Félicie Costantino

Léa Bahier

Luis Coronel Tarancón

Ariane Leboime

François Vidal

Lamouri Bessalah

Maxime Breban

Maria Antonietta D'agostino

Maria-Antonietta D'agostino
email: maria-antonietta.dagostino@aphp.fr

Maria Antonietta

D ' Agostino

COVID-19 in French patients with chronic inflammatory rheumatic diseases: Clinical features, risk factors and treatment adherence

Keywords: COVID-19, spondyloarthritis, psoriatic arthritis, rheumatoid arthritis, nonsteroidal anti-inflammatory drugs

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

Introduction

Since the beginning of the current COVID-19 pandemic, concerns have arisen about patients with immune mediated inflammatory diseases, in particular chronic inflammatory rheumatic diseases (CIRDs) [START_REF] Pope | What Does the COVID-19 Pandemic Mean for Rheumatology Patients?[END_REF][START_REF] Richez | Implications of COVID-19 for the management of patients with inflammatory rheumatic diseases[END_REF]. Indeed, patients with CIRDs are known to have an increased risk of infection, in particular viral infections, because of immune dysregulation caused by the disease itself and also linked to their comorbidities and immune-modulating treatments [START_REF] Gabriel | Epidemiological studies in incidence, prevalence, mortality, and comorbidity of the rheumatic diseases[END_REF].

With the growing knowledge on the COVID-19, several risk factors have been identified including male sex, age over 65, severe obesity, hypertension, diabetes, cardiovascular diseases, chronic kidney disease, liver disease and chronic respiratory diseases [START_REF] Zheng | Risk factors of critical & mortal COVID-19 cases: A systematic literature review and meta-analysis[END_REF], which may cause increased incidence of SARS-CoV-2 infection or worse outcomes in case of COVID-19. These risk factors are frequently increased in CIRDs, in particular cardiovascular comorbidities. Uncertainties on the possibility of an increased risk of COVID-19 due to the rheumatic disease in itself with associated comorbidities still exist. However, last reports have shown that patients with CIRDs do not seem to present an increased incidence of COVID-19, neither a worse outcome, in case of infection, as compared to the general population [START_REF] Favalli | Incidence of COVID-19 in patients with rheumatic diseases treated with targeted immunosuppressive drugs: what can we learn from observational data? Arthritis & Rheumatology[END_REF][START_REF] Rosenbaum | Biologics, spondylitis and COVID-19[END_REF][START_REF] Quartuccio | Prevalence of COVID-19 among patients with chronic inflammatory rheumatic diseases treated with biologic agents or small molecules: A population-based study in the first two months of COVID-19 outbreak in Italy[END_REF].

Management of CIRDs patients in pandemic periods may vary according to countries guidelines, severity of the disease, presence of comorbidities and lastly according to patients' fears, making sometimes challenging and conflicting priorities for these patients: should they interrupt immunomodulatory drugs to decrease the risk of viral infection, with the risk of disease flare? Should they attend health care visits with the risk of being infected?

We therefore aimed at exploring how patients with CIRDs coped with their disease during the lockdown period, and to identify possible predictive factors of SARS-CoV-2-infection and the global burden of COVID-19 on these patients.

Methods

Study population

We focused on the three more frequently seen CIRDs: spondyloarthritis (SpA), rheumatoid arthritis (RA) and psoriatic arthritis (PsA). The study population came from two recruitment procedures. On one hand, all adult (>18 years-old) patients seen since 2019 in the rheumatology department of Ambroise Paré Hospital in Paris close suburb (either in outpatient or inpatient clinics) with a diagnosis of SpA, RA or PsA were asked to complete a questionnaire focused on their experiences around COVID19 symptoms, testing and medication access during the lockdown period. The questionnaire was sent via an e-mail on April 18, 2020 and responses were collected until May 21, 2020. On the other hand, the same questionnaire was proposed during the same period to all patients registered on the Spondy+® platform, a secure e-health platform for SpA patients developed to give them update information on their disease, to allow them to self-monitor disease activity and possibility to participate to research studies. The Spondy+® patients participating to our survey have previously provided a medical certificate from their rheumatologist or general practitioner confirming the SpA diagnosis.

The study was conducted according to the current regulations of the International Conference on Harmonization guidelines and the principles of the Declaration of Helsinki.

Study variables -COVID-19 survey

Participants were asked to fill a questionnaire of 39 questions divided in several sections: sociodemographic characteristics, disease characteristics and treatments, comorbidities, presence of symptoms suggesting viral infection, eventual confirmed diagnosis of COVID-19 (nasopharyngeal swan or CT-scan), patient's contacts with subjects diagnosed with COVID-19, adherence to the ongoing rheumatological therapy and any changes in their rheumatology treatment plan since the beginning of the COVID-19 pandemic, as well as reported changes on their disease activity or on the management of their follow up visits [Appendix A ; See the supplementary material associated with this article online].

Statistical analyses

Statistical analysis was performed using R version 3.6.1 (http://www.r-project.org/).

Results

Study population.

We were able to contact by email 948 (83.7%) of the 1133 RA, SpA and PsA patients seen in 2019 in our Rheumatology department, of whom 429 (45.3%) responded to the questionnaire.

We also obtained 251 (17.1%) answers from the 1471 Spondy+® patients. As 25 patients from the Hospital have also answered to the questionnaire on Spondy+®, we finally obtained 655 unique answers (Figure 1). Factors independently associated with the non-response to the questionnaire were age (younger), SpA diagnosis and recruitment through Spondy+® platform (Appendix A, table S1).

Demographic and clinical characteristics of the included patients are detailed in Table 1. The population was predominantly female (61.8%) with a mean (± standard deviation) age of 51.0 ± 13.4 years. Most of the patients were from European ancestry (87.3%) and lived in Paris or in its suburban area (64.6%). Geographical distribution of the responders is shown in Appendix A, figure S1. Frequency of SpA (72.3%) was higher than that of RA (19.7%) or PsA (8.0%) and the mean disease duration was 12.5 ± 10.2 years. Anti-inflammatory drugs were taken by 62% of the patients (non-steroidal anti-inflammatory drugs (NSAIDs): 45.8%, corticosteroids: 16.3%). The vast majority of them (72.8%) were treated with disease modifying anti-rheumatic drug (DMARDs): conventional DMARDs (cDMARDs): 24.12%, biological DMARDs (bDMARDs): 61.7%, targeted synthetic DMARDs: 1.5%. Only a minority presented some comorbidity (35.6%), high blood pressure being the most frequent (18.6%).

SARS-CoV-2 exposure, symptoms and diagnoses

Approximately one third of the patients reported a possible or confirmed exposure to SARS-CoV-2. The most frequently reported circumstance was the contact with a confirmed or suspected case. More than half of the responders experienced at least one symptom potentially associated with COVID-19 since January 2020. The most frequently reported symptoms were headache (30.7%), rhinorrhea (22.5%) and dry cough (18.8%). Details of exposure and symptoms are given in Table 2. Twelve cases of COVID-19 were confirmed (1.8%, 95%CI: 3 and their symptoms are detailed in Appendix A, table S2.

Factors independently associated with an infection (confirmed or suspected) were age (odds ratio (OR) (95%CI): 0.96 (0.93-0.99), p-value=0.01), current smoking (OR (95%CI): 0.17 (0.03-0.59), p-value=0.02) and potential exposure to SARS-CoV-2 (OR (95%CI): 2.25 (1.15-4.44), p-value=0.02). Among the collected symptoms reported by patients with at least one of suspicious symptom of infection, anosmia and agueusia presented thhe highest positive predictive value for COVID-19 diagnosis (55% and 62% respectively, Appendix A, table S3).

Consequences on COVID-19 pandemic on rheumatic disease management

Overall, more than 30% of patients suspended or decreased the dosage of one of their drugs. Table 4. NSAIDs were the most frequently modified (41.6%), followed by bDMARDs (17.4%), cDMARDs (12.0%) and then corticosteroids (11.2%). Modifications were mostly due to fear of contagion (79.3%) or symptoms suggestive of infection (17.8%). Discontinuation for drug shortage or prescription problems was rare (2.9%). The only factor significantly associated with treatment modification for fear of contagion was NSAIDs use (Table 5). Almost two third of the patients who modified their treatment reported a flare of disease activity.

Details on modifications are given in

Other consequences of COVID-19 pandemic on rheumatic disease management were delayed or cancelled appointments, which occurred for 47% of patients (Table 4).

