H =11).

A small proportion of pollen from Sussex (10%), and nectar from both regions was contaminated with thiamethoxam (SU=10%; H=20%). Pollen from Hertfordshire colonies also contained acetamiprid (10%) and, more frequently, thiacloprid (40%).

Honeybees: In total, 175 pollen and nectar samples were collected from honeybee hives in Sussex and Hertfordshire between April and June May 2014, with over two thirds (68%) found to be contaminated with neonicotinoids. Total residue concentrations in nectar (N= 85, mean± SD = 0.64 ± 0.84 ng/g, median=0.20 ng/g, max= 4.23 ng/g) were approximately three times the concentrations detected in pollen (N= 90, mean± SD = 0.20 ± 0.32 ng/g, median ≤0.12 ng/g, max=1.74 ng/g), with 40% of nectar samples containing more than one residue, compared to just 9% of pollen samples.

Alongside thiamethoxam, which was highly prevalent in both pollen (61% of samples) and nectar only once in pollen (Table 2). Imidacloprid and thiacloprid were detected in a very small percentage of samples (4-5%) and acetamipirid was not detected.

Seasonal differences: Frequency of neonicotinoid detection in pollen (Cochran's Q=24.67, df=2, p<0.001) and nectar (Q=20.38, df=2, p<0.001) sampled from honeybee colonies in 2014 changed significantly across the season. The highest frequency and concentration of neonicotinoid residues were detected in April (Fig. 3), when nearly all nectar samples collected from hives in Hertfordshire (H) and Sussex (SU) were contaminated with neonicotinoids (H=100%, mean H ± SD =1.46±0.66 ng/g; median=1.17 ng/g; SU=93%, mean SU =0.95 ±1.13 ng/g, median ≤0.12 ng/g).

Likewise, almost all pollen samples contained neonicotinoid residues (H=80%, mean H =0.41±0.47 ng/g, median ≤0.12 ng/g; SU=100%, mean SU =0.23±0.19 ng/g, median ≤0.12 ng/g) in April.

Between study areas in each region in 2014, showed that arable crops accounted for 55% of land cover in Hertfordshire (9% oilseed rape), and 32% in Sussex (5% oilseed rape, Figure S4).

Species-specific differences:

A comparison of residue concentrations in pollen and nectar collected from adjacent honeybee (HB) and bumblebee (BB) nests located in rural habitats in Hertfordshire and Sussex revealed significantly higher concentrations of neonicotinoid exposure for honeybees compared to bumblebees (Table 1, 2, U 18, 18 = 112, p=0.04; mean HB =0.17±0.39 ng/g, median ˂MDL, max=1.38 ng/g; mean BB ≤0.12 ng/g, median ˂MDL, max ≤0.12 ng/g).

DISCUSSION

In December 2013, an EU-wide moratorium on the use of certain neonicotinoids on bee-attractive flowering crops was implemented by the European Commission, which in early 2018 was subsequently expanded to include all field crops. To monitor bees' exposure to neonicotinoids during the intital transitional period from pre-to post-ban, between 2013 and 2015 we collected more than 400 pollen and nectar samples from bumblebee and honeybee colonies located in rural and peri-urban habitats in three regions across the UK, finding just over half of all samples to be contaminated with neonicotinoids. While combined total concentrations of neonicotinoids in pollen collected by rural bumblebees declined post-ban from an average of 5.1 ng/g in 2013, to 0.06 ng/g in 2015, suggesting a positive impact of the moratorium, neonicotinoid concentrations detected in samples collected from peri-urban bumblebee colonies remained largely unchanged between 2013 and 2015, indicating that the risk of exposure for peri-urban bees was not altered during the transitional period, and that more could be done to mitigate the risk for bees foraging in such habitats.

Across all samples, the highest neonicotinoid residue concentrations were detected in 2013, in pollen samples collected from rural bumblebee colonies in Sussex. Concentrations of up to 38.77 ng/g of thiamethoxam were detected, with the average total neonicotinoid concentrations of 5.1 ng/g similar to that detected by previous studies conducted prior to the moratorium [START_REF] Scheper | Environmental Factors Driving the Effectiveness of European Agri-Environmental Measures in Mitigating Pollinator Loss--a Meta-Analysis[END_REF][START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF][START_REF] Botías | Contamination of Wild Plants near Neonicotinoid Seed-Treated Crops, and Implications for Non-Target Insects[END_REF] , and within the range demonstrated to have negative impacts on bumblebee physiology [START_REF] Moffat | Chronic Exposure to Neonicotinoids Increases Neuronal Vulnerability to Mitochondrial Dysfunction in the Bumblebee (Bombus Terrestris)[END_REF][START_REF] Moffat | Neonicotinoids Target Distinct Nicotinic Acetylcholine Receptors and Neurons, Leading to Differential Risks to Bumblebees[END_REF] , foraging efficiency [START_REF] Feltham | Field Realistic Doses of Pesticide Imidacloprid Reduce Bumblebee Pollen Foraging Efficiency[END_REF] and colony growth [START_REF] Moffat | Neonicotinoids Target Distinct Nicotinic Acetylcholine Receptors and Neurons, Leading to Differential Risks to Bumblebees[END_REF] . Pre-ban (2013), the frequency of neonicotinoid contamination was extremely high for pollen sampled from bumblebee colonies in both rural and peri-urban habitats in Sussex (74% and 84% of pollen samples respectively, mean=3.74 ng/g). As predicted, pollen samples collected from nests near Stirling in 2013 were contaminated to a lesser degree (23-30% of pollen samples), and with lower concentrations (mean=0.20 ng/g). This likely reflects the fact that across Scotland, neonicotinoid use in 2013/2014 was approximately four times lower than in South East England (4, 186 kg, over 78, 345 ha vs. 16, 820 kg, over 197,507 ha [START_REF]FERA PUS STATS for insecticides[END_REF] ), though differences in the growth season and therefore timing of neonicotinoid application between regions may also have played a role.

Pollen and nectar samples collected from honeybee colonies in 2014, post-implementation of the ban, but when any winter-sown oilseed rape may still have been seed-treated with neonicotinoids, also had a high prevalence of neonicotinoid contamination (68% samples). Contamination was highest in April when oilseed rape was flowering (93% samples), and declined throughout the season, a phenomenon observed in several earlier studies [START_REF] Botías | Neonicotinoid Residues in Wildflowers, a Potential Route of Chronic Exposure for Bees[END_REF][START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF][START_REF] Botías | Quantifying Exposure of Wild Bumblebees to Mixtures of Agrochemicals in Agricultural and Urban Landscapes[END_REF][START_REF] Woodcock | Neonicotinoid Residues in UK Honey despite European Union Moratorium[END_REF] , and hypothesised to arise from temperature increases and photo-degradation of neonicotinoid residues in plant tissues as the season progresses [START_REF] Bonmatin | Environmental Fate and Exposure; Neonicotinoids and Fipronil[END_REF] .

