Safety and efficacy of Ayush 64 tablets as an adjunct therapy and its effect...

[Full title: Safety and efficacy of Ayush 64 tablets as an adjunct therapy and its effect on bio- markers in mild to moderate covid-19 patients]

[Find the meaning and references behind the names: Suresh, Hassan, Lal, Kumar, Ayu, Prasanna, Narasimha, China, Ayush, Ambika, Rao, Med, Rich, Suhas]

251 Journal of Ayurvedic and Herbal Medicine 2021; 7(4): 251-256 Research Article ISSN: 2454-5023 J. Ayu. Herb. Med. 2021; 7(4): 251-256 Received: 13-10-2021 Accepted: 29-11-2021 © 2021, All rights reserved www.ayurvedjournal.com DOI: 10.31254/jahm.2021.7407 *Corresponding author: Dr. Nataraj HR Associate Professor, Department of Agadatantra, SDM College of Ayurveda & Hospital, Hassan- 573201, Karnataka, India Email: drnatarajhr@sdmcahhassan.org Safety and efficacy of Ayush 64 tablets as an adjunct therapy and its effect on biomarkers in mild to moderate covid-19 patients Nataraj HR 1 , Suresh R M 2 , Chennaveerappa 2 , Prasanna Narasimha Rao 1 , Suhas Kumar Shetty 1 , Gopkrishna S 1 , Ambika S 3 , Padmanabha Lal 3 1 Faculty, Department of Agadatantra, SDM College of Ayurveda & Hospital, Hassan- 573201, Karnataka, India 2 Faculty, Hassan Institute of Medical Sciences, Hassan, Karnataka- 573201, India 3 Post Graduate Scholars, SDM college of Ayurveda & Hospital, Hassan, Karnataka- 573201, India ABSTRACT Background: COVID- 19 has emerged as the latest pandemic that erupted in the Wuhan City of People’s Republic of China in December 2019, which is affecting human health and economy across the world. The ongoing COVID-19 outbreak in developed countries also highlights the fact that developed countries and rich populations are not immune to infectious disease outbreaks. Coronaviruses (CoVs) are enveloped, single-stranded, positive-sense RNA viruses that belong to the Coronaviridae family. SARS-CoV-2 is a member of the beta CoV genus, which also includes SARS-CoV-1 and MERS-CoV. The lack of approved effective drug therapeutic protocols for CoVs would make treating newly emerged COVID-19 infections globally difficult. Objective: A clinical study was conducted to evaluate the safety and efficacy of AYUSH 64 a poly herbal drug as an adjunct therapy to standard of care in mild to moderate Covid 19 patients Materials and methods: A prospective, open-label, randomized, parallel assignment, single-center clinical study with pre-test and post-test design was conducted at Covid hospital, Hassan institute of medical sciences between September 2020 to December 2020. A total 60 diagnosed cases (22-75 years of age) of Covid 19 were randomly allocated to both the groups. Control group received standard of care (SOC) as prescribed by the ICMR/WHO and state government, Trial Group received one week intervention of AYUSH 64 tablets at the dose of 2 gm/day along with standard of care Assessment of parameters viz. improvement in the symptoms, hematology, liver function, kidney function tests, acute phase reactants, Serum ferritin, Di dimer, LDH, and hsCRP were analyzed on day zero, day three and day seven. Results: One-week intervention of AYUSH-64 along with SOC helped to recover from Covid 19 symptoms. The intervention was safe on blood and biochemical parameters. Trial group has shown significant reduction in acute phase reactants viz hsCRP, LDH, Di dimer and Ferritin compare to the control group. No serious drug adverse effects were observed during the study. Conclusion : AYUSH-64 along-with standard care in mild to moderate covid 19 patients is safe and efficacious and this may be used as add-on to standard care for early recovery and better outcome. Keywords: Ayush 64, Acute phase reactants, Covid 19, Covid bio markers. INTRODUCTION COVID-19 has emerged as the latest pandemic that erupted in the People's Republic of China's Wuhan City in December 2019, affecting human health and the global economy. According to the WHO Coronavirus disease 2019 (COVID-19) Situation Report, approximately 158,651,638 cases are reported globally, with 3,299,764 deaths as of May 11 th, 2021. The ongoing COVID-19 outbreak in developed countries also highlights the fact that developed countries and rich populations are not immune to infectious disease outbreaks. Coronaviruses (CoVs) are enveloped, single-stranded, positive-sense RNA viruses that belong to the Coronaviridae family. SARS-CoV-2 is a member of the beta CoV genus, which also includes SARS-CoV-1 and MERS-CoV. The lack of approved effective drug therapeutic protocols for CoVs would make treating newly emerged COVID-19 infections globally difficult. Drug repurposing, defined as identifying alternative uses for approved or investigational drugs outside of their defined indication, could be a possible way to overcome the time constraints of research and development required to design a therapeutic drug to combat the pathogen. As a result, drug repurposing or repositioning can facilitate faster clinical decisions at a lower cost than de novo drug development [1] . Though drug repurposing is sometimes based on chance observations, target-based drug repurposing is dependent on prior knowledge of the precise molecular or cellular element recognized by the proposed drug [2, 3] . Ayurveda and traditional systems of medicine in India have been treating infectious and