Discussion

To date, the impact on inflammatory rheumatic conditions and their associated treatment on incidence and severity of COVID-19 remain uncertain. In our survey performed in France during the lockdown period, frequency of COVID-19 was 1.8% (95%CI: 1.0-3.2%) if we considered only confirmed cases and 6.9% (95%CI: 5.1-9.2%) when high suspicious cases were added. This frequency is higher than that estimated in French general population (4.4%, range: 2.8-7.2) but similar to the estimated frequency of Paris region, the most affected region of the country (9.9%, range: 6.6 -15.7) where 87% of our respondents lived [START_REF] Salje | Estimating the burden of SARS-CoV-2 in France[END_REF]. These results are consistent with those reported by Favalli et al. in Lombardy with no increased incidence of COVID-19 in a cohort of rheumatic patients as compared to the general population [START_REF] Favalli | Incidence of COVID-19 in patients with rheumatic diseases treated with targeted immunosuppressive drugs: what can we learn from observational data? Arthritis & Rheumatology[END_REF]. In line with them, we also did not observe more severe infections than expected in the general population with no death observed, five hospitalisations, and only one admission in intensive care unit [START_REF] Salje | Estimating the burden of SARS-CoV-2 in France[END_REF].

Non-smoking was associated with a higher risk of infection. Although counter-intuitive, this observation is consistent with several studies reporting a lower proportion of current smokers among hospitalised COVID-19 patients than expected based on population smoking rates [START_REF] Farsalinos | COVID-19 and the nicotinic cholinergic system[END_REF][START_REF] Farsalinos | Systematic review of the prevalence of current smoking among hospitalized COVID-19 patients in China: could nicotine be a therapeutic option?[END_REF]. The fact that age was negatively associated with risk of infection was also unexpected. One possible explanation is the fact that younger people had more social contact than the elderly ones. Of interest, we did not find any association between anti-rheumatic treatments (NSAIDs, corticosteroids, cDMARD or bDMARD) and risk of infection which supports the recent recommendations established by scientific societies to maintain the ongoing treatment [START_REF] Landewé | EULAR provisional recommendations for the management of rheumatic and musculoskeletal diseases in the context of SARS-CoV-2[END_REF][START_REF] Mikuls | American College of Rheumatology Guidance for the Management of Rheumatic Disease in Adult Patients During the COVID-19 Pandemic: Version 1[END_REF][START_REF] Richez | Managing patients with rheumatic diseases during the COVID-19 pandemic: The French Society of Rheumatology answers to most frequently asked questions up to May 2020[END_REF].

Almost one third of the respondents reduced or discontinued their ongoing therapy with NSAIDs, corticosteroids, cDMARDs or bDMARDs. This observation is higher to frequencies previously reported [START_REF] Favalli | Incidence of COVID-19 in patients with rheumatic diseases treated with targeted immunosuppressive drugs: what can we learn from observational data? Arthritis & Rheumatology[END_REF][START_REF] Rosenbaum | Biologics, spondylitis and COVID-19[END_REF][START_REF] Fragoulis | Treatment adherence of patients with systemic rheumatic diseases in COVID-19 pandemic[END_REF]. Discontinuation or reduction of ongoing therapy lead to a subjective disease flare-up in two thirds of the patients. If some of these modifications were secondary to suspicious symptoms of possible SARS-Cov2 infection or to confirmed infection, the majority of them (79.3%) were linked to fear of contagion especially in patients treated with NSAIDs. This high rate of NSAIDs discontinuation might have been encouraged by a warning of the French health authorities published on 14 March and signalling serious side-effects in COVID-19 patients treated with NSAIDs [START_REF]Actualisation recommandations Covid 19[END_REF], in line with other regulatory agencies [START_REF] Day | Covid-19: European drugs agency to review safety of ibuprofen[END_REF]. However, there is no scientific data to support an increased risk of SARS-CoV-2 infection or COVID-19 severity with NSAIDs [START_REF] Moore | Does Ibuprofen Worsen COVID-19?[END_REF]. Factors independently associated with the non-response to the questionnaire were younger age, SpA diagnosis and recruitment through Spondy+® platform.

The cross-sectional survey design may be a limitation. Unfortunately we might have missed infected patients who were not able to respond to the survey because they were hospitalized or died. The high number of suspicion of COVID-19 without diagnosis confirmation is another limitation. However it reflected the situation in France during the lockdown period with a limited availability of PCR-based diagnostic tests.

In conclusion, this survey confirms that the incidence and severity of COVID-19 are not increased in patients with RA, SpA and PsA. It also adds evidence of the safety of antirheumatic drugs use (including NSAIDs, corticosteroids, cs-and b-DMARDs) during this period. This observation, together with the high rate of increased disease activity in case of treatment discontinuation, suggests that these drugs should be maintained.

1.0- 3 .

 3 2%) by nasopharyngeal polymerase chain reaction (PCR) testing, and/or by CT-scan (with typical COVID-19 abnormalities): 4 patients have both positive PCR test and CT-scan, 3 PCR test positive only and 5 positive CT-scan only. None of them died. Five patients required hospitalisation, including one admission in intensive care unit. Four required lowflow oxygen supplementation. In addition to these confirmed cases, 33 patients had a strong suspicion of COVID-19 (5.0%, 95% confidence interval (CI): 3.6-7.1%) but without PCR test confirmation (negative PCR test (n=7) or no PCR test performed (n=26). Characteristics of confirmed and suspected cases as compared to non-infected patients are summarized in Table

Figure 1 :

 1 Figure 1: Flow chart

Table 1 .

 1 Characteristics of the study population

	Characteristic	All (n =655)	Spondyloarthritis (n = 474)	Rheumatoid arthritis (n = 129)	Psoriatic arthritis (n=52)
	Sociodemographic characteristics,
	-Gender, n (%) of men	250 (38.2)	206 (43.5)	22 (17.1)	22 (42.3)
	-Age, mean ± sd	51.0 ± 13.4	48.1 ± 12.2	60.5 ± 12.8	54.1 ± 13.8
	-Health professionnals, n (%)	94 (14.3)	76 (16.0)	11 (8.5)	7 (13.5)
	-Profession with public contact, n (%)	271 (41.4)	208 (43.9)	40 (31.0)	23 (44.2
	Medical history
	-Rheumatic disease duration, mean ± sd	12.5 ± 10.2	13.0 ± 10.2	12.0 ± 10.6	8.4 ± 8.4
	-Smokers, n (%)	103 (15.7)	75 (15.8)	20 (15.5)	8 (15.4)
	-BMI > 25, n (%)	307 (46.9)	218 (46.0)	59 (45.7)	30 (52.7)
	-At least one comorbidity*, n (%)	233 (35.6)	150 (31.3)	65 (50.4)	18 (34.6)
	-Number of comorbidities*, mean ± sd	0.5 ± 0.8	0.4 ± 0.7	0.7 ± 1.0	0.5 ± 0.8
	Treatment
	-NSAIDs, n (%)	300 (45.8)	249 (52.5)	26 (20.2)	25 (48.1)
	-Corticosteroids, n (%)	107 (16.4)	28 (5.9)	74 (57.4)	5 (9.6)
	daily dose ≥ 10 mg/day	21 (3.2)	10 (2.1)	9 (7.0)	2 (3.8)
	-cDMARDs, n (%)	158 (24.1)	58 (12.2)	83 (64.3)	17 (32.7)
	Methotrexate, n (%)	132 (20.1)	42 (8.9)	75 (58.1)	15 (28.9)
	-bDMARDs, n (%)	404 (61.7)	299 (63.1)	74 (57.4)	31 (59.6)
	TNF blockers, n (%)	306 (46.7)	255 (53.8)	33 (25.6)	18 (34.6)
	IL-17 blockers, n (%)	55 (8.4)	43 (9.07)	0 (0)	12 (23.1)
	IL-6 blockers, n (%)	18 (2.7)	0 (0)	17 (13.2)	1 (1.92)
	tsDMARDs, n (%)	10 (1.5)	4 (0.8)	4 (3.1)	2 (3.8)
	sd: standard deviation; BMI: body mass index; NSAIDs: non-steroidal anti-inflammatory drugs; cDMARDs:
	conventional disease modifying anti-rheumatic drugs; bDMARDs : biological conventional disease modifying

anti-rheumatic drugs; tsDMARDs: targeted synthetic conventional disease modifying anti-rheumatic drugs. * among the following comorbidities (past or present): cardiovascular disease (stroke, heart failure), hypertension, diabetes, asthma, renal failure, respiratory insufficiency, cancer, hematologic malignancy, HIV, HBV or HCV infection, cirrhosis, sickle cell anemia, splenectomy, organ transplant.

Table 2 .