During this early part of the year, concentrations detected in honeybee-collected nectar averaged 1.2 ng/g, close to the average maximum concentration detected in seed-treated crop nectar, as reported by Godfray et al. [START_REF] Godfray | A Restatement of the Natural Science Evidence Base Concerning Neonicotinoid Insecticides and Insect Pollinators[END_REF] (1.9 ng/g, averaged from 20 published studies). Concentrations in pollen were considerably lower (0.32 ng/g, average maximum concentration in seed-treated crop pollen=6.1 ng/g 32 ), likely reflecting honeybees' preference for collecting nectar from oilseed rape. For both bumblebees and honeybees, early spring is a period when the colony might be expected to be particularly vulnerable [START_REF] Seeley | Survival of Honeybees in Cold Climates: The Critical Timing of Colony Growth and Reproduction[END_REF][START_REF] Wu-Smart | Sub-Lethal Effects of Dietary Neonicotinoid Insecticide Exposure on Honey Bee Queen Fecundity and Colony Development[END_REF] , and levels detected in pollen were within the range known to impair honeybee foraging performance [START_REF] Cresswell | A Meta-Analysis of Experiments Testing the Effects of a Neonicotinoid Insecticide (Imidacloprid) on Honey Bees[END_REF] , immune function [START_REF] Di Prisco | Neonicotinoid Clothianidin Adversely Affects Insect Immunity and Promotes Replication of a Viral Pathogen in Honey Bees[END_REF] and alter gene expression pathways [START_REF] Christen | Molecular Effects of Neonicotinoids in Honey Bees (Apis Mellifera)[END_REF] .

Furthermore, as observed in several previous studies [START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF][START_REF] Tosi | A 3-Year Survey of Italian Honey Bee-Collected Pollen Reveals Widespread Contamination by Agricultural Pesticides[END_REF][START_REF] Colwell | Honey Bee-Collected Pollen in Agro-Ecosystems Reveals Diet Diversity, Diet Quality, and Pesticide Exposure[END_REF] , many of the samples we screened were found to contain more than one neonicotinoid residue, which gives rise to the potential for additive or synergistic effects. Tosi et al. [START_REF] Tosi | A 3-Year Survey of Italian Honey Bee-Collected Pollen Reveals Widespread Contamination by Agricultural Pesticides[END_REF] found when screening honeybee pollen collected from multiple apiaries across Italy for 66 different pesticides, that the frequency of detection actually peaked in summer months. Though here we did not screen for the presence of other chemical classes such as fungicides, there is evidence to suggest that exposure to certain fungicides can make bees more susceptible to the adverse effects of neonicotinoids [START_REF] Iwasa | Mechanism for the Differential Toxicity of Neonicotinoid Insecticides in the Honey Bee, Apis Mellifera[END_REF] .

Although the concentration of neonicotinoids in pollen and nectar sampled from rural bumblebee colonies declined between 2013 and 2015, bumblebees from both rural and peri-urban habitats were nevertheless still exposed to neonicotinoids following the implementation of the ban.

Indeed 47% of nectar and 36% of pollen samples collected from rural colonies in 2015 contained neonicotinoid residues, a similar frequency as observed for peri-urban nests (33% nectar, 64% pollen), albeit at lower concentrations (mean concentration detected in pollen from rural nests = 0.06 ng/g vs. 1.29 ng/g detected in peri-urban pollen in 2015). This echoes the findings of Woodcock et al. [START_REF] Woodcock | Neonicotinoid Residues in UK Honey despite European Union Moratorium[END_REF] who screened honey samples submitted by beekeepers across the UK, and found that while samples harvested in 2014 were more likely to be contaminated (52% samples), 22.9% of samples harvested post-ban in 2015 also contained neonicotinoids. Similarly, a worldwide study of honey contamination spanning five years between 2012 and 2016, found 75% of 198 samples to contain neonicotinoids, with the highest prevalence in honey from North America, Asia and Europe [START_REF] Mitchell | A Worldwide Survey of Neonicotinoids in Honey[END_REF] .

Not only did exposure to neonicotinoids change for rural bees between 2013 and 2015, so did the chemical type. Across all samples, thiamethoxam was the most frequently detected, which is unsurprising given that, prior to the moratorium, it was the active ingredient in the mostly commonly used seed dressing on oilseed rape across Great Britain. Indeed, of fifteen farmers growing wintersown oilseed rape within a 5 km radius of our experimental bee colonies that we interviewed in 2014, twelve had used seeds dressed with a thiamethoxam-based formulation (Cruiser®). Clothianidin, a metabolite of thiamethoxam and still in use as a seed-dressing on non-flowering cereal crops, was also frequently detected in honeybee nectar (69% samples), but only once in pollen, and was rarely detected in any samples collected from bumblebee colonies. Post-ban, acetamiprid and thiacloprid, the use of which is unaffected by the moratorium, were detected more often and at higher levels than thiamethoxam. For nectar samples collected from rural bumblebee colonies, thiacloprid concentrations actually significantly increased between 2013 and 2015. Thiacloprid is an active ingredient in many bug sprays sold in garden centres, and a recent study in which ornamental 'beefriendly' plants were screened for multiple pesticide and fungicide residues found more than 70% of plants contained neonicotinoids, with thiacloprid present in almost half [START_REF] Lentola | Ornamental Plants on Sale to the Public Are a Significant Source of Pesticide Residues with Implications for the Health of Pollinating Insects[END_REF] .

Imidacloprid was detected in a moderate proportion (10%) of samples collected from bumblebee nests throughout the duration of the study. Considering that use of imidacloprid in arable farming has dramatically declined in the UK (50% and 90% decline in weight of imidacloprid applied to cereals and oilseeds respectively between 2012 and 2014, PUS Stats database, Table S6), having been replaced by thiamethoxam and clothianidin, it is somewhat concerning that it was detected to such an extent. Woodcock et al. [START_REF] Woodcock | Neonicotinoid Residues in UK Honey despite European Union Moratorium[END_REF] also noted that imidacloprid was present in honey samples harvested in 2014 at a rate 'disproportional to its use' and Tosi et al. [START_REF] Tosi | A 3-Year Survey of Italian Honey Bee-Collected Pollen Reveals Widespread Contamination by Agricultural Pesticides[END_REF] detected imidacloprid in 9.1% of honeybee-collected pollen sampled from multiple apiaries across Italy in 2014 at mean concentrations of 2 ng/g, raising concerns about the persistence of this chemical in agroenvironments. As previously observed when screening pollen from bumblebee colonies [START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF] and wild bumblebees collected in peri-urban areas [START_REF] Botías | Quantifying Exposure of Wild Bumblebees to Mixtures of Agrochemicals in Agricultural and Urban Landscapes[END_REF] , the highest concentrations of imidacloprid were detected in peri-urban colonies, at levels up to 11.16 ng/g in 2015 (mean=1.13 ng/g). . Again, this may originate from use by the horticulture industry, since screening of ornamental plants detected imidacloprid in 38% of samples [START_REF] Lentola | Ornamental Plants on Sale to the Public Are a Significant Source of Pesticide Residues with Implications for the Health of Pollinating Insects[END_REF] . An alternative, yet untested source, is the use of imidacloprid for flea control in domestic pets and as ant poison.

Honeybees in Hertfordshire were exposed to significantly higher neonicotinoid concentrations in nectar compared to Sussex honeybees, which is most likely explained by the fact that, in 2014, there was almost double the percentage cover of treated oilseed crops (9% land cover in Hertfordshire vs. 5% in Sussex), and generally a higher percentage of arable land cover (55%) compared to Sussex (32%).

Overall, honeybee samples had higher concentrations of neonicotinoids compared to bumblebees. This contrasts with findings from an earlier study conducted in 2013 where the reverse was found to be true [START_REF] David | Widespread Contamination of Wildflower and Bee-Collected Pollen with Complex Mixtures of Neonicotinoids and Fungicides Commonly Applied to Crops[END_REF] . However in the previous study, colonies of each species were not placed in identical locations, therefore in addition to differences in foraging range and flower preferences [START_REF] Leonhardt | The Same, but Different: Pollen Foraging in Honeybee and Bumblebee Colonies[END_REF][START_REF] Wood | Pollinator-Friendly Management Does Not Increase the Diversity of Farmland Bees and Wasps[END_REF] , colonies may simply have been in proximity to a different range of plant species. Clearly more paired sampling of both species is required to establish whether there are consistent differences in exposure.