[Find the meaning and references behind the names: Ili, Main, Full]

252 Journal of Ayurvedic and Herbal Medicine|October-December|2021 non-infectious diseases equally with high success rates, treating patients through an individualized person-to-person approach based on the patient's needs. The Central Council for Research in Ayurvedic Sciences, India's apex body for Ayurvedic research and development under the Ministry of AYUSH, has developed a poly-herbal drug called AYUSH 64 after extensive pharmacological, toxicological, and clinical studies. AYUSH 64 was found to be safe and non-toxic in experimental studies at a dose of 500 mg/kg weight for 12 weeks. Aside from its antimalarial activity, it has been shown to be effective in fevers of unknown a etiology, filarial lymphangitis, and liver function derangement [4, 5, 6] . Taking cues from the clinical experiences of physicians who had successfully used AYUSH-64 for the treatment of Influenza-like illness (ILI), the Central Council for Research in Ayurveda and Siddha conducted a pilot study (CCRAS). This sparked the idea of repurposing AYUSH 64 for use in the management of Covid-19 positive cases who also present with influenza-like symptoms due to respiratory tract affliction. Saptaparna ( Alstonia scholaris R. Br.), Katuki ( Picrorhiza kurroa Royle ex. Benth), Kiratatikta ( Swertia Chirata Pexbex. Karst), and Kuberaksha ( Caesalpinia crista Linn.) are among the Ingredients of AYUSH 64. Anti-inflammatory and immunomodulatory activities have been demonstrated in studies on the ingredients of AYUSH-64. Total alkaloids from Saptaparna have been shown in animal studies to inhibit the production of inflammatory cytokines TNFand IL-8 in the BALF and lung [7] Swertia chirata inhibited the expression of Vpr in Hela cells harbouring the TREx plasmid encoding full-length Vpr, and crude extract of the whole Swertia chirata plant inhibited the expression of Vpr in Hela cells harbouring the TREx plasmid encoding full-length Vpr (TREx-HeLa-Vpr cells) [8, 9] . OBJECTIVES Primary Objective To compare the efficacy and safety of a combination regimen of Standard of Care (SOC) plus a selected standardised Ayurvedic drug (AYUSH 64 as adjuvant) in the treatment of mild to moderate COVID-19 to that of a standalone SOC regimen (active control). Secondary Objective To determine the therapeutic effect of a combination of SOC plus a standardized Ayurvedic drug (AYUSH 64) on surrogate markers of COVID - 19 disease severity and complications, and recovery pertaining to broad based domain of acute phase reactants. OUTCOMES Primary Outcomes 1 Improvement in selected laboratory parameters: blood hemoglobin, differential and total leukocyte counts, liver enzymes, renal functions, acute phase reactants, serum ferritin. STUDY DESIGN Table 1: Study design Study Type Interventional (Clinical Trial) Enrollment size 60 (30 Patients in each group) Allocation Randomized Intervention Model Parallel Assignment Intervention Model Description The main trial was an open label, randomized, Controlled trial which was conducted in the patients of COVID-19 in selected site Hassan institute of Medical sciences Hassan. Randomization was 1:1, stratified by severity of illness, to either with or without AYUSH-64 for 7 days. All patients were also received a supportive care/treatment according to ICMR/WHO guidelines for COVID-19 Purpose Treatment Masking None (open label) Control Controlled Timing Prospective No. of Groups Two Interventions Group I: (Ayurveda as add-on to standard care as per guidelines) AYUSH 64 Dose 2 Tablets (500 mg each) twice daily Dosage form Tablets Route of Administration Oral Time of Administration Twice a day after food Anupana Water Duration of therapy 7 days Group-II: Conventional standard therapy as per ICMR/WHO guidelines ASSESMENT PARAMETERS Clinical assessment: Daily monitoring of Symptoms like Fever, breathless ness, Loss of taste and smell, headache, Myalgia, General Debility, Vomiting, Diarrhea, Headache Daily monitoring of Body temperature, SPO 2, Blood pressure Clinical Laboratory Parameters The following laboratory tests were performed on the day of admission, day three and day seven in both the groups