 2 SARS-CoV-2 exposure and symptoms

	Characteristic	All (n = 655)	Spondyloarthritis (n = 474)	Rheumatoid arthritis (n = 129)	Psoriatic arthritis (n = 52)
	Exposure, at least one, n (%)	211 (33.2)	170 (35.9)	30 (23.2)	11 (21.1)
	-Contact with a traveler in high incidence	83 (12.7)	70 (14.8)	9 (7.0)	4 (7.7)
	country*, n (%)
	-Contact with a confirmed case, n (%)	58 (8.9)	49 (10.4)	6 (4.7)	3 (5.8)
	-Contact with a suspected case, n (%)	153 (23.4)	125 (26.4)	20 (15.5)	8 (15.4)
	-Travel in a high incidence country*, n (%)	35 (5.3)	28 (5.9)	6 (4.7)	1 (1.9)
	Symptoms, at least one, n (%)	388 (59.2)	290 (61.2)	67 (51.9)	31 (59.6)
	-Headache, n (%)	201 (30.7)	157 (33.1)	25 (19.4)	19 (36.5)
	-Rhinorrhea, n (%)	147 (22.5)	106 (22.4)	25 (19.4)	16 (30.8)
	-Dry cough, n (%)	123 (18.8)	101 (21.3)	16 (12.4)	6 (11.5)
	-Fever, n (%)	117 (17.9)	89 (18.8)	17 (13.2)	11 (21.1)
	-Myalgia, n (%)	115 (17.6)	91 (19.3)	15 (11.6)	9 (17.3)
	-Diarrhea, n (%)	106 (16.2)	87 (18.3)	12 (9.3)	7 (

Table 3 .

 3 Factors associated with SARS-CoV-2 infection (confirmed or suspected) in the multivariate regression analysis

	Characteristic	No infection	Infection (n= 45)	Univariate analysis	Multivariate analysis
		(n = 610)	Suspected (n = 33)	Confirmed (n = 12)	OR (95%CI)	OR (95%CI)
	Gender
	-Female, n (%)	372 (91.6)	24 (6.2)	9 (2.2)	-
	-Male, n (%)	238 (95.2)	9 (3.6)	3 (1.2)	0.57 (0.28-1.09)
	Age, mean ± sd	51.5 ± 13.3	44.8 ± 13.0	46.4 ± 14.6	0.96 (0.94-0.99)**	0.97 (0.94-0.99)**
	Disease
	-Rheumatoid arthritis, n (%)	122 (94.6)	3 (2.3)	4 (3.1)	-
	-Psoriatic arthritis, n (%)	47 (90.4)	4 (7.7)	1 (1.9)	1.85 (0.53-6.10)
	-Spondyloarthritis, n (%)	441 (92.8)	26 (5.7)	7 (1.5)	1.30 (0.60-3.28)
	Health professionals
	-No, n (%)	529 (94.1)	23 (4.3)	9 (1.6)	-
	-Yes, n (%)	81 (86.2)	10 (10.6)	3 (3.2)	2.65 (1.30-5.16)**	1.71 (0.78-3.61)
	Profession with public contact
	-No, n (%)	367 (95.3)	12 (3.4)	5 (1.3)	-
	-Yes, n (%)	243 (89.7)	21 (7.7)	7 (2.6)	2.49 (1.35-4.73)**	1.73 (0.87-3.48)
	Smokers,
	-No, n (%)	509 (92.0)	31 (5.8)	12 (2.2)	-
	-Yes, n (%)	101 (98.1)	2 (1.9)	0 (0)	0.23 (0.04-0.78)*	0.18 (0.03-0.61)*
	BMI, mean ± sd	25.6 ± 5.3	25.0 ± 4.7	25.0 ± 5.0	0.98 (0.91-1.03)
	At least one comorbidity †, n (%)
	-No, n (%)	389 (92.2)	26 (6.2)	7 (1.6)
	-Yes, n (%)	221 (94.8)	7 (3.0)	5 (2.1)	0.64 (0.31-1.23)
	Number of comorbidities †, mean ± sd	0.5 ± 0.8	0.3 ± 0.7	0.6 ±0.8	0.81 (0.49-1.23)
	NSAIDs
	-No, n (%)	313 (92.9)	17 (5.0)	7 (2.1)	-
	-Yes, n (%)	297 (93.4)	16 (5.0)	5 (1.6)	0.92 (0.50-1.69)

Table 5 :

 5 factors associated with treatment modification for fear of contagion in the multivariate analysis NSAIDs: non-steroidal anti-inflammatory drugs; cDMARDs: conventional disease modifying anti-rheumatic drugs; bDMARDs : biological conventional disease modifying anti-rheumatic drugs; tsDMARDs: targeted synthetic conventional disease modifying anti-rheumatic drugs. † among the following comorbidities (past or present): cardiovascular disease (stroke, heart failure), hypertension, diabetes, asthma, renal failure, respiratory insufficiency, cancer, hematologic malignancy, HIV, HBV or HCV infection, cirrhosis, sickle cell anemia, splenectomy, organ transplant * p≤0.05; ** p≤0.01; *** p≤0.001

	Characteristic	No treatment modification	Treatment modification	Univariate analysis	Multivariate analysis
		(n= 431)	(n=224)	OR (95%CI)	OR (95%CI)
	Gender
	-Female	305 (75.3)	100 (24.7)	-
	-Male	185 (74.0)	65 (26.0)	1.07 (0.74-1.54)
	Age, mean ± SD	52.1 (13.5)	47.8 (12.5)	0.98 (0.96-0.99)***	0.99 (0.98-1.01)
	Disease
	-Rheumatoid arthritis	111 (86.0)	18 (14.0)	-	-
	-Psoriatic arthritis	39 (75.0)	13 (25.0)	2.06 (0.91-4.57)	1.26 (0.52-2.98)
	-Spondyloarthritis	340 (71.7)	134 (28.3)	2.43 (1.45-4.28)***	1.19 (0.63-2.33)
	Health professionals
	-No, n (%)	417 (74.3)	144 (25.7)	-
	-Yes, n (%)	73 (77.7)	21 (22.3)	0.83 (0.48-1.38)
	Profession with public contact
	-No, n (%)	296 (77.1)	88 (22.9)	-
	-Yes, n (%)	194 (71.6)	77 (28.4)	1.34 (0.93-1.90)
	Smokers,
	-No, n (%)	270 (76.1)	85 (23.9)	-
	-Yes, n (%)	220 (73.3)	80 (26.7)	1.16 (0.81-1.65)
	BMI, mean ± SD	25.8 (5.6)	24.8 (4.2)	0.96 (0.92-0.99)*	0.98 (0.93-1.01)
	At least one comorbidity †, n (%)
	-No, n (%)	301 (71.3)	121 (28.7)	-	-
	-Yes, n (%)	189 (81.1)	44 (18.9)	0.58 (0.39-0.85)**	0.82 (0.52-1.26)
	NSAIDs

sd: standard deviation; BMI: body mass index;

Acknowledgements: we thank Dr. Benjamin Hagège, Dr. Ilaria Padovano, Dr. Andreea Todorutiu for their contribution on the survey and selection of the patients. We also thank Frédéric Durand-Salmon for giving us the possibility to use Spondy+, a patient support program developed by BEPATIENT.

Funding: This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Conflict of interest:

The authors declare no conflicts of interest.

Appendix A. Supplementary data

Supplementary data (Tables S1-S3, Figure S1) associated with this article can be found in the online version at …


Rakhi Sood

Stefano Giancola

Anna Donnadio

Marta Zatoń

Nicolas Donzel

Jacques Rozière

Deborah J Jones

Sara Cavaliere
email: sara.cavaliere@umontpellier.fr

Active Electrospun Nanofibers as an Effective Reinforcement for Highly Conducting and Durable Proton Exchange Membranes

Keywords: Proton exchange membrane fuel cell, reinforced membrane, active fibrous reinforcement, electrospinning, triazole functionalization

Mechanical reinforcement of proton exchange membranes is a great challenge allowing the reduction of their thickness, with the advantages of lower resistance, improved water transport, decreased fuel crossover and high durability, which are crucial for fuel cells. We describe herein a new class of reinforced membranes based on nanofibers of polysulfone (PSU) functionalized with 4-heptyl-1,2,3-triazole (PSUT) likely interacting via hydrogen bonds or ionic cross-linking with a short-side-chain type perfluorosulfonic acid (Aquivion ® ) matrix. PSUT with two functionalization degrees was electrospun into webs which were impregnated with Aquivion ® to afford composite membranes that presented higher dimensional stability in water, increased mechanical strength and Young modulus in comparison to reinforced membranes of non-functionalized PSU and pristine ionomer membrane, without any decrease in proton conductivity. Membrane-electrode assemblies incorporating an Aquivion ® membrane reinforced with PSUT exhibited 5 times superior durability than those with a pristine Aquivion ® membrane without any radical scavenger. These features may be ascribed to the specific interaction between basic fibers and acidic