On the basis of evidence published post-2013, the European Food Standards Agency recently concluded that neonicotinoids do indeed pose a risk to bees [START_REF] Efsa | Evaluation of the Data on Clothianidin, Imidacloprid and Thiamethoxam for the 402 Updated Risk Assessment to Bees for Seed Treatments and Granules in the EU[END_REF] , and in 2017 the EU commission proposed extending the moratorium to include all field crops (barring permanent greenhouse crops), which was passed by the European Union in early 2018 [START_REF][END_REF][START_REF][END_REF][12] . Here we have shown for the first time Table 2 Frequency of detection (% samples), mean (± standard deviation (SD)), median and maximum concentrations of five neonicotinoids (TMX=thiamethoxam, CLO= clothianidin, IMC= imidacloprid, ACT=acetamiprid, THC=thiacloprid) and the combined total concentration of neonicotinoids detected in pollen and nectar sampled from bumblebee colonies located in rural and peri-urban habitats in three different regions; Stirling, Hertfordshire (Herts) and Sussex. Samples were collected across three years (2013-2015). Multi-residue samples are those where more than one type of neonicotinoid was detected. MQL= Method quantification limit, MDL=Method detection limit, nt= not tested, ≤ less than or equal to. 3 Frequency of detection (% samples), mean (± standard deviation (SD)), median and maximum concentrations of five neonicotinoids (TMX=thiamethoxam, CLO= clothianidin, IMC= imidacloprid, ACT=acetamiprid, THC=thiacloprid) and the combined total concentration of neonicotinoids detected in honeybee nectar and pollen sampled from colonies located in in Sussex (N=15) and Hertfordshire (Herts, N=15) between April and June. Multiresidue samples are those where more than one type of neonicotinoid was detected. MQL= Method quantification limit, MDL=Method detection limit, nt= not tested, ≤ less than or equal to.

FIGURESFigure 1

 1 FIGURES

Figure 2

 2 Figure 2 Total neonicotinoid concentrations (Thiamethoxam, clothianidin, imidacloprid, acetamiprid and thiacloprid combined) detected in A) Pollen and B) Nectar samples collected from bumblebee colonies in Rural (White, N Pollen samples=45; Nectar=33) and Peri-urban (Grey, N Pollen samples= 30; Nectar=25) habitats across the region of Sussex in the years 2013 and 2015. Concentrations are plotted on a square-root scale. Black horizontal bars show median values. Box limits denote the first and third quantiles, and boxplot whiskers extend to 1.5 times the interquartile range. Outliers are represented by solid black circles.

  13, 12 = 58.5, p= 0.067, mean 2013 ˂MDL, median ˂MDL, mean 2015 =0.05±0.13 ng/g, median ˂MDL), again there was no difference in the concentrations detected in pollen and nectar collected from peri-urban nests between 2013 and 2015. Exact Test p=1.00, Sussex=14%; Stirling 12.5%) and concentrations (U 27,15 =200, p=0.931; mean SU =0.11±0.37 ng/g, median SU ˂MDL, mean ST =0.13±0.47 ng/g, median ST ˂MDL).

	Environmental Science & Technology
	nectar (U 14, 19 =98, p=0.134; mean 2013 = 0.20±0.51 ng/g, median ˂MDL, mean 2015 =0.10±0.15 ng/g,
	median ˂MDL). When considering just those neonicotinoids affected by the moratorium
	(thiamethoxam, clothianidin and imidacloprid), the same effect is observed, with a significant
	decrease in residue concentrations in pollen (U 23, 22 = 389, p ˂0.001, mean 2013 = 5.02±11.32 ng/g,
	median ≤0.12 ng/g, mean 2015 =0.05±0.14 ng/g, median ˂MDL) but not nectar between 2013 and 2015
	2 1 =8.987, p=0.003, n=7) and (U 14, 19 = 140, p=0.676; mean 2013 = 0.20±0.51 ng/g, median ˂MDL, mean 2015 ˂MDL , median ˂MDL).
	nectar samples (80%, Nectar χ 2 1 =6.152, p=0.013, n=5) from peri-urban nests were contaminated In contrast, concentrations of thiacloprid, which was unaffected by the ban, increased significantly in
	(N=12), frequently with multiple residues (40% nectar samples, 29% of pollen). Again, thiacloprid nectar between 2013 and 2015 (U 14, 19 = 84, p= 0.013, mean 2013 ˂MDL, median ˂MDL,
	mean 2015 =0.09±0.15 ng/g, median ˂MDL). Concentrations of thiacloprid in pollen remained
	unchanged over this period (U 23, 22 =267, p=0.627, mean 2013 =0.08±0.31 ng/g, median ˂MDL, mean 2015
	Regional differences in exposure In 2013, pollen collected from bumblebee colonies in Sussex (SU)
	To compare the changing risk of exposure to peri-urban and rural bees over the transitional period was more frequently contaminated (χ 2 1 =15.62, p<0.001, Sussex=79%; Stirling=27%), with
	from pre-to post-moratorium, we compared residue concentrations in 2013 and 2015 for Sussex significantly higher concentrations of neonicotinoids than pollen collected from colonies in Stirling

(up to 9.32 ng/g in pollen, mean=1.34±3.52 ng/g, median≤ 0.04 ng/g) and thiamethoxam (up to 3.48 ng/g in pollen, mean= 0.76±1.52, median=0.10 ng/g) and were detected at the highest concentrations.

In 2015, the frequency of neonicotinoid detection was similar for nectar collected from rural and peri-urban bumblebee colonies in Sussex (χ 2 1 =0.158, p=0.691, Rural=47%, Peri-urban=33%) as were the concentrations present (Mann-Whitney U 19, 12 =130.5, p=0.469, mean R =0.10±0.15 ng/g, median R ˂MDL, mean PU =0.08±0.17 ng/g, median PU ˂MDL). While the frequency of detection (Rural=35%, Peri-urban=64%), proportion of samples with multiple residues (Rural=9% vs. Peri-urban=18%) and mean concentration of neonicotinoids were higher in pollen from peri-urban nests, the difference was not significant (χ 2 1 =1.238, p=0.266, U 22, 11 = 75.5, p=0.06, mean R =0.06±0.14 ng/g, median R ˂MDL, mean PU =1.29±3.30 ng/g, median PU ˂MDL). Both habitats were contaminated predominantly with thiacloprid (up to 0.44 ng/g, mean±SD=0.04±0.11 ng/g, median ˂MDL), and imidacloprid (up to 11.16 ng/g in peri-urban nests, mean±SD=0.21±1.40 ng/g, median ˂0.14), though a small proportion of peri-urban samples also contained acetamiprid (4% up to 1.4 ng/g, mean≤0.03 ng/g, median ˂MDL). bumblebee colonies only. For pollen collected from rural colonies there was a significant decrease in overall combined residue concentrations between years (Mann-Whitney U 23, 22 =385, p=0.002, mean 2013 = 5.10±11.40 ng/g, median ≤0.12 ng/g, mean 2015 =0.06±0.14 ng/g, median ˂MDL), but not for ACS Paragon Plus Environment < MDL, median ˂MDL). For peri-urban nests, there was no significant difference in overall residue concentrations in either pollen (U 19, 11 =124, p=0.408, mean 2013 = 2.11±4.56 ng/g, median=0.12 ng/g, mean 2015 =1.29±0.14 ng/g, median ≤0.04 ng/g) or nectar (U 13, 12 =62.5, p=0.276, mean 2013 = 0.02±0.05 ng/g, median ˂MDL, mean 2015 =0.08±0.17 ng/g, median ˂MDL), samples collected between 2013 and 2015. When considering either the banned neonicotinoids only (Pollen, U 19, 11 =134.5, p=0.188; mean 2013 = 0.63±1.64 ng/g, median ≤0.12, mean 2015 =1.14±3.33 ng/g, median ˂MDL; Nectar U 13, 12 = 76, p= 0.898, mean 2013 ˂MDL, median ˂MDL, mean 2015 ˂MDL, median ˂MDL) or thiacloprid, which was unaffected by the ban (Pollen U 19, 11 = 104, p= 1, mean 2013 = 1.47±4.41 ng/g, median ˂MDL, mean 2015 ˂MDL, median ˂MDL, Nectar U (ST) (Mann-Whitney U 42,26 =276, p<0.001; mean SU ±SD=3.74±9.01 ng/g, median SU ≤0.12 ng/g mean ST ±SD =0.20±0.49 ng/g, median ST ˂MDL). Nectar was contaminated at similar frequencies (Fisher's