[Find the meaning and references behind the names: Real]

253 Journal of Ayurvedic and Herbal Medicine|October-December|2021 Confirmatory Diagnostic Test : Nasal and / or Throat swab for real time RT - PCR for SARS - CoV - 2, Haemogram, Liver function test, Kidney Function, Blood Sugar Level, Urine Routine, C-Reactive protein titer, LDH, Ferritin, D-Dimer, Serum Electrolytes RESULTS Between September 2020 and December 2020, a total 78 participants were screened, out of which 60 were enrolled in the study (Fig 1). Eligible subjects were allocated to two groups using simple randomization after obtaining the consent. In Group one the patients were given AYUSH 64 for 7 days only along with Standard of care and in group two only standard of care was administered and response was recorded on Day one and Day 7 in both the groups. Figure 1: CONSORT 2010 Flow diagram Table 2: Clinical features n=60 S. No Symptoms Frequency 1 Fever 18 2 Cough 23 3 Sore throat 13 4 Shortness of breath 14 5 Diarrhoea 04 6 Headache 11 7 Loss of taste 13 8 Tiredness 30 9 Skin rashes 00 Table 3: Trial group paired t test statistics S. No Assessment BT AT (Day 7) SD SE T test value P value 1 SpO 2 90.2 91.4 3.53 0.644 2.43 <0.05 2 Hb% 14.27 14.4 0.993 0.181 1.11 0.50 3 WBC 9686 11988 5.52 1.00 2.21 0.03 4 Platelet 1.91 lakh 2.66 lakh 76.08 13.89 8.54 0.001 5 RBC 4.88 5.22 0.5046 0.0921 7.24 0.009 6 LDH 419.66 287.86 152.60 27.86 4.731 0.000 7 Ferritin 766.85 432.19 414.52 75.68 8.23 0.0001 Enrollment • Assesed for eligibility (n=78) • Excluded (n=18) • Not meeting inclusion criteria (n=16) • Declined to participate (n=02) • other reasons(n=0) Allocation • Allocated to Group 1 (SOC+AYUSH 64) n=30 • Alocated to group 2 (only SOC) n=30 Follow up • Lost follow up in Group 1 n=0 • lost folowup in Group 2 n=04 • Discontinued intervention reasons n=04 shifted to other covid care centre Analysis • Analysis in Group 1 n=30 • Analysis in group 2 n=26 • Excluded from analysis in group 2 n=4 as after treatment data was not available

[Find the meaning and references behind the names: Seed, Zhao, Yun]