Introduction

The development of membranes with low protonic resistance, gas permeability and cost, and high durability is crucial for the widespread adoption of proton exchange membrane fuel cells (PEMFC) for clean energy conversion [START_REF] Kusoglu | New Insights into Perfluorinated Sulfonic-Acid Ionomers[END_REF][START_REF] Zatoń | Design of Heterogeneities and Interfaces with Nanofibers in Fuel Cell Membranes BT -Handbook of Nanofibers[END_REF]. In order to optimize the performance of PEMFCs, reduction in the resistance of the polymer electrolyte membrane (PEM) via reduction of its thickness is widely suggested [START_REF] Subianto | Physical and chemical modification routes leading to improved mechanical properties of perfluorosulfonic acid membranes for PEM fuel cells[END_REF]. However, this approach brings up degradation issues as thin membranes are weaker and more gas permeable and generally exhibit earlier mechanical failure than thicker membranes [START_REF] Qiu | Mechanical failure and mitigation strategies for the membrane in a proton exchange membrane fuel cell[END_REF]. Several physical and chemical approaches have been investigated to increase membrane durability [START_REF] Subianto | Physical and chemical modification routes leading to improved mechanical properties of perfluorosulfonic acid membranes for PEM fuel cells[END_REF], including the use of polymer reinforcements [START_REF] Bahar | Ultra-thin integral composite membrane[END_REF][START_REF] Xiao | Physically stable and high performance Aquivion/ePTFE composite membrane for high temperature fuel cell application[END_REF][START_REF] Tang | Mechanical properties of a reinforced composite polymer electrolyte membrane and its simulated performance in PEM fuel cells[END_REF][START_REF] Shoibal | Advances in Proton Exchange Membrane Technology[END_REF], inorganic scaffolds [START_REF] Casciola | Zirconium phosphate reinforced short side chain perflurosulfonic acid membranes for medium temperature proton exchange membrane fuel cell application[END_REF][START_REF] Donnadio | Promising Aquivion Composite Membranes based on Fluoroalkyl Zirconium Phosphate for Fuel Cell Applications[END_REF][START_REF] Donnadio | Double filler reinforced ionomers: a new approach to the design of composite membranes for fuel cell applications[END_REF], thermal annealing [START_REF] Alberti | Annealing of Nafion 1100 in the Presence of an Annealing Agent: A Powerful Method for Increasing Ionomer Working Temperature in PEMFCs[END_REF][START_REF] Giancola | Study of Annealed Aquivion® Ionomers with the INCA Method[END_REF] and chemical cross-linking [START_REF] Zatoń | Design of Heterogeneities and Interfaces with Nanofibers in Fuel Cell Membranes BT -Handbook of Nanofibers[END_REF]. For instance, expanded polytetrafluoroethylene (ePTFE) impregnated with Nafion ® or other perfluorosulfonic acids (PFSA) is currently employed in commercial PEMFC membranes for automotive applications (e.g. Gore-Select ® membrane) with high durability and mechanical properties. [START_REF] Bahar | Ultra-thin integral composite membrane[END_REF][START_REF] Tang | Mechanical properties of a reinforced composite polymer electrolyte membrane and its simulated performance in PEM fuel cells[END_REF][START_REF] Shoibal | Advances in Proton Exchange Membrane Technology[END_REF]. However, PTFE suffers from mechanical softening at fuel cell operation temperatures, and its good solvent properties for oxygen lead to high gas crossover affecting the membrane stability.

Another strategy to mechanically reinforce ionomer membranes is the incorporation of a macroporous non-woven nanofiber web into the polymer matrix, allowing for the preparation of composite membranes with controlled 3D design and with significantly improved mechanical properties. In this regard, electrospinning has emerged as a versatile technique for the synthesis of robust and highly porous fibrous webs with targeted morphology, controlled fiber diameters and possibility of upscale [START_REF] Cavaliere | Electrospun Materials for Proton Exchange Membrane Fuel Cells and Water Electrolysis[END_REF]. Proton exchange membranes reinforced with electrospun nanofibers are possible candidates to disrupt the usual conductivity/durability trade-off and exceed the performance of state-of-the-art membranes [START_REF] Zatoń | Design of Heterogeneities and Interfaces with Nanofibers in Fuel Cell Membranes BT -Handbook of Nanofibers[END_REF][START_REF] Subianto | Electrospun Nanofibers for Low-Temperature Proton Exchange Membrane Fuel Cells[END_REF][START_REF] Sood | Electrospun nanofibre composite polymer electrolyte fuel cell and electrolysis membranes[END_REF][START_REF] Boaretti | Mechanically stable nanofibrous sPEEK/Aquivion® composite membranes for fuel cell applications[END_REF]. Pintauro and coworkers pioneered this approach [START_REF] Ballengee | Composite Fuel Cell Membranes from Dual-Nanofiber Electrospun Mats[END_REF][START_REF] Choi | Nanofiber network ionexchange membranes[END_REF][START_REF] Powers | Electrospun tri-layer membranes for H2/Air fuel cells[END_REF]. For instance, nanostructured membranes based on electrospun nanofibers of a robust polymer i.e. polysulfone (PSU) embedded into PFSA were prepared for fuel cells and water electrolyzers, where rigid nonwoven provide mechanical strength and low hydrogen crossover, and PFSA provide high proton conductivity [START_REF] Ballengee | Composite Fuel Cell Membranes from Dual-Nanofiber Electrospun Mats[END_REF][START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF].

Inducing specific ionic interactions at the fiber/matrix interface by combining a polymer bearing basic sites with an ionomer bearing sulfonic/phosphonic acid functionalities substantially increases the impact of the nanofiber web on membrane properties [START_REF] Wang | Enhanced proton conductivities of nanofibrous composite membranes enabled by acid-base pairs under hydrated and anhydrous conditions[END_REF][START_REF] He | Synergistic proton transfer through nanofibrous composite membranes by suitably combining proton carriers from the nanofiber mat and pore-filling matrix[END_REF][START_REF] Wang | Porous Nafion nanofiber composite membrane with vertical pathways for efficient through-plane proton conduction[END_REF][START_REF] Jones | Reinforced ionomer membranes, fabrication and application in energy conversion devices[END_REF][START_REF] Wang | Constructing Amino-Functionalized Flower-Like MOFs Nanofibers in Sulfonated Poly(ether sulfone) Proton Exchange Membrane for Simultaneously Enhancing Interface Compatibility and Proton Conduction[END_REF].

Wang et al. [START_REF] Wang | Enhanced proton conductivities of nanofibrous composite membranes enabled by acid-base pairs under hydrated and anhydrous conditions[END_REF] reported improved stiffness and dimensional stability for membranes based on sulfonated poly(ether ether ketone) (sPEEK) nanofibers embedded into chitosan matrix attributed to acid-base interactions. Higher proton conductivity was also obtained for the composite PEM with respect to cast sPEEK and chitosan alone. This behavior was ascribed to the resulting ion pairs on the fiber/ionomer interface generating structurally ordered protonhopping sites. He et al. [START_REF] He | Synergistic proton transfer through nanofibrous composite membranes by suitably combining proton carriers from the nanofiber mat and pore-filling matrix[END_REF] investigated the effect of different interfacial interactions (acidacid, acid-base, acid-neutral, base-base, base-neutral) on membranes based on poly(vinyl alcohol) (PVA) nanofibers either non-modified or functionalized with -NH2 or -SO3H embedded into basic chitosan and acidic sPEEK. The acid-base interaction was demonstrated as the most effective in enhancing membrane mechanical properties and conductivity [START_REF] He | Synergistic proton transfer through nanofibrous composite membranes by suitably combining proton carriers from the nanofiber mat and pore-filling matrix[END_REF].

Furthermore, membranes comprising electrospun polybenzimidazole nanofiber webs impregnated with Aquivion ® thereby coupling acid-base ionic interactions along with the scaffolding effect of nanofibers demonstrated exceptional mechanical stability, low gas crossover and high durability upon long-term fuel cell operation [START_REF] Jones | Reinforced ionomer membranes, fabrication and application in energy conversion devices[END_REF].

Since the development of copper-catalyzed azide-alkyne cycloaddition (CuAAC) modular ligation [START_REF] Rostovtsev | A Stepwise Huisgen Cycloaddition Process: Copper(I)-Catalyzed Regioselective "Ligation" of Azides and Terminal Alkynes[END_REF][START_REF] Tornøe | Peptidotriazoles on Solid Phase: [1,2,3]-Triazoles by Regiospecific Copper(I)-Catalyzed 1,3-Dipolar Cycloadditions of Terminal Alkynes to Azides[END_REF], a large variety of materials containing 1,2,3-triazole groups with advanced macromolecular design (e.g. telechelic polymers, block copolymers, dendrimers etc.) have been reported [START_REF] Lutz | 1,3-Dipolar Cycloadditions of Azides and Alkynes: A Universal Ligation Tool in Polymer and Materials Science[END_REF][START_REF] Mudraboyina | 1,2,3-Triazolium-based poly(ionic liquid)s with enhanced ion conducting properties obtained through a click chemistry polyaddition strategy[END_REF][START_REF] Sood | 1,2,3-Triazolium-based poly(acrylate ionic liquid)s[END_REF][START_REF] Sood | Triethylene glycol-based poly(1,2,3-triazolium acrylate)s with enhanced ionic conductivity[END_REF][START_REF] Akbey | 1H Solid-State NMR Investigation of Structure and Dynamics of Anhydrous Proton Conducting Triazole-Functionalized Siloxane Polymers[END_REF][START_REF] Johnson | Construction of Linear Polymers, Dendrimers, Networks, and Other Polymeric Architectures by Copper-Catalyzed Azide-Alkyne Cyclo-addition "Click" Chemistry[END_REF]. However, only a few studies have investigated acid-base blends based on triazole functionalized polymers [START_REF] Ahn | Facile preparation of blend proton exchange membranes with highly sulfonated poly(arylene ether) and poly(arylene ether sulfone) bearing dense triazoles[END_REF][START_REF] Sen | Nafion/poly(1-vinyl-1,2,4-triazole) blends as proton conducting membranes for polymer electrolyte membrane fuel cells[END_REF][START_REF] Yue | Improved performance of acidbase composite of phosphonic acid functionalized polysulfone and triazolyl functionalized polysulfone for PEM fuel cells[END_REF][START_REF] Li | Acid-base blend membranes consisting of sulfonated poly(ether ether ketone) and 5-amino-benzotriazole tethered polysulfone for DMFC[END_REF]. Acid-base blend membranes based on crosslinked sulfonated poly(arylene ether) and triazole-containing poly(arylene ether sulfone) demonstrated high proton conductivity, mechanical strength and dimensional stability ascribed to the acid-base interaction. In another approach, polysulfone tethered with aminobenzotriazole side-groups was combined with sulfonated poly (ether ether ketone) (sPEEK) to afford an acid-base polymer composite membrane [START_REF] Li | Acid-base blend membranes consisting of sulfonated poly(ether ether ketone) and 5-amino-benzotriazole tethered polysulfone for DMFC[END_REF], leading to enhanced performance in fuel cell compared with membrane electrode assemblies (MEAs) based on Nafion ® and sPEEK.