Table 2 )

 2 Regional higher in Hertfordshire hives (U 44, 42 =1301, p≤0.001, mean H =0.88±0.81, median=0.75 ng/g, mean SU =0.40±0.80 ng/g, median ≤0.10 ng/g). Crop mapping of the five 5 km 2

	At the final sampling point in June, neonicotinoid concentrations detected in samples from
	both regions were below the limit of quantification, and were significantly lower than in May (Pollen
	Z 30 =0.55, p=0.003; Nectar Z 27 =0.73, p<0.001). The frequency of neonicotinoid detection in both
	pollen (30% samples, McNemar test, p=0.002) and nectar (34% samples, p=0.002) was also
	significantly lower than the previous month (

April and May, there was a similar frequency of neonicotinoid detection in both pollen (April= 90%, May=73%, McNemar test, p=0.287) and nectar (April=81%, May=80% p=0.760). While the concentration of neonicotinoid residues in pollen remained the same as the previous month (Wilcoxon signed-rank test, Z 30 =0.28, p=0.120, mean April =0.32±0.37 ng/g, median ≤0.12 ng/g mean May =0.22±0.33, median ≤0.12 ng/g), neonicotinoid concentrations in nectar, previously high in comparison to pollen, declined significantly between April and May (Z 26 =0.75, p<0.001; mean April =1.20±0.95 ng/g, median= 1.06 ng/g, mean May =0.65±0.72, median=0.27 ng/g). differences: While overall neonicotinoid concentrations in pollen contamination did not differ between Hertfordshire and Sussex (Mann-Whitney U 45, 45 =1014, p=0.100, mean H =0.23±0.36, median ≤0.12 ng/g, mean SU = 0.17±0.27, median ≤0.12 ng/g), concentrations in nectar were significantly

ACS Paragon Plus EnvironmentEnvironmental Science & Technology

ACS Paragon Plus Environment

Environmental Science & Technology Table 1 Number of honeybee and bumblebee colonies placed in each habitat type (Peri-urban vs. Rural), in each region (Sussex, Stirling, Hertfordshire (Herts)) across the three years of the study (2013)(2014)(2015). The specific dates colonies were sampled for pollen and nectar are listed.

ACS Paragon Plus Environment

Environmental Science & Technology ACKNOWLEDGEMENTS We are grateful to BBSRC (BB/K014498/1; BB/J014915/1) and Defra (Research Project PS2372) for funding this work and for the farmers and land-owners who allowed us to work on their property and shared their pesticide usage data. We thank Bill Hughes, Mark Crumpton, Steve Kennedy and Luciano Scandian for beekeeping equipment, assistance and advice, and Pete Kennedy and Martyn Stenning for useful discussions regarding experimental design and logistics. Thanks also to Julia ACS Paragon Plus Environment Environmental Science & Technology Horwood and Daniel Ingram for technical advice and assistance and to Grace Twiston-Davies for digitising crop maps.

Notes

The authors declare no competing financial interest.


Laure Gossec
email: laure.gossec@aphp.fr

Anna Moltó

Xavier Romand
email: xromand@chu-grenoble.frpgaudin@chu-grenoble.fr

Déborah Puyraimond-Zemmour

Matthieu Lavielle
email: matthieu.lavielle@wanadoo.fr

Catherine Beauvais
email: catherine.beauvais@sat.aphp.fr

Eric Senbel
email: eric.senbel@wanadoo.fr

René-Marc Flipo
email: rene-marc.flipo@chru-lille.fr

Sophie Pouplin
email: sophie.pouplin@chu-rouen.fr

Christophe Richez
email: christophe.richez@chu-bordeaux.fr

Anna Molto
email: anna.molto@aphp.fr

Alain Saraux
email: alain.saraux@chu-brest.fr

Maryse Mézières

Loriane Gutermann
email: loriane.gutermann@aphp.frmaxime.dougados@aphp.fr

Philippe Gaudin

Daniel Wendling
email: dwendling@chu-besancon.fr

Maxime Dougados

Maryse Mezieres
email: ircmezieres@gmail.com

Prof Laure Gossec

Recommendations for the assessment and optimization of adherence to disease-modifying drugs in chronic inflammatory rheumatic diseases: A process based on literature reviews and expert consensus

Keywords: adherence, compliance, rheumatoid arthritis, spondyloarthritis, recommendations

à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

1. Non-adherence to medications in chronic inflammatory rheumatic diseases is a frequent and deleterious issue. 2. Using an evidence-based approach followed by expert consensus, recommendations to assess and improve medication adherence in CIRDs have been developed. 3. Adherence should be assessed at each outpatient visit, at least using an open question. 4. Patient information and education, and patient/physician shared decision, are key to optimize adherence. 5. Formalized education sessions, motivational interviewing and cognitive behavioral therapy may be useful to improve adherence.

Introduction

Drug adherence is defined by the World Health Organization (WHO) as "the extent to which a person's behaviourtaking medication, following a diet, and/or executing lifestyle changes, corresponds with agreed recommendations from a health care provider" [START_REF]Adherence to long-term therapies: evidence for action[END_REF]. Drug adherence is usually conceived as drug intake and is usually considered satisfactory for adherence rates above 80% (meaning that more than 80% of the prescribed doses are absorbed by the patient) [START_REF] Treharne | Compliance' is futile but is 'concordance' between rheumatology patients and health attainable?[END_REF]. Adherence is different from persistence, which is defined as the length of time a specific drug is taken without interruption.

Depending on the definition of drug adherence, the methodology used and the drugs concerned, adherence is variable but overall insufficient [START_REF] Van Den Bemt | Medication adherence in patients with rheumatoid arthritis: a critical appraisal of the existing literature[END_REF][START_REF] Curtis | Adherence and Persistence with Methotrexate in Rheumatoid Arthritis: A Systematic Review[END_REF]. In 2003, the WHO reported that, in developed countries, 50% of patients with chronic disease were adherent to prescribed treatment [START_REF]Adherence to long-term therapies: evidence for action[END_REF]. In the field of rheumatic and musculoskeletal disorders, and in particular chronic inflammatory rheumatic diseases (CIRDs) such as rheumatoid arthritis, or connective tissue diseases, this non-optimal adhesion is confirmed. In these diseases, adherence to disease-modifying drugs is only moderate over the long term. Adherence has been mostly studied in RA and reported rates of adherence are 30 to 80% [START_REF] Van Den Bemt | Medication adherence in patients with rheumatoid arthritis: a critical appraisal of the existing literature[END_REF][START_REF] Scheiman-Elazary | The rate of adherence to antiarthritis medications and associated factors among patients with Rheumatoid Arthritis: A Systematic Literature Review and Metaanalysis[END_REF]. Non-optimal patient adherence compromises therapeutic efficacy. In RA in particular, poor adherence has been shown to be linked to increased disease activity (6)(7)[START_REF] Contreras-Yáñez | Inadequate therapy behavior is associated to disease flares in patients with rheumatoid arthritis who have achieved remission with disease-modifying antirheumatic drugs[END_REF][START_REF] Bluett | Impact of inadequate adherence on response to subcutaneously administered anti-tumour necrosis factor drugs: results from the Biologics in Rheumatoid Arthritis Genetics and Genomics Study Syndicate cohort[END_REF]. Furthermore, non-adherence may lead to complications, unnecessary treatment switches and heightened costs [START_REF] Sokol | Impact of medication adherence on hospitalization risk and healthcare cost[END_REF].

