A multi-center collaborative double-blind study on clinical evaluation of AYUSH-SL in patients receiving mass drug administration for management of chronic filarial lymphedema—study protocol

Santanu K Tripathi; Renu Singh; Gurucharan Bhuyan; Achintya Mitra; Lalita Sharma; Chagam R Murlikrishna; Kshirod K Ratha; Sujata Dhoke; Bharti Gupta; Adarsh Kumar; Narayanam Srikanth

doi:10.4103/jras.jras_46_21

PROTOCOL

Year : 2021 | Volume : 5 | Issue : 3 | Page : 139-147

A multi-center collaborative double-blind study on clinical evaluation of AYUSH-SL in patients receiving mass drug administration for management of chronic filarial lymphedema—study protocol

Santanu K Tripathi¹, Renu Singh², Gurucharan Bhuyan³, Achintya Mitra⁴, Lalita Sharma², Chagam R Murlikrishna⁵, Kshirod K Ratha³, Sujata Dhoke⁵, Bharti Gupta⁶, Adarsh Kumar², Narayanam Srikanth²
¹ Department of Clinical and Experimental Pharmacology, Calcutta School of Tropical Medicine, Kolkata, India
² CCRAS, New Delhi, India
³ Central Ayurveda Research Institute (CARI), Bhubaneswar, Odisha, India
⁴ Central Ayurveda Research Institute (CARI), Kolkata, India
⁵ Regional Ayurveda Research Institute (RARI), Vijayawada, Andhra Pradesh, India
⁶ Central Ayurveda Research Institute (CARI), Punjabi Bagh, New Delhi, India

Date of Submission	17-Aug-2021
Date of Acceptance	11-Feb-2022
Date of Web Publication	22-Mar-2022

Correspondence Address:
Dr. Renu Singh
Central Council for Research in Ayurvedic Sciences (CCRAS), 61–65, opp. D' Block, Janakpuri Institutional Area, Janakpuri, New Delhi 110058
India

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jras.jras_46_21

Abstract

BACKGROUND: Lymphatic filariasis (LF) is caused by Wuchereria bancrofti, Brugia malayi, or B. timori, the parasitic nematodes that are transmitted through mosquitoes. The World Health Organization has recommended mass drug administration (MDA) as preventive chemotherapy strategy. In Ayurveda, LF has been narrated under the heading Shlipada, and the literature also includes detailed therapeutic measures. AYUSH-SL is a formulation prepared based on the classical information of herbs used in treating LF. MATERIALS AND METHODS: This is a multi-center, prospective, parallel-group, placebo-controlled, double blind interventional study. A total of 180 participants are planned to be recruited, and the duration of the study will be 2.5 years. Diagnosed cases of filariasis (as evidenced by a positive immune-chromatographic card test or enzyme-linked immunosorbent assay or microfilariae (MF) test report) of either sex within the age group of 18–65 years present with features of filarial lymphedema of Grade I and Grade II (lower limbs) will be included in the trial. The study group will be treated with MDA and AYUSH-SL tablet. The control group will receive MDA and a matching placebo. The duration of the therapy will be of 12 weeks, and a last follow-up will be done at the 24^th week. OUTCOME MEASURES: The primary outcome measure is reduction of edema computed for analysis by percentage reduction from the baseline at 4^th, 12^th, and 24^th week. The secondary outcome measures include improvement in quality of life (QoL), which will be assessed using the Lymphatic Filariasis Specific QoL Questionnaire, and prevention of recurrence of acute episodes, which will be assessed by probing the presence and severity of episodes in the past 4 weeks. DISCUSSION: The deformities and disability due to filarial lymphedema vanquish physically as well as mentally leading to hardships in normal life. Management of filarial lymphedema is required for preventing disease progression and episodes of adenolymphangitis. The study is expected to provide evidence on the coded Ayurveda drug AYUSH-SL in the management of chronic filarial lymphedema.

Keywords: Adenolymphangitis, elephantiasis, lymphatic filariasis, lymphedema, Shlipada

How to cite this article:
Tripathi SK, Singh R, Bhuyan G, Mitra A, Sharma L, Murlikrishna CR, Ratha KK, Dhoke S, Gupta B, Kumar A, Srikanth N. A multi-center collaborative double-blind study on clinical evaluation of AYUSH-SL in patients receiving mass drug administration for management of chronic filarial lymphedema—study protocol. J Res Ayurvedic Sci 2021;5:139-47

How to cite this URL:
Tripathi SK, Singh R, Bhuyan G, Mitra A, Sharma L, Murlikrishna CR, Ratha KK, Dhoke S, Gupta B, Kumar A, Srikanth N