We earlier reported the synthesis of 1,2,3-triazole-functionalized polysulfone with different aliphatic and aromatic triazole ring substituents via microwave assisted-CuAAC [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF]. In this work we describe an active reinforcement based on 1,2,3-triazole functionalized nanofibers embedded in PFSA Aquivion ® . High chemical stability, antioxidant activity, aromatic character and ability to form hydrogen bonds [START_REF] Thibault | A versatile new monomer family: Functionalized 4-vinyl-1,2,3-triazoles via click chemistry[END_REF][START_REF] Iha | Applications of Orthogonal "Click" Chemistries in the Synthesis of Functional Soft Materials[END_REF][START_REF] Dubey | A click chemistry strategy to synthesize geraniol-coupled 1,4-disubstituted 1,2,3-triazoles and exploration of their microbicidal and antioxidant potential with molecular docking profile[END_REF] are relevant characteristics of the 1,2,3triazole group for fuel cell membrane durability, that potentially impact the overall properties of the corresponding composite membranes. Moreover, this flexible approach demonstrates the possibility of modulating membrane properties by adjusting the number and type of triazole moieties [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF]. We investigate the effectiveness of the triazole functionality in strengthening the impact of the polysulfone electrospun non-wovens on the PFSA matrix.

Polysulfone modified with different degrees of 4-heptyl-1,2,3-triazole was synthesized by microwave assisted-CuAAC and electrospun. The resulting fibrous webs were impregnated with Aquivion ® with EW of 830 g eq -1 (Aq830) to prepare nanocomposite acid-base PEMs.

The reinforced membranes were characterized for their proton conductivity, mechanical properties and dimensional stability under humidified conditions, and they were integrated into MEAs which were characterized in single fuel cell for their performance and durability.

Experimental Section

Materials

Chlorotrimethylsilane (TMCS, C3H9ClSi, ≥ 97.0 %), p-formaldehyde powder (HO(CH2O)nH, 95 %), tin tetrachloride (SnCl4, 98 %), sodium azide (NaN3, ≥ 99%), diisopropylethylamine (DIPEA, [(CH3)2CH]2NC2H5, 99 %), 1-nonyne (CH3(CH2)6C≡CH, 99%), copper(I) iodide triethylphosphite (CuI•P(OEt)3, 97 %), N,N,N′,N′′,N′′pentamethyldiethylenetriamine (PMDETA, [(CH3)2NCH2CH2]2NCH3, 99 %) and all solvents were purchased from Aldrich and used as received. A 24 wt% Aquivion ® dispersion in water (EW=830 g eq -1 ) was purchased from Solvay Specialty Polymers. Bisphenol A polysulfone (PSU) granules (Mn=80 KDa and Mw=102 KDa) were purchased from Goodfellow.

Synthesis of polysulfone functionalized with 4-heptyl-1,2,3-triazole (PSUT)

Polysulfone (PSU) functionalized with 4-heptyl-1,2,3-triazole (Figure S1 of Supporting Information, SI) was synthesized with a procedure based on chloromethylation and azidation as earlier reported [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF].

The chloromethyl-functionalized polysulfone (PSUCl) with a degree of functionalization of 30 mol% (PSUCl(0.3), 0.89 g) and 90 mol% (PSUCl(0.9), 0.95 g) functional groups per repeat unit presented the following 1 H NMR (400 MHz, DMF; ppm) chemical shifts: δ 8.05-6.90 (aromatic backbone), 4.75 (brs, 2H, CH2Cl), 1.70 (brs, 6H, C-CH3). 13 For the triazole functionalization, a solution of PSUAz(0.3) (100 mg, 0.062 mmol of azide functionalities), with 1-nonyne (11.5 mg, 0.093 mmol), CuI•P(OEt)3 (2.2 mg, 0.0062 mmol) and DIPEA (8 mg, 0.062 mmol) in DMF (5 mL) reacted under microwaves (Sairem MiniFlow 220SS) (20 W, 120 °C, 30 minutes under constant stirring). The 4-epthyl-1,2,3triazole-functionalized polysulfone PSUT(30 mol%) (93 mg) was recovered as previously described [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF]. An identical procedure was used combining PSUAz(0.9) (100 mg, 0.

Preparation of polymer electrospun nanofibers

Nanofibers of 4-heptyl-1,2,3-triazole functionalized polysulfone (PSUT) were prepared by electrospinning polymer solutions with a standard syringe using a grounded rotating drum collector configuration (Linari Engineering) under ambient conditions. Fibers were collected onto a rotating drum with a rotation speed of 800 rpm and a translational speed of 10 mm s -1 .

PSUT(30 mol%) and PSUT(90 mol%) nanofibers were obtained by electrospinning respectively a 13 wt% and 14 wt% polymer solution in anhydrous DMAc with an applied potential of 10 kV, a needle collector distance of 12 cm and a flow rate of 0.2 mL min -1 .

Non-modified polysulfone nanofibers were prepared by electrospinning a 18 wt% PSU solution in anhydrous dimethylacetamide (DMAc) and acetone in a 7:3 volume ratio as reported elsewhere [START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF].

Membrane preparation 2.4.1. Composite nanofiber membranes

Reinforced membranes embedding 12 wt% of fibers were prepared by impregnating electrospun webs with a 12 wt% dispersion of Aquivion ® (equivalent weight, EW = 830 g eq - 1 ) in a mixture of water/propanol/DMAc (weight ratio: 38 %/47 %/3 %). A layer of Aquivion ® dispersion was cast with a doctor blade. The electrospun web was then placed on the top of this layer and another layer of ionomer dispersion was cast onto it. Afterwards, the composite membrane was left to dry overnight at room temperature and then at 80 °C for 5h

to remove remaining solvent. Membranes were hot pressed at 160 °C at 980.6 kPa for 15 min and then thermally treated at 160 °C for 45 min. After, these were first washed in 1 M H2SO4

for 1 h at 80 °C for the reactivation of sulfonic acid sites, then 4-5 times in Milli-Q water at room temperature until neutral pH was achieved, and finally in Milli-Q water at 80 °C for one hour. Membrane thicknesses of 50 and 30 µm were set respectively for ex situ and fuel cell characterization.

Non-reinforced Aquivion ® membranes

A 16 wt% dispersion of Aquivion ® (EW = 830 g eq -1 ) in a mixture of water /propanol/DMAc (weight ratio: 50 %/24 %/10 %) was cast to obtain membranes of thickness similar to that of reinforced films. The membranes were left overnight at room temperature and then dried at 80 °C for 12h. Non-reinforced membranes were hot pressed, thermally treated and washed in the same way as for reinforced films.

Characterization of functionalized polymers

NMR spectra were recorded on a Bruker Avance HD 400 MHz spectrometer equipped with a BBFO (Broad Band Fluorine Observation) probe in DMF-d6 using tetramethylsilane (TMS) as reference residual hydrogenated solvent.

Characterization of nanofiber webs and membranes

Reinforced membrane fiber content

The fiber content of the reinforced membranes was evaluated by the weight ratio between the web and the membrane after drying at 80 °C for 5h.