Thus, adherence is a key issue when dealing with chronic patients and there is a need for new care models addressing the problem of medication adherence, integrating this problem into the patient care process.

However, several questions remain unanswered. Firstly, assessment of non-adherence is an issue. Physicians tend to overestimate how well patients take their medication as prescribed.

This can lead to missed opportunities to change medications, solve adverse effects, or propose the use of reminders in order to improve patients' adherence. Thus, better assessment of non-adherence would be useful. Adherence to treatment can be assessed by several methods, direct (such as blood tests) and indirect (such as questionnaires) [START_REF] Zaugg | Providing physicians with feedback on medication adherence for people with chronic diseases taking long-term medication[END_REF].

However to date, no gold standard method has been defined and clinicians may feel at a loss on how and when to measure adherence in their patients [START_REF] Lehmann | Assessing medication adherence: options to consider[END_REF][START_REF] Hansen | Comparison of methods to assess medication adherence and classify nonadherence[END_REF]. Secondly, as patients' lack of adherence to treatments is multifactorial, it would be useful to better understand the determinants of non-adherence [START_REF] Pasma | Factors associated with adherence to pharmaceutical treatment for rheumatoid arthritis patients: a systematic review[END_REF][START_REF] Voshaar | Barriers and facilitators to disease-modifying antirheumatic drug use in patients with inflammatory rheumatic diseases: a qualitative theory-based study[END_REF][START_REF] Wong | Medication adherence in patients with rheumatoid arthritis: why do patients not take what we prescribe?[END_REF][START_REF] Delestras | Comparison between two generic questionnaires to assess satisfaction with medication in chronic diseases[END_REF]. And finally, a better understanding of interventions

to improve adherence and their efficacy is needed [START_REF] Salt | Adherence to disease-modifying antirheumatic drugs in patients with rheumatoid arthritis: a narrative review of the literature[END_REF].

The objective of the present initiative was to develop recommendations to facilitate in daily practice, the management of non-adherence to disease-modifying drugs, in patients with CIRDs.

Methods

This process included literature reviews and a consensus process in France, in accordance with previous Rencontres d'Experts en Rhumatologie (RER) and 3E (Evidence, Expertise, Exchange) initiatives [START_REF] Gossec | Collection and management of selected comorbidities and their risk factors in chronic inflammatory rheumatic diseases in daily practice in France[END_REF][START_REF] Gossec | Dissemination and evaluation of the 3E initiative recommendations for use of methotrexate in rheumatic disorders: results of a study among 2233 rheumatologists[END_REF] Of note, the recommendations were developed by a group of rheumatologists from a single country (France), the steering committee was issued from 9

university hospitals and one private practice, and the project was conducted thanks to an unrestricted grant from Abbvie France.

Decisions on target population and target aspects of adherence

A face-to-face meeting of the steering group took place in October 2016. The group included a convenor (MD), 2 methodologists (LG and AM), 3 fellows for the literature review (XR, DP and ML), 9 rheumatologist experts, one pharmacist and one rheumatology nurse. The steering group decided to elaborate these recommendations with the objective to facilitate the daily practice of health care providers in charge of this aspect of patient's management (e.g. nurses, general practitioners, pharmacists and rheumatologists). It was decided to limit the project scope to CIRDs, defined here as RA, spondyloarthritis (SpA), psoriatic arthritis (PsA), connective tissue diseases as well as some other inflammatory diseases such as crystal-induced arthritis, vasculitis and auto-inflammatory diseases. In terms of treatments, the project scope was limited to disease-modifying anti-rheumatic drugs, DMARDs (defined here as mainly conventional synthetic DMARDs, biologics and targeted synthetic DMARDs, but also checking the literature for data on glucocorticoid intake, non-steroidal drugs in SpA, and uric-lowering drugs as well as colchicine for crystal-induced arthritis). It was also decided to focus on 3 aspects of adherence, namely how to assess it, which patients to target more specifically for assessment (i.e., risk factors of non-adherence), and interventions to improve adherence (supplementary table 1).

The process then comprised 3 steps.

Systematic literature reviews

Between 1).

Data were extracted and meta analyses were considered but given heterogeneity, were not performed [START_REF] Lavielle | Methods to improve medication adherence in patients with chronic inflammatory rheumatic diseases: a systematic literature review[END_REF]. The most relevant information to answer each main question was discussed and reviewed with the steering committee and presented in a condensed form during the consensus process. Consensus process

In October 2017, during a 2-day face-to-face meeting, recommendations were developed through consensus of 104 rheumatologists/pharmacists/nurse experts (with a high predominance of rheumatologists). The final recommendations were anonymously evaluated by the participants for agreement and ease of applicability (on a Likert scale 1-5 were 5 is highest).

Dissemination, evaluation, implementation and update

For the dissemination and evaluation steps, the steering committee decided that the results of this initiative would be evaluated and disseminated via small, regional, face to face meetings throughout France over one year. Finally, these recommendations should be updated systematically after a 5 years period or before in case some new relevant information in this area is available sooner.

Results

The consensus process led to 5 overarching principles and 10 recommendations regarding adherence (Table 1). The French version is presented as Online supplementary Table 1.

There was high agreement within the Task Force regarding these points: the agreement with the recommendations was high (range of means, 3.9-4.5 on a 1-5 scale) but ease of applicability was lower (range of means, 2.7-4.4).

Overarching principles (Table 1)

A-Drug adherence covers 2 complementary notions: compliance, i.e., treatment intake as prescribed, and persistence, i.e., maintenance of intake over time.

This overarching principle reminds the reader of the definition of drug adherence. Drug adherence refers to both a notion of drug intake and of long-term adherence, as recognised by the WHO [START_REF]Adherence to long-term therapies: evidence for action[END_REF][START_REF] Brown | Medication Adherence: WHO Cares?[END_REF]. This point is for information and was not the subject of debate.

B-Non-adherence to disease-modifying anti rheumatic drugs is frequent. It can be detrimental, leading to lower drug efficacy and potential cost increases.

This principle is again informative and was not debated. Many studies have shown that nonadherence is a detrimental process: it can lead to periods of disease intensification which in turn are detrimental [START_REF] Ho | Clinical and economic impact of non-adherence to antidepressants in major depressive disorder: A systematic review[END_REF]. In several chronic diseases, the management of patients not adhering to their treatment is associated with higher health costs compared to that of adherent patients [START_REF] Sokol | Impact of medication adherence on hospitalization risk and healthcare cost[END_REF]. In RA, poor patient adherence can compromise treatment effectiveness, decrease quality of life and appears to increase health care costs by around 33% [START_REF] Vera | Economics of non-adherence to biologic therapies in rheumatoid arthritis[END_REF][START_REF] Pasma | Does non-adherence to DMARDs influence hospital-related healthcare costs for early arthritis in the first year of treatment?[END_REF].

C-In non-adherence, factors known as "unintentional" (simply forgetting, ...) and "intentional"

(linked to the patient's beliefs and fears, ...), are often intertwined.