254 Journal of Ayurvedic and Herbal Medicine|October-December|2021 8 Hs-CRP 75.25 24.46 44.82 8.183 6.75 0.01 9 Trop-T 0.019 0.177 3.825 0.6984 1.65 0.23 10 D-DIMER 2094.9 1301.6 1849.28 337.6 6.78 0.001 Table 4: control group paired t test statistics S. No Assessment BT AT (Day 7) SD SE T test value P value 1 SpO 2 88.88 89.15 3.86 0.757 0.43 0.34 2 Hb% 14.16 13.99 0.869 0.170 1.11 0.31 3 WBC 10918 11955 5.11 1.00 2.27 0.23 4 Platelet 2.32 lakh 2.49 lakh 85.59 16.78 8254 0.02 5 RBC 5.02 5.08 0.344 0.0674 4.24 0.053 6 LDH 407.4 388.8 186.04 36.48 0.508 0.61 7 Ferritin 628 640 344.07 67.47 8.23 0.03 8 Hs-CRP 48.51 33.08 53.21 10.43 5.75 0.01 9 Trop-T 0.026 0.007 2.671 0.5239 1.65 0.23 10 D-DIMER 1891.33 2007.27 1996.21 391.48 6.78 0.04 DISCUSSION Including both groups maximum subjects were males (80%), common age group 28-54 (70% of the subjects) all subjects tested negative for HCV, HBSAg, and HIV. Among all the subjects (n=56) most common symptom observed is general tiredness (91%) followed by cough (60%), fever (51%), shortness of breath (48%), loss of taste (41%) and sore throat (37%) whereas diarrhea and headache were observed in least number of subjects. When compared with in the group both the groups have shown significant changes in terms of improvement in oxygen saturation levels in study duration. Statistically significant increase was observed in both groups in Hb%, RBC, and significant decrease was observed in LDH (p value 0.000), Ferritin (p value 0.0001), and hsCRP (p value 0.01) in trial group when compared to the control group. Highly significant increase in WBC (p value 0.03) and Highly significant decrease in Di dimer (p value 0.04) values observed only in trial group. Di dimer values are indicative of severe infection and altered coagulopathy and one of the associate indicators for the assessment of severity in covid 19 patients which is complimented by decreased APTT in trial group on day seven. Decreased values of di dimer and hsCRP, LDH in trial group along with increased WBC values it evident that AYUSH 64 positively complimented along with SOC in reducing acute phase reactants and inflammatory markers in mild to moderate covid 19 patients. Hematological parameters including liver function test and kidney function tests were within the normal limits during the treatment period and no significant change was observed in either of them in both the groups. Secondary complications of covid 19 were not observed in any participant during the study period in both the groups. The ingredients in AYUSH-64 have been shown to have antiinflammatory and immune modulatory properties. The aqueous extract of A. scholaris bark induced the cellular immune response in BALB/c mice at 50 mg/kg body weight once a day for 7 days, while 100 mg/kg body weight inhibited the delayed type of hypersensitivity reaction [10] . Yun-Li Zhao et al . investigated the effects of A. scholaris indole alkaloids and total alkaloids on post-infectious cough in mice and airway inflammation in rats, respectively. Indole alkaloids suppressed inflammatory cells, cytokines (IL-6), and antioxidant balance. TNFand IL-8 production in bronchoalveolar lavage fluid and lung was inhibited by total alkaloids [11, 12] . The biopolymeric fraction RLJ-NE-205 derived from the rhizomes of P. kurroa improved the immune system of mice by increasing lymphocyte proliferation and cytokine levels (IL-4 and IFN-gamma) in serum, phagocytic index, and CD 4/CD 8 population [13] . Pretreatment with P. kurroa rhizome extract demonstrated anti-inflammatory activity by suppressing macrophage-derived cytokines (TNF-, IL-1, IL-6, IL-10) and mediators via NF-B signalling [14] . Picroside II, an active ingredient from P. scrophulariiflora, has been shown to have promising antiinflammatory effects in cells and animals by lowering TNF-, IL-1, and IL- 6 concentrations. When compared to lipopolysaccharide (LPS) stimulation, it inhibited the activation of the p 65 NF-B signalling pathway. Pathologic changes in lung tissues had been alleviated, and the lung wet/dry weight ratio had been reduced [15] . At IC 50 concentrations, Swertia chirata inhibited NF-kB/DNA interactions and reduced pro-inflammatory IL-8 expression in cystic fibrosis cells [16] , the two main xanthones from S. chirata , bellidifolin and swerchirin, inhibit the production of the pro-inflammatory cytokines IL-6 and TNF-. Bellidifolin inhibited prostaglandin E 2 (PGE 2) production by suppressing cyclooxygenase-2 protein expression (COX- 2) [17] . In HeLa cells harbouring the TREx plasmid encoding full-length Vpr (TREx-HeLa-Vpr cells), a CHCl 3 soluble crude extract of S. chirata inhibited the expression of Viral protein R (Vpr) [18] . A crude extract of the swertia plant was found to have antiviral properties against the Herpes simplex virus type-1 [19] . TNF-, IL-1, and IL-6 mRNA expression were reduced in the hippocampus and frontal cortex brain areas of rats after administration of methanolic extracts of C. crista [20] Caesalpinia bonducella seed