Fiber and membrane morphology

Morphology of electrospun nanofibers and membranes was analyzed by field emissionscanning electron microscopy (FE-SEM) using a Hitachi S-4800 microscope after coating with a Pt layer of few nanometers (≈ 2-3 nm) to provide electrical conductivity. Freezefractured membrane cross-sections were prepared by immersing and cutting samples in liquid nitrogen. Fiber diameters were calculated using the ImageJ software. For each web, the dimension of more than 100 nanofibers was evaluated. A Gaussian function was used to fit the fiber diameter distribution of both PSUT(30 mol%) and PSUT(90 mol%) and to evaluate the average fiber diameter. TEM-EDX analysis of reinforced membrane cross-sections was carried out using a JEOL 2200FS (Source: FEG) microscope operating at 200 kV equipped with a CCD camera Gatan USC (16 MP). Quantitative EDX analysis was performed by evaluating the mass percentage of fluorine at different points between a region comprising only Aquivion ® to a region with only nanofibers.

Membrane conductivity

Proton conductivity of the membranes was measured in-plane at 80 °C and 110 °C as a function of RH on 5 cm x 0.5 cm membrane strips in the frequency range 10 Hz-100 kHz with 100 mV signal amplitude by four-probe impedance measurements by using an Autolab, PGSTAT30 potentiostat/galvanostat equipped with a frequency response analysis module.

The RH was controlled by using stainless steel sealed-off cells that comprise two communicating cylindrical compartments held at different temperatures, the cold compartment containing water, and the hot compartment housing the membrane under test.

RH values were calculated from the ratio between the pressures of saturated water vapor (p) at the temperatures of the cold (Tc) and hot (Th) compartment:

% = p Tc ℎ * 100

Stress-strain tests

Stress-strain mechanical tests were performed by using a Zwick Roell Z1.0 instrument with a 200 N static load cell equipped with a climatic chamber that operated in the RH range of 30-95 % (± 0.5 %) and in the temperature range of 10-80 °C (± 0.5 °C). Measurements were performed on 100 mm x 5 mm film strips. Before the measurements at room temperature, samples were equilibrated for five days in vacuum desiccators at 53 % RH and RT (25 °C).

The investigations at higher temperature and humidity were performed after sample equilibration in the climate chamber at 70 °C and 80 % RH for one day. An initial grip separation of 10.000 ± 0.002 mm and a crosshead speed of 30 mm min -1 were used. At least five replicate film strips were analyzed. The data were analyzed by the TestXpert V11.0

Master software and are presented as mean ± standard deviation (SD).

Membrane dimensional swelling in water

Gravimetric water uptake (WU, %) was determined by weighing the membranes after immersion in deionized water at 80 °C for 15 h (Wf) and then after drying them at 120 °C for 5 h (Wi). Before weighing, excess water was removed from the surface of hydrated samples by lightly pressing the membranes between two filter papers. Membrane water uptake was calculated using the following relationship:

WU % = Wf -Wi Wi * 100
Dimensional swelling in water was determined by measuring the dimensions of rectangular samples of membrane after immersion in liquid water at 80 °C for 15 h as well as after drying at 120 °C for 5 h. The dimensional change was calculated using the following equation:

% = Dw -Dd Dd * 100
where Dw and Dd are the dimensions (volume, area or thickness) of respectively the wet and dry sample.

Membrane ion exchange capacity

The ion exchange capacity (IEC) of the membranes was determined by acid-base titrations by using a Radiometer automatic titrimeter (TIM900 TitraLab and ABU91 Burette)

operating with the equilibrium point method. Before titration, the membranes were heated at 120 °C for 5 h to remove water and determine the weight of the dried sample. The dry membranes were equilibrated overnight at RT with a 0.1 M NaCl aqueous solution to exchange Na + ions with the membrane protons. The solutions were then titrated with 0.1 M NaOH without removing the membranes [START_REF] Casciola | Zirconium phosphate reinforced short side chain perflurosulfonic acid membranes for medium temperature proton exchange membrane fuel cell application[END_REF].

Membrane permeability

Permeation measurements were performed using a fuel cell test station with the membranes under evaluation held in a single fuel cell fixture (active surface area of 8 cm 2 )

equipped with a micro GC, gas flow mixer and pressure monitor. The membranes were placed in the single cell hardware, gas sealed at 1 bar and conditioned to ensure equilibrium state of the ionomer. The hydrogen permeability was determined at 80 % relative humidity at 80 °C.

2.6.8. Fuel cell polarization curve measurements and durability tests I-V curves were recorded on membrane electrode assemblies (MEAs) (8 cm 2 ) prepared by incorporating the proton exchange membrane between two electrodes (Sigracet 10BC from Baltic Fuel Cells, nominal platinum loading of 0.5 mgPt cm -2 ) and two propylene sub-gaskets.

MEAs were hot-pressed at 160 °C and 6.3 MPa for 3 minutes. Prior to measurements, MEAs were conditioned in the fuel cell at 80 °C, 100 % RH and 2.5 bar absolute pressure for at least 12 hours, at a current density of 400 mA cm -2 until constant potential was obtained. I-V curves were recorded on a Bio-Logic VMP3B-80 A booster. The fuel cell was fed with H2

and air using a constant flux of 150 and 500 mil min -1 , respectively. Accelerated stress test (AST) were performed on MEAs based on reinforced and non-reinforced Aquivion ® membranes using standard 25 cm 2 single cell hardware in FuelCon fuel cell station. Baltic Fuel Cells electrodes with a nominal loading of 0.2 mgPt cm -2 (anode) and 0.5 mgPt cm -2 (cathode) were used in this study. Break-in procedure consisted of cycling operation at 0.4 V and 0.8 V for 5 min for 4 hours at 80 °C and 100 % RH in H2/air. After break-in, MEAs were exposed to wet/dry cycling between 0 and 100 % RH in 10-minute cycles. Durability testing was performed at OCV at 90 °C in H2/air with gas flow rates of 420 and 1450 sccm respectively.

Results and Discussion

Electrospun polymer nanofibers

Electrospinning of 4-heptyl-1,2,3-triazole-functionalized polysulfone afforded regular and homogenous nanofibers with thin diameter and small size distribution. Thin nanofibers are indeed advantageous due to their high surface area ensuring enhanced polymer/ionomer interface. For this study, triazole functionalized polysulfone with a degree of functionalization of 30 mol% (PSUT(30 mol%)) and 90 mol% (PSUT(90 mol%)) functional group per repeat unit were taken into consideration which allowed the effect of the amount of interfacial triazole moieties on membrane properties to be investigated. In Figure 1 are depicted the SEM micrographs of fiber webs of non-functionalized (PSU) and functionalized polysulfone prepared with the optimized electrospinning conditions, as well as the histograms of the related fiber diameter distribution. Randomly oriented and non-welded nanofibers were obtained in all cases. This web morphology was targeted since it ensures a strengthening of the membrane throughout its thickness. In particular, the absence of fiber welding allows the web to expand during its impregnation with the PFSA dispersion leading to a homogeneous membrane architecture [START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF]. Diameters of PSUT(30 mol%) (Figure 1 e) and PSUT(90 mol%) (Figure 1 f) fibers range from 100 to 700 nm with an average fiber diameter (AFD) of 313 ± 6 nm and 256 nm ± 8 nm respectively. Thus, the degree of functionalization seems to have a negligible effect on the nanofiber morphology. Nonetheless, a higher proportion of thin nanofibers (diameter ca. 100 nm) was obtained for the triazole functionalized polysulfone with the higher degree of substitution. In comparison, PSU fiber diameters range from 200 to 900 with AFD of 390 nm (Figure 1 d). The difference in diameter can be ascribed to the different electrospinning conditions (18 wt% solution of the non-functionalized polymer in a in DMAc/acetone (7v/3v) binary solvent system [START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF], while PSUT was electrospun at lower concentration (13-14 wt%) and in pure DMAc).

Membrane morphology

In the subsequent step, the electrospun webs of functionalized and pristine polysulfone were used to prepare reinforced membranes by impregnation with Aquivion ® (EW = 830 g eq -1 ). In this study, fiber content was fixed at 12 wt%. Membranes reinforced by triazole functionalized polysulfone are referred to as Aq-PSUT(n) where n is the degree of functionalization of PSUT. For comparison, non-reinforced Aquivion ® and Aquivion-PSU nanofiber reinforced membranes are labelled as Aq and Aq-PSU respectively. In Figure 2 are represented the SEM images of cross-sections of membranes reinforced with functionalized and non-functionalized PSU fiber webs.

In all cases, fully dense and crack-free films were obtained, indicating that PFSA impregnation was successful. Moreover, both the ionomer and the nanofibers are homogenously distributed throughout membrane cross-section.

In the present study, a clear difference emerges between membranes reinforced with the functionalized and the non-functionalized polysulfone fibers that may be seen in the higher magnification micrographs of Figure 2. For the non-functionalized PSU/Aquivion ® membrane a void space is observed between the fibers and the surrounding ionomer (Figure 2 d). This might be a result of the freeze-fracture process during preparation of samples for SEM or of the drying of the membrane under vacuum conditions of the analysis. These results give a preliminary corroboration of the presence of acid-base ionic crosslinking between 1,2,3-triazole moieties on the functionalized polymer fibers and sulfonic acid sites of the Aquivion ® matrix, unlike for polysulfone fibers.