This principle refers to causes of non-adherence and was again for explanatory purposes.

Patients often cite unintentional causes of non-adherence such as having 'forgotten' to take the drug. But health professionals should be aware that this may 'hide' other reasons for nonadherence.

D-Knowledge both of the disease and of the treatment, and patients' perceptions of the benefit/risk of the treatment are key elements in drug adherence.

This overarching principle and the next one both introduce later recommendations on how to improve adherence since they refer to causes of non-adherence. Patient beliefs play a key role in non-adherence [START_REF] Horne | Understanding patients' adherence-related beliefs about medicines prescribed for long-term conditions: a meta-analytic review of the Necessity-Concerns Framework[END_REF]. Studies have shown that non-adherence is more frequent if the perceived necessity of a drug is lower than its perceived risk [START_REF] Horne | Understanding patients' adherence-related beliefs about medicines prescribed for long-term conditions: a meta-analytic review of the Necessity-Concerns Framework[END_REF].

E-In the context of shared decision-making/therapeutic alliance, caregiver-patient communication about treatment is a key factor in drug adherence.

Given the elements above and also as a separate item, patient-physician interactions play an important role in non-adherence. All studies on adherence across diseases point out the importance of the quality of the patient-physician relationship and communication around drugs. Empathy also plays a role, some studies having indicated that empathy can contribute to better disease control [START_REF] Cobos | White coat hypertension: improving the patient-health care practitioner relationship[END_REF].

Thus, adherence is a notion which refers to therapeutic alliance, a key notion in chronic disease management.

Recommendations (Table 1)

When and how to assess adherence? (recommendations 1-4)

1. Adherence should be assessed at each patient visit. It must be systematic if the treatment target is not reached and before any therapeutic change.

Healthcare providers are often unaware of adherence problems for their patients. Thus, providing physicians with feedback on medication adherence has the potential to prompt changes that improve their patients' adherence to prescribed medications. A recent Cochrane review did not evidence improvements in patient outcomes in studies with provision of feedback to physicians regarding their patients' adherence to prescribed medication [START_REF] Zaugg | Providing physicians with feedback on medication adherence for people with chronic diseases taking long-term medication[END_REF]. However, the group felt that being aware of adherence and communicating about this issue with patients was key.

The consensus was that adherence should be assessed regularly, ideally at each outpatient visit. If this cannot be done, adherence should at least be assessed when the disease state is unsatisfactory, i.e., when the treatment target has not been reached and when a treatment change is being considered [START_REF] Smolen | Treating rheumatoid arthritis to target: 2014 update of the recommendations of an international task force[END_REF][START_REF] Smolen | Treating axial spondyloarthritis and peripheral spondyloarthritis, especially psoriatic arthritis, to target: 2017 update of recommendations by an international task force[END_REF].

2. Adherence should be evaluated during outpatient visits by at least one open question.

The literature on assessment of adherence was mainly issued from RA studies and questionnaires were found to be most frequently used in studies (online supplementary Table 2). The questionnaires are discussed in Recommendation 2. The group felt it was not possible to reach consensus on a single questionnaire to be used in everyday practice, and furthermore assessment of adherence using a questionnaire was not thought to be feasible in the clinic for every patients. Thus the consensus was to use an open question though this was not data-driven. A minimal assessment of non-adherence can be obtained through an open question such as 'how did it go with your treatment since the last visit?'. Examples of open questions proposed by the steering committee, can be seen in Table 2.

3. The assessment of adherence, particularly in the context of multidisciplinary care, can be carried out by more complete methods than an open question alone (self-reported questionnaires, dispensation data, etc.).

When the situation allows it (e.g. in a day hospitalization or when multidisciplinary care is provided), more complete questionnaires may be useful. The literature review allowed us to compare different questionnaires and in particular the Compliance Questionnaire on Rheumatology (CQR), Medication Adherence Report Scale (MARS), Morisky Medication

Adherence Scale (MMAS) and Medication Adherence Self-report Inventory (MASRI), as seen in online supplementary Table 2. The discussion focused on the MMAS which has been widely used in the literature across several chronic diseases, but given that it is copyrighted, not validated for use in RA, and the items were found to be repetitive, no consensus was reached on which questionnaire should be put forward [START_REF] Morisky | Predictive validity of a medication adherence measure in an outpatient setting[END_REF].

4. Adherence to hydroxychloroquine can be verified by a blood test and explaining the results to the patient can improve adherence.

In some cases, other ('more objective') assessments of adherence can be applied. Where available easily, patient-level dispensation data (available in some countries through the health insurance records) may allow to check it the patient has been given his/her drugs.

Pharmacists may contribute to this assessment of adherence, as also explained below (31).

For hydroxychloroquine, adherence can be verified by a blood test and explaining the results to the patient can improve adherence, as has been shown in lupus [START_REF] Costedoat-Chalumeau | Hydroxychloroquine in systemic lupus erythematosus: results of a French multicentre controlled trial (PLUS Study)[END_REF].

Factors associated to non-adherence (recommendation 5)

5. When assessing adherence, risk factors for nonadherence should be examined, in particular those related to the patient (young subject, fear of side effects, mood disorders,...), treatment (polymedication,...) and environment (caregiver-patient relationship, ...).

The literature review evidenced multiple factors associated to non-adherence, with differing results across studies and across diseases, making the interpretation of the findings difficult.

Overall, around 240 different factors related to adherence were evidenced in the literature though they could be broadly regrouped as patient-related, treatment-related and patienthealthcare provider relationship related [START_REF] Michetti | Impact of Treatment-Related Beliefs on Medication Adherence in Immune-Mediated Inflammatory Diseases: Results of the Global ALIGN Study[END_REF][START_REF] Betegnie | Why Do Patients with Chronic Inflammatory Rheumatic Diseases Discontinue Their Biologics? An Assessment of Patients' Adherence Using a Self-report Questionnaire[END_REF][START_REF] Zwikker | Perceived need to take medication is associated with medication non-adherence in patients with rheumatoid arthritis[END_REF][START_REF] Curtis | Patient's experience with subcutaneous and oral methotrexate for the treatment of rheumatoid arthritis[END_REF]. It was difficult to reach a consensus on this part of the project. For this reason, it was proposed (see recommendation 1) to screen all patients for non-adherence. However, particular attention should be given to patients who have characteristics corresponding to the 'non-adherent profile' (Table 3): people who are younger, worried of side effects, do not see the necessity of the treatment, and are in psychological distress are more prone to non-adherence.

Interventions to improve adherence (recommendations 6-10)

All studies reporting the results of interventions to improve adherence in CIRDs were analysed in the systematic literature review [START_REF] Lavielle | Methods to improve medication adherence in patients with chronic inflammatory rheumatic diseases: a systematic literature review[END_REF]. Interventions were classified as educational, behavioural, cognitive behavioural or multicomponent interventions [START_REF] Greenley | Practical Strategies for Enhancing Adherence to Treatment Regimen in Inflammatory Bowel Disease[END_REF]. In all, 22 studies were analysed: most (18) were performed on RA patients. The results were mitigated since of 13 randomized controlled trials (1535 patients), only 5 were positive (774 patients) (10, 24, 25).

6. In order to optimize drug adherence, the patient should be an actor in his disease and his care within the framework of a shared decision (therapeutic alliance). This is a generic recommendation which refers to the same concept as ocverarching principles D and E. The literature indicates a link between patient involvement and drug adherence though the data is observational rather than in the format of randomized trials.

However, the experts felt this was a key point which should be emphasized.

7. In order to optimize drug adherence, any prescription for antirheumatic treatment must be accompanied by patient information and education.