[Find the meaning and references behind the names: Liu, Srikanth, Sharma, Singh, Raj, Gupta, Abraham, Wang, Patil, Huang, Sid, Cherian, Ito, Rasa, Ama, Dutt, Basu, Ngwe, Verma, Satti, Nil, Chen, Yang, Sama]

255 Journal of Ayurvedic and Herbal Medicine|October-December|2021 extract increased antibody production and delayed type hypersensitivity in rats in a dose-dependent manner, indicating promising immune stimulant properties [21] . A water-soluble glucoarabinan isolated from the alkaline extract of the endosperm of C. bonducella seeds demonstrated immune stimulant activity by activating splenocytes and thymocytes [22] . In experimental albino rats, whole ethanolic seed extract of C. bonducella seeds exhibited antiinflammatory, antipyretic, and analgesic activities [23, 24] . According to Ayurvedic theory, weakened agni in jvara causes the formation of ama (proinflammatory undigested substance), which results in amavastha and a weakened immune system. The Ayurvedic treatment aims to correct agni status through agnideepana and amapachana , resulting in niramavastha (absence of ama), i.e. reversing the pathology and improving immune status. Tikta rasa ( Agnideepak and Amapachak ) is one of main rasa in the ingredients in AYUSH 64; hence AYUSH 64 takes on the role of amapachaka and changes the status of agni from sama to nirama . Niramavastha which has the effect of restoring normal metabolism and strengthening the immune system. The seven-day treatment of AYUSH 64, combined with standard care, resulted in symptom reduction in the majority of patients. Symptoms such as fever, cough, and headache, as well as difficulty breathing and general debility, had significantly decreased by the seventh day of intervention. However, AYUSH 64 for a longer period of time may have produced better results in terms of metabolism restoration and immune system strengthening. In terms of herb – drug interactions, AYUSH 64 was used earlier in conjunction with standard antimalarial treatment. The seven-day administration of AYUSH 64 in conjunction with standard care was well tolerated in this study, and no adverse events were recorded. Limitations This is pre and post-test design study with a small sample size and was focused on mild to moderate covid 19 patients only clinical safety related biochemical parameters. However, for substantial evidence, a larger multicentric study with a robust design with standard diagnostic investigations and cytokine response will be helpful to validate the efficacy of this drug. CONCLUSION AYUSH 64 along-with standard care in mild to moderate covid 19 patients is safe and efficacious it may be used as an add-on to standard care for early recovery and better outcome Clinical Trial Registry Clinical Trial Registry of India (CTRI), number: CTRI/2020/07/035245. Source of support CCRAS, Ministry of AYUSH, GoI. Provided trial drug AYUSH 64 for the study. Conflict of Interest None declared. Financial support Nil. Acknowledgment The authors are thankful to Director, CCRAS, Ministry of AYUSH, Government of India for providing trial drug AYUSH 64 for this study, Director and District surgeon Hassan institute of medical sciences, Hassan for providing necessary resources for the study. REFERENCES 1 Cherian SS, Agrawal M, Basu A, Abraham P, Gangakhedkar RR, Bhargava B. Perspectives for repurposing drugs for the coronavirus disease 2019. The Indian journal of medical research. 2020;151(2-3):160. 2 Pushpakom S, Iorio F, Eyers PA, Escott KJ, Hopper S, Wells A, et al . Drug repurposing: Progress, challenges and recommendations. Nat Rev Drug Discov 2019; 18: 41-58. 3 Nosengo N. New tricks for old drugs. Nature 2016; 534: 314-6. 4 Sharma KD, Kapoor ML, Vaidya SP, Sharma L. A clinical trial of “AYUSH 64”(a coded antimalarial medicine) in cases of malaria. Jour Res A y Sid. 1981;2(4):309-26. 5 Anonymous. 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Journal of bioscience and bioengineering. 2019;128(4):445-9. 10 Iwo MI, Soemardji AA, Retnoningrum DS. Immunostimulating effect of pule ( Alstonia scholaris LR Br., Apocynaceae) bark extracts. Clinical hemorheology and microcirculation. 2000;23(2, 3, 4):177-83. 11 Zhao YL, Yang ZF, Shang JH, Huang WY, Wang B, Wei X et al . Effects of indole alkaloids from leaf of Alstonia scholaris on post-infectious cough in mice. Journal of ethnopharmacology. 2018;218:69-75. 12 Zhao YL, Shang JH, Pu SB, Wang HS, Wang B, Liu L et al . Effect of total alkaloids from Alstonia scholaris on airway inflammation in rats. Journal of ethnopharmacology. 2016;178:258-65. 13 Gupta A, Khajuria A, Singh J, Bedi KL, Satti NK, Dutt P et al . Immunomodulatory activity of biopolymeric fraction RLJ-NE-205 from Picrorhiza kurroa. International Immunopharmacology. 2006;6(10):1543-9. 14 Kumar R, Gupta YK, Singh S, Raj A. Anti-inflammatory effect of Picrorhiza kurroa in experimental models of inflammation. Planta medica. 2016;82(16):1403-9. 15 Shen B, Zhao C, Chen C, Li Z, Li Y, Tian Y, Feng H. Picroside II protects rat lung and A 549 cell against LPS-induced inflammation by the NF- κB pathway. Inflammation. 2017;40(3):752-61. 16 Guerrini A, Mancini I, Maietti S, Rossi D, Poli F, Sacchetti G et al . Expression of pro‐inflammatory interleukin‐8 is reduced by ayu rvedic decoctions. Phytotherapy Research. 2014;28(8):1173-81. 17 Hu TY, Ju JM, Mo LH, Ma L, Hu WH, You RR et al . Anti-inflammation action of xanthones from Swertia chirayita by regulating COX-2/NF- κB/MAPKs/Akt signaling pathways in RAW 264.7 macrophage cells Phytomedicine. 2019;55:214-21.