This could be promoted by PFSA/PSUT co-penetration during the membrane hot-pressing procedure at temperature higher than the glass transition temperature (Tg) [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF] and the α transition temperature of Aquivion ® (≈ 125 °C) [START_REF] Kusoglu | New Insights into Perfluorinated Sulfonic-Acid Ionomers[END_REF]. Quantitative TEM-EDX analysis shed further information on the PFSA/PSUT blend region at the fiber/ionomer interface (Figure 4).

Using fluorine as a marker to evaluate the penetration of Aquivion ® into the fiber, it is observed that for Aq-PSU the fluorine signal sharply decreases to almost 0 mass% at the fiber edge (Fig. 4 The proton conductivity of all the reinforced and non-reinforced membranes increases with temperature and relative humidity as expected for PFSA [START_REF] Donnadio | Double filler reinforced ionomers: a new approach to the design of composite membranes for fuel cell applications[END_REF][START_REF] Subianto | Reactive coaxial electrospinning of ZrP/ZrO2 nanofibres[END_REF][START_REF] Jones | Membrane materials and technology for low temperature fuel cells[END_REF]. The conductivity of nonreinforced Aquivion ® extends from 60 mS cm -1 (50 % RH) to 180 mS cm -1 (95 % RH) at 80 °C and reached 260 mS cm -1 at 95 % RH and 110 °C. The presence of PSU nanofibers leads to a slight decrease of the conductivity of the pure ionomer in agreement with the reduction of the concentration of proton carriers due to the presence of the non-conducting reinforcement [START_REF] Ballengee | Composite Fuel Cell Membranes from Dual-Nanofiber Electrospun Mats[END_REF][START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF]. Some loss in conductivity could be expected upon the introduction of triazole functionalized polysulfone nanofiber reinforcement, due to possible proton transfer from sulfonic acid groups in their interaction with basic groups of the fiber surface. No such conductivity reduction was observed however, and indeed there is negligible effect of the degree of functionalization. This implies that protons are not immobilized and still available for transport.

Experimental ion exchange capacity (IEC) of composite membranes reinforced with functionalized polysulfone were determined and compared to those of Aq-PSU and Aq830

(Table 1) The IEC of reinforced membranes is slightly lower than that of pristine Aquivion ® (ca 15 % lower IEC for 12 wt% of inert polymer fiber content). Independently of their degree of functionalization, Aq-PSUT reinforced membranes have the same IEC as Aq-PSU membranes, around 1 mequiv•g -1 . These results are in agreement with proton conductivity determined from in-plane measurements for the different membranes. On the contrary, in acid-base blend of triazole-functionalized polymer and sulfonated ionomer IEC values are lower than the theoretical values calculated assuming that only the ionomer provides dissociable protons [START_REF] Li | Acid-base blend membranes consisting of sulfonated poly(ether ether ketone) and 5-amino-benzotriazole tethered polysulfone for DMFC[END_REF]. The IEC values obtained for the nanocomposite membranes could be explained considering that the interaction does not involve all triazole moieties, but only those at the extended Aquivion ® /PFSA interface. This assumption is supported by the fact that the degree of functionalisation of the PSU (30-90 mol% triazole) does not seem to play a significant role in the properties of the corresponding fiber reinforced membranes.

Mechanical properties

The mechanical properties of reinforced functionalized and non-functionalized polysulfone membranes were determined at 20 °C and 45 % RH and at 70 o C and 80 % RH. The obtained results are summarized in Table 2 and stress-strain curves are presented in Figure S2 (Supporting Information).

Table 2. Mechanical properties of reinforced and non-reinforced membranes at 20 °C and 45

% RH and at 70 °C and 80 % RH.

Membrane

20°C- 45%RH 70°C- 80%RH 20°C- 45%RH 70°C- 80%RH 20°C- 45%RH 70°C- 80%RH 20°C- 45%RH 70°C- 80RH% 20°C- 45%RH 70°C- 80%RH Aq 196±7 107±9 10±2 5±1 11±1 6±1 54±2 81±3 500±9 439±18 Aq-PSU 349±4 203±10 / / 15±1 9±1 9.9±1.3 11.0±2.0 125±7 51±3 Aq-PSUT(30 mol%) 390±10 259±5 18±1 11±2 20±1 13±1 135±19 164±11 2853±25 1891±11 Aq-PSUT(90 mol%) 385±14 220±18 17±1 9±1 18±1 10±1 59±19 89±30 1274±12 643±8
All reinforced membranes presented a significantly higher Young modulus (E) than reference Aquivion ® due to the presence of the fiber reinforcement. Elastic modulus followed the order Aq-PSUT(30 mol%)>Aq-PSUT(90 mol%)>Aq-PSU>Aq and decreased with the increase in temperature-relative humidity as expected for PFSA based membranes [START_REF] Kusoglu | New Insights into Perfluorinated Sulfonic-Acid Ionomers[END_REF]. At 70 °C and 80 % RH the membrane reinforced with PSUT(30 mol%) nanofibers displayed a Young modulus of 259 N•mm -2 , the membrane reinforced with PSU, 203 N•mm -2 , and the non-reinforced Aquivion ® , 107 N•mm -2 . The higher stiffness of triazole functionalized PSU reinforced membranes with respect to Aq-PSU can be explained by ionic interactions made possible by the presence of triazole moieties. This effect became more remarkable at higher temperature and RH confirming the effectiveness of the functionalized fibers in enhancing membrane mechanical strength in conditions similar to those of actual fuel cell operation.

Moreover, Aq-PSUT(30 mol%) presented Young modulus higher than Aq-PSUT(90 mol%):

390 N•mm -2 vs. 385 N•mm -2 at RT, and 259 N•mm -2 vs. 220 N•mm -2 at 70 °C and 80 % RH.

This could be related to a lower mechanical strength of the PSUT(90 mol%) with respect to the PSUT(30 mol%). We previously reported [START_REF] Sood | 1,2,3-Triazole-Functionalized Polysulfone Synthesis through Microwave-Assisted Copper-Catalyzed Click Chemistry: A Highly Proton Conducting High Temperature Membrane[END_REF] that an increase in the degree of 4-heptyl-1,2,3-triazole functionalization on PSU resulted in a decrease in glass transition temperature (Tg =138 °C for PSUT(30 mol%) vs Tg =115 °C for PSUT(90 mol%), which is related to the crystallinity and therefore to the mechanical strength.

Both yield stress and breaking strength followed the same trend of the elastic modulus, and decreased by increasing temperature and relative humidity. Modulus of toughness (MT) was also estimated in order to quantify energy absorption capability of the membrane before breaking, and it followed the sequence: Aq-PSUT(30 mol%)>Aq-PSUT(90 mol%)>Aq>Aq-PSU. This result further confirmed the strength of the interaction between the ionomer and the PSUT nanofibers and the efficacy of the latter as mechanical reinforcement. No clear trend is observed in the mechanical properties, going from Aq-PSUT (30 mol%) to Aq-PSUT (90 mol%). This, combined with the proton conductivity results, suggests that the increase in the functionalization degree of the polymer does not increase the interaction between fibers and Aquivion ® , which is consistent with this interaction involving only the region at the fiber/ionomer interface.

We can conclude that Aq-PSUT(30 mol%) presented the highest strength of the membranes investigated (non-reinforced, reinforced with PSU and PSUT (90 mol%)). Furthermore, its properties are much improved over those of the non-reinforced membranes at 70 °C/80 %RH than at ambient conditions.

Membrane dimensional water swelling

The dimensional change of reinforced and non-reinforced membranes from a dry to a waterswollen state is reported in Table 3. Lower water uptake and volumetric water swelling was observed for all the composite membranes in comparison to non-reinforced Aquivion ® .

Membranes reinforced with functionalized polysulfone fibers undergo less change in dimension than the PSU reinforced membranes. This behavior is, once again, attributed to the ionic interactions taking place between the sulfonic acid sites of the PFSA and the 1,2,3triazole moieties of PSUT(30 mol%) and PSUT(90 mol%) fibers, resulting in an improved dimensional stability limiting membrane water uptake. The dimensional change of all fiber reinforced membranes is markedly anisotropic, with remarkably lower in-plane (area) variation and higher through-plane (thickness) variation compared to non-reinforced Aquivion ® . For instance, Aq-PSUT(30 mol%), Aq-PSUT(90 mol%) and Aq-PSU have area swelling of respectively 15 %, 26 % and 18 % while for the non-reinforced Aquivion ®, the area swelling is 81 %. Aquivion ® presents the lowest thickness swelling (22 %), against 61% for Aq-PSU, 54 % for Aq-PSUT(30 mol%) and 45 % for Aq-PSUT(90 mol%). Moreover, the area swelling was observed to be isotropic in all reinforced membranes (same increase in sample length and width), indicating uniform in-plane reinforcing effect of the fibers. The electrospun web thus constrains the ionomer towards swelling in the through-plane direction, due to the in-plane fiber directionality and the weak through-plane fiber interconnectivity [START_REF] Ballengee | Composite Fuel Cell Membranes from Dual-Nanofiber Electrospun Mats[END_REF][START_REF] Giancola | Composite short side chain PFSA membranes for PEM water electrolysis[END_REF]. Area stresses are considered predominant and one of the main cause of premature membrane mechanical failure [START_REF] Qiu | Mechanical failure and mitigation strategies for the membrane in a proton exchange membrane fuel cell[END_REF]. Further reduction in area swelling is desirable, and this can be achieved increasing the amount of electrospun nanofibers in the membrane over the 12 wt% of this work. In-plane membrane dimensional stabilization is thus considered crucial and a significantly enhanced lifetime has been reported

for reinforced membranes thanks to lower in-plane swelling and enhanced mechanical strength [START_REF] Powers | Electrospun tri-layer membranes for H2/Air fuel cells[END_REF].