In the literature, educational interventions had the highest level of evidence, corresponding to 8 of the 13 randomized controlled trials and 4 (50%) were positive. Briefly, educational interventions aim to enhance patient knowledge of the disease, the benefits and mechanisms of action of the medication regimen, the consequences of non-adherence, and potential side effects of treatment. The expert consensus was that given that not all patients need formalised, organised therapeutic education and not all patients have access to therapeutic education, the recommendation should be strong for the encouragement to educate patients (ie the term 'must') but should be inclusive of all formats of information and education, rather than limited to formalised patient education (ie the inclusive term of 'patient information and education).

Recommendations 6 and 7 reflect that patient information and education, and patient/physician shared decision, are key to optimize adherence and should be systematic.

8. The detection of nonadherence to medication must lead to the implementation of a specific intervention (therapeutic education, motivational interview, cognitive behavioural methods, etc.) to improve adherence.

In the literature, on top of patient therapeutic education, other interventions such as behavioural interventions, cognitive behavioural interventions or motivational interviewing

were also assessed [START_REF] Lavielle | Methods to improve medication adherence in patients with chronic inflammatory rheumatic diseases: a systematic literature review[END_REF]. In some cases, the strategies shown to be effective were complex and difficult to implement in clinical practice.

The consensus was that other techniques such as formalized education sessions, motivational interviewing and cognitive behavioral therapy may be useful although it was not possible to hierarchize these interventions or to define profiles of patients who would potentially most benefit from one or the other.

9. The patient information and education process, individual or collective, must be carried out repeatedly by one or several health professionals (doctors, pharmacists, specialized nurses...) alone or in a team.

In the published studies of patient education, patients were informed or educated by different actors (physicians, pharmacists, nurses…).( 21) Given this literature and given also feasibility issues (not all rheumatologists have access to formalized group education sessions for their patients), it was felt the recommendation should reflect that all education processes are acceptable. However, we wanted to emphasize that all health professionals should be involved in adherence. In particular, apart from rheumatologists, healthcare teams including nurses are important. Pharmacists will increasingly be involved as well. Advice by pharmacists on drug management has been shown to promote drug adherence, both in rheumatology and in other chronic disease settings [START_REF] Ryan | Interventions to improve safe and effective medicines use by consumers: an overview of systematic reviews[END_REF][START_REF] Kulchaitanaroaj | Cost-utility analysis of physician-pharmacist collaborative intervention for treating hypertension compared with usual care[END_REF]. The role of pharmacists may vary of course according to the healthcare system.

10. The patient information and education process can be supplemented by tools such as brochures and multimedia to improve therapeutic adherence.

In the published studies, supports and contents of the educational interventions were heterogeneous. ( 21) Thus the recommendations note the possibility to use different supporting materials in the education process.

Over the past years, the group felt e-health has taken on increasing importance. Patients seek information online and websites or apps can participate in promoting adherence [START_REF] Bosworth | Health Information Technology: Meaningful Use and Next Steps to Improving Electronic Facilitation of Medication Adherence[END_REF].

The group was not able to direct patients towards a specific online resource but it was reflected in the recommendations that e-health might be of value.

Discussion

Using an evidence-based approach followed by expert consensus, the current work has allowed the development of pragmatic recommendations regarding the assessment and management of non-adherence in CIRDs. This initiative should improve the assessment and optimization of adherence in daily practice.

Adherence is a key issue in chronic diseases but strangely has not perhaps received up to now, the focus it deserves. Non-adherence is frequent and costly at the patient level and at the society level. In its 2003 report on medication adherence, the World Health Organization quoted the statement by Haynes et al that "increasing the effectiveness of adherence interventions may have a far greater impact on the health of the population than any improvement in specific medical treatments" (1). Indeed, among patients with chronic illness, approximately 50% do not take medications as prescribed [START_REF]Adherence to long-term therapies: evidence for action[END_REF][START_REF] Treharne | Compliance' is futile but is 'concordance' between rheumatology patients and health attainable?[END_REF][START_REF] Van Den Bemt | Medication adherence in patients with rheumatoid arthritis: a critical appraisal of the existing literature[END_REF][START_REF] Curtis | Adherence and Persistence with Methotrexate in Rheumatoid Arthritis: A Systematic Review[END_REF] This poor adherence to medication leads to increased morbidity and death and increased costs [START_REF] Sokol | Impact of medication adherence on hospitalization risk and healthcare cost[END_REF][START_REF] Cutler | Economic impact of medication non-adherence by disease groups: a systematic review[END_REF]. However, today many more publications are found centred on the prescription of a drug (e.g., management recommendations) than on the intake of the drug (i.e., adherence). It is possible that this relative lack of interest is due to the difficult situation which it puts the physician in, or the fact that many healthcare professionals persist in seeing the nonadherent patient as a problematic patient (rather than, perhaps a problematic prescription) [START_REF] Fung | Patient-reported adherence to statin therapy, barriers to adherence, and perceptions of cardiovascular risk[END_REF][START_REF] Kvarnström | Barriers and facilitators to medication adherence: a qualitative study with general practitioners[END_REF][START_REF] Suh | Medication nonadherence in Korean patients with rheumatoid arthritis: the importance of belief about medication and illness perception[END_REF]. In the present work, we felt that a focus on adherence, with the view to develop practical and helpful recommendations, was needed.

The present study has strengths and weaknesses. Well-validated procedures to develop recommendations were followed [START_REF] Gossec | Collection and management of selected comorbidities and their risk factors in chronic inflammatory rheumatic diseases in daily practice in France[END_REF][START_REF] Gossec | Dissemination and evaluation of the 3E initiative recommendations for use of methotrexate in rheumatic disorders: results of a study among 2233 rheumatologists[END_REF][START_REF] Van Der Heijde | Update of the EULAR standardised operating procedures for EULAR-endorsed recommendations[END_REF]. However, categories of evidence and grades of recommendations were not determined, which is a limitation. Systematic literature reviews were performed and the consensus process allowed everyone's voice to be heard. However, it was not possible to include patients in the process for regulatory reasons. For this reason, the process was followed with a dissemination process among patient associations in France, which is ongoing. Although a strict process was followed, it was in some cases difficult to develop precise and applicable recommendations. This was the case in particular when trying to identify patient characteristics related to non-adherence [START_REF] Michetti | Impact of Treatment-Related Beliefs on Medication Adherence in Immune-Mediated Inflammatory Diseases: Results of the Global ALIGN Study[END_REF][START_REF] Betegnie | Why Do Patients with Chronic Inflammatory Rheumatic Diseases Discontinue Their Biologics? An Assessment of Patients' Adherence Using a Self-report Questionnaire[END_REF][START_REF] Zwikker | Perceived need to take medication is associated with medication non-adherence in patients with rheumatoid arthritis[END_REF][START_REF] Curtis | Patient's experience with subcutaneous and oral methotrexate for the treatment of rheumatoid arthritis[END_REF]. Indeed, it appears it is impossible to define a single non-adherent patient profile. It is possible that nonadherence is more related to a moment / a prescription than to a patient profile; or that all these elements play a part. For this reason, we feel non-adherence should be assessed systematically. It was also not possible to recommend a single questionnaire to assess adherence, since all available questionnaires were found to be unsatisfactory for the context of usual care [START_REF] Morisky | Predictive validity of a medication adherence measure in an outpatient setting[END_REF]. The inclusive nature of our group is a strength, however, the funding of the project by pharmaceutical industry could be perceived as a pontetial weakness, though adherence was addressed as a generic concept rather than drug-related, throughout the process.

Dissemination and implementation of recommendations is often an issue. In the present case, dissemination was planned in the project from the start through face to face meetings where the literature and recommendations are shared with rheumatologists nationwide. Even so, implementation during patient visits should be further assessed.