[Find the meaning and references behind the names: Ramesh, Islam, Ravi, Mandal, Vincent]

256 Journal of Ayurvedic and Herbal Medicine|October-December|2021 18 Woo SY, Win NN, Oo WM, Ngwe H, Ito T, Abe I et al . Viral protein R inhibitors from Swertia chirata of Myanmar. Journal of bioscience and bioengineering. 2019;128(4):445-9. 19 Verma H, Patil PR, Kolhapure RM, Gopalkrishna V. Antiviral activity of the Indian medicinal plant extract, Swertia chirata against herpes simplex viruses: A study by in-vitro and molecular approach. Indian Journal of Medical Microbiology. 2008;26(4):322-6. 20 Ravi SK, Ramesh BN, Mundugaru R, Vincent B. Multiple pharmacological activities of Caesalpinia crista against aluminium-induced neurodegeneration in rats: relevance for Alzheimer’s disease. Environmental toxicology and pharmacology. 2018;58:202-11. 21 Shukla S, Mehta A, Mehta P, Vyas SP, Shivaprasad HN. In vivo immunomodulatory activities of the aqueous extract of bonduc nut Caesalpinia bonducella seeds. Pharmaceutical biology. 2010;48(2):227-30. 22 Mandal EK, Mandal S, Maity S, Behera B, Maiti TK, Islam SS. Structural studies of an immunostimulating gluco-arabinan from seeds of Caesalpinia bonduc. Carbohydrate polymers. 2013;92(1):704-11. 23 Shukla S, Mehta A. In vivo anti-inflammatory, analgesic and antipyretic activities of a medicinal plant, Caesalpinia bonducella F. Pakistan journal of pharmaceutical sciences. 2015 Jul 2;28. 24 Shukla S, Mehta A, Mehta P, Vyas SP, Shukla S, Bajpai VK. Studies on antiinflammatory, antipyretic and analgesic properties of Caesalpinia bonducella F. seed oil in experimental animal models. Food and Chemical Toxicology. 2010;48(1):61-4. HOW TO CITE THIS ARTICLE Nataraj HR, Suresh RM, Chennaveerappa, Rao PN, Shetty SK, Gopkrishna S, Ambika S, Lal P. Safety and efficacy of Ayush 64 tablets as an adjunct therapy and its effect on biomarkers in mild to moderate covid-19 patients. J Ayu Herb Med 2021;7(4):251-256. DOI: 10.31254/jahm.2021.7407 Creative Commons (CC) License- This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY 4.0) license. This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. (http://creativecommons.org/licenses/by/4.0/).

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