Aq-PSUT(30 mol%) displayed the best combination of mechanical strength and ductility, dimensional stability and proton conductivity and was selected for integration into a membrane electrode assembly (MEA) and single fuel cell tests.

Membrane permeability

The hydrogen permeation values measured for Aq, Aq-PSU and Aq-PSUT(30 mol%) were as follows 2.4 x 10 -13 mol/s cm kPa, 2.1 x 10 -13 mol/s cm kPa and 1.6 x 10 -13 mol/s cm kPa. It can be concluded that the presence of the reinforcement decreased membrane permeability.

Furthermore, PSUT nanofibers proved to be an efficient barrier to hydrogen, the corresponding membranes demonstrating the lowest hydrogen permeation. In order to confirm the high performance of PSUT reinforcement in membrane durability enhancement the pristine and composite membranes were subjected to wet dry cycling at OCV.

Fuel cell polarization curves and durability

I-V curves of membrane electrode assemblies (MEAs) with membranes of Aq, Aq-PSU and Aq-PSUT(30 mol%) of thickness 30±1 µm, at 80 ºC, 100 % RH and 2.5 bar absolute (H2/air) are reported in Figure 6 a. In all cases, high OCV, with similar values ranging between 0.936 for Aq and 0.947 V for Aq-PSUT(30 mol%) were obtained. This indicates excellent gas barrier properties for all membranes. Moreover, similar polarization curves were obtained to ca. 0.6 A cm -2 for all MEAs as expected from the similar membrane proton conductivity described above, with differences in the mass transport region at higher current density, leading to maximum power densities of Aq, 0.56 W cm -2 with non-reinforced Aquivion ® , 0.52 W cm -2 with Aq-PSUT(30 mol%), and 0.48 W cm -2 with Aq-PSU.

The MEAs incorporating the reinforced and non-reinforced membranes were subjected to an accelerated ageing test protocol designed to combine chemical and mechanical degradation by performing wet-dry cycles while holding at OCV [START_REF] Mukundan | Membrane Accelerated Stress Test Development for Polymer Electrolyte Fuel Cell Durability Validated Using Field and Drive Cycle Testing[END_REF]. The voltage decay with time was recorded (Figure 6). A membrane exposed to repeated shrinking-swelling cycles eventually loses its mechanical integrity, while the free radicals produced at open circuit voltage attack the ionomer and cause chemical degradation according to pathways that are now well understood [START_REF] Zatoń | Current understanding of chemical degradation mechanisms of perfluorosulfonic acid membranes and their mitigation strategies: a review[END_REF].

The pristine Aquivion ® membrane demonstrated a lifetime of only 75 hours before the

Conclusion

This work demonstrated the effect of an active reinforcement, i.e. nanofibers interacting by hydrogen bond or ionic crosslinking with the ionomer, on mechanical properties and durability of proton exchange membranes. The reinforcement based on 4-heptyl-1,2,3-triazole functionalized PSU nanofibers resulted in a remarkable improvement in membrane mechanical strength and dimensional stability with respect to non-reinforced Aquivion ® and Aquivion ® reinforced by non-functionalized polysulfone nanofibers. The interaction at the fiber/ionomer interface between triazole (fiber) and -SO3H (ionomer), evidence for which was obtained from electron microscopy observation of a local polymer blend-like region, might be at the origin of this improved properties set. The proton conductivity of triazole functionalized PSU Aquivion ® membranes is closely similar to that of non-reinforced Aquivion ® . The significantly greater MEA durability enabled by the use of the Aq-PSUT(30 mol%) membrane (in the absence of any radical scavenging component) affords new possibilities for further improving fuel cell performance by reducing the membrane thickness. This work demonstrates the flexible approach of active fibrous reinforcement to achieve highly conducting, dimensionally stable and durable fuel cell membranes, with applications too in other energy conversion devices (electrolyzers), and opens up a range of perspectives on tuning the fiber/ionomer interface.

  C NMR (100 MHz, DMF; ppm): δ 162.70-118.16 (aromatic backbone), 42.58 (C-CH3), 41.48 (CH2Cl), 30.60 (C-CH3) (Figure S1, SI). The methylazide-functionalized polysulfone PSUAz presented the following 1 H NMR (400 MHz, DMF; ppm) chemical shifts: δ 8.05-6.90 (aromatic backbone), 4.46 (brs, 2H, CH2N3), 1.70 (brs, 6H, C-CH3). 13 C NMR (100 MHz, DMF; ppm): δ 161.82-117.90 (aromatic backbone), 49.75 (CH2N3), 42.27 (C-CH3), 30.43 (C-CH3) (Figure S1, SI).

Figure. 1 .

 1 Figure. 1. FE-SEM micrographs of electrospun nanofibers of PSU (a), PSUT(30 mol%) (b) and PSUT(90 mol%) (c) selected to be incorporated into the Aquivion ® matrix. Histograms of fiber diameter distribution of PSU (d), PSUT(30 mol%) (e) and PSUT(90 mol%) (f).

Figure. 2 .

 2 Figure. 2. SEM micrographs of freeze-fractured cross-sections of Aq-PSU (a,d), Aq-PSUT(30 mol%) (b,e) and Aq-PSUT(90 mol%) (c,f).In contrast, under the same measurement conditions, Aquivion ® appears to be in tight contact with the triazole functionalized polymer fibers, with the appearance of a halo around

Figure 3 .

 3 Figure 3. SEM micrographs at different magnifications of the cross-section of the composite membrane Aq-PSUT (30 mol%).

  a and b) However, for Aq-PSUT, a gradual transition is observed with a signal from fluorine persisting even inside the fiber (Fig.4 c and d), consistent with an Aquivion ® /PSUT(30 mol%) blend region at the PFSA/fiber interface. It is known that hydrogen bonds can enhance the miscibility of otherwise immiscible blends.[START_REF] Kuo | Hydrogen-bonding in polymer blends[END_REF]

Figure 4 . 3 . 3 .Figure 5 .

 4335 Figure 4. TEM micrograph of the Aq-PSU (a) of the Aq-PSUT(30 mol%) (c) and the related fluorine mass distribution (b, d). On TEM micrographs are indicated the analyzed region (diameter of the nanofiber, light blue line) and the corresponding fluorine mass distribution (blue line).

Figure 6 .

 6 Figure 6. a) Polarization curves of MEAs with membranes Aq (■, a), Aq-PSU (♦, b) and Aq-PSUT(30 mol%) (▲, c) at 80 ºC, 100 % RH and 2.5 bar abs (H2/air), and corresponding power density Aq(□, a'), Aq-PSU(◊, b') and Aq-PSUT(30 mol%) (Δ, c'); b) OCV hold at 90 °C as a function of time for MEAs incorporating PSUT(30 mol%) nanofiber reinforced Aquivion ® (c), PSU reinforced Aquivion ® (b), non-reinforced Aquivion ® membrane (a).

Table 1 .

 1 Experimental ion exchange capacity (IEC) values of Aquivion ® and reinforced membranes with 12 wt% fiber content.

	Membrane	IEC
		(mequiv•g -1 )
	Aq830	1.17
	Aq-PSU	1.01
	Aq-	0.98
	PSUT(30 mol%)
	Aq-	1.02
	PSUT(90 mol%)

Table 3 .

 3 Water uptake (WU) and dimensional water swelling of pristine Aquivion ® membrane and PSU and functionalized PSUT reinforced Aquivion ® membranes between dry and swollen states in liquid water at 80 °C for 15h.

	Membrane	WU (%) Volume swelling (%) Area swelling (%) Thickness swelling (%)
	Aq	73±3	121±4	81±3	22±4
	Aq-PSU	53±3	91±4	18±2	61±4
	Aq-PSUT(30 mol%) 45±3	77±4	15±2	54±4
	Aq-PSUT(90 mol%) 48±3	83±4	26±2	45±4

Acknowledgements: The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013)/ERC Grant Agreement n. 306682. SC acknowledges the financial support from the French IUF.