Some overlap can be noted between some of the overarching principles, and some of the recommendations which deal with shared decision-making. The group felt strongly that the implication of patients as actors and shared decision making were such key points when discussing adherence, that they had to be emphasized. Shared decision making is still an ongoing subject of research. Overall, we felt interventions to improve drug adherence were disappointing at the group level, as has been also found by other authors [START_REF] Zaugg | Providing physicians with feedback on medication adherence for people with chronic diseases taking long-term medication[END_REF][START_REF] Salt | Adherence to disease-modifying antirheumatic drugs in patients with rheumatoid arthritis: a narrative review of the literature[END_REF][START_REF] Lavielle | Methods to improve medication adherence in patients with chronic inflammatory rheumatic diseases: a systematic literature review[END_REF][START_REF] Ryan | Interventions to improve safe and effective medicines use by consumers: an overview of systematic reviews[END_REF][START_REF] Galo | What are the effects of medication adherence interventions in rheumatic diseases: a systematic review[END_REF][START_REF] Depont | Interventions to Improve Adherence in Patients with Immune-Mediated Inflammatory Disorders: A Systematic Review[END_REF]. There are several potential explanations: perhaps interventions are not targeting the right patients; perhaps the interventions are not sufficiently tailored. Perhaps also nonadherence, due to its multifactorial nature, is difficult to act upon [START_REF] Fung | Patient-reported adherence to statin therapy, barriers to adherence, and perceptions of cardiovascular risk[END_REF][START_REF] Kvarnström | Barriers and facilitators to medication adherence: a qualitative study with general practitioners[END_REF][START_REF] Suh | Medication nonadherence in Korean patients with rheumatoid arthritis: the importance of belief about medication and illness perception[END_REF]. Overall, nonpharmacological interventions pose specific methodological problems thus the mitigated results of trials should be interpreted with caution [START_REF] Page | Assessing risk of bias in studies that evaluate health care interventions: recommendations in the misinformation age[END_REF].

As treatment options become more numerous in the field of CIRDs, in a financially-limited context in many countries, adherence to medication should become a priority to improve the quality of care for patients with chronic diseases.

Data extraction

Data were collected by one person using a specific CRF developed for this study. 3 fellows performed the literature review each one dealing with one of the subjects described in the research question: 1Methods used to assess adherence and psychometric properties of each method; 2: Factors shown to be associated to low adherence; 3: Efficacy of interventions: For all studies, we collected data regarding the design of the studies: randomized or non randomized, controlled or not, length of follow up, number of patients in each group (intervention and control). Characteristics of the population were also collected: disease studied, disease duration, activity of the disease, age at baseline, gender, treatment studied.

The type of measurement used for adherence and adherence results were also collected.

We then collected for subject 3, information on the intervention under evaluation: type of intervention aiming to improve medication adherence (educational…), main components and supports of this intervention, actors of the intervention, patients targeted by the intervention (systematic for all patients or targeted on patients considered to be non-adherent or at high risk of non-adherence). Tools used to measure adherence were noticed and finally, results on adherence were collected in each group of patient if available.

•

Figure 1 .

 1 Figure 1. Systematic literature review: flowchart

supplementary Table 1).

	searched were performed in CIRDs but were also enlarged as non systematic reviews, to
	cover other diseases where non-adherence is an issue, namely diabetes, high blood
	pressure, HIV infection and osteoporosis.
	A systematic review of the literature in CRIDs was performed according to the Cochrane
	guidelines The data were issued from PubMed Medline, Embase, Cochrane central register
	of clinical trials; up to February 2017. Associations of key words around the disease names
	and "medication adherence" or "patient compliance" were used (The search was completed using congress abstracts from the American and European
	congresses of 2016 and 2017; and several websites were searched: Clinicaltrials.gov, World
	Health Organisation and French High Authority of Health. Finally, hand search was
	performed using the references of the most relevant studies provided by the initiative's
	scientific committee of experts in the field. Inclusion criteria were studies of adults, published
	in English or in French, on the diseases considered which were: RA, SpA, PsA, connective
	tissue diseases including systemic lupus, crystal related diseases including gout and
	chondrocalcinosis, vasculitis including ANCA-associated vasculitis, giant cell arteritis and
	polymyalgia rheumatica, and Still's disease. Pharmacological medications considered were:
	conventional synthetic DMARDs, biological DMARDs, immunosuppressive drugs
	(cyclophosphamide…), non-steroidal anti-inflammatory drugs in SpA, glucocorticoids,
	colchicine and urate-lowering therapy. Articles were screened and all studies fitting the
	inclusion criteria and related to methods to measure non-adherence, risk factors classified in
	5 domains according to WHO and management options for non-adherence with their
	reported efficacy classified in 5 modalities (educational, behavioral, cognitive behavioral,
	multicomponent interventions or others) were selected. The flow chart shows this selection
	process (figure
	December 2016 and February 2017, 3 fellows performed systematic reviews of
	methods (including questionnaires) to measure non-adherence, risk factors for non-
	adherence and management options for non-adherence with their reported efficacy. The

Table 1 . Overarching principles and recommendations regarding drug adherence in

 1

	CIRDs
	Overarching principles

Table 2 . Examples of open questions to assess adherence

 2 How did it go when taking your treatment since the last visit? What difficulties did you meet when taking your treatment since the last visit??

	How is it going with your medication?
	How do you feel regarding your treatment?
	How do you manage not to forget to take your treatment?
	I have the impression that you have issues with this treatment
	I see that the treatment is less effective, what do you think? Maybe there is a problem with
	the way you take your medication?
	What do you do when you forget to take your treatment?

Table 3 . Profile of the 'non-adherent' patient according to the literature Factor Characteristic

 3

	Age	Young age
	Beliefs and knowledge	Concerns about side effects
		Low perceived need of treatment
		Low knowledge regarding disease
	Treatments	Polymedication
	Comorbidities	Presenting with comorbidities (in RA and SpA) or not (in
		gout and lupus)
	Psychological status	Mood disorders
	Patient physician relationship Poor doctor-patient relationship

Maxime DOUGADOS has received honorarium fees from Abbvie for his participation as the convenor of this initiative. Xavier ROMAND, Déborah PUYRAIMOND-ZEMMOUR and Matthieu LAVIELLE have received honorarium fees from Abbvie for their participation as fellows of this initiative. All the other coauthors have received honoraria from Abbvie as members of the scientific committee.

This study was conducted thanks to an unrestricted grant from

Supplementary data Table 1. Details of the methods of the systematic literature review

Methodologystep

Information

Choice

of diseases RA, SpA, PsA, connective tissue diseases including systemic lupus, crystal related diseases including gout and chondrocalcinosis, vasculitis including ANCA-associated vasculitis, giant cell arteritis and polymyalgia rheumatica, and Still's disease.

Choice of treatments

Pharmacological medications considered were: conventional synthetic DMARDs, biological DMARDs, immunosuppressive drugs (cyclophosphamide…), non-steroidal anti-inflammatory drugs in SpA, glucocorticoids, colchicine and urate-lowering therapy

Research questions

WSubject 1

WWhat tools are used to easily assess adherence by health care professionals in CIRDs? What are their performances? Check for questionnaires (specific? generic? from daily practice?), scores, dispensation data, pharmaceutical files. What are their validity? sensitivity? ease of use? Performance of generic vs. specific questionnaires?

Risk of bias assessment

Given the observational nature of almost all the studies, is was not possible to assess risk of bias.

Data analysis

Analysis was descriptive Supplementary data Table 2 The + represent a semi quantitative summary of the available literature with more + meaning higher / better results (from O to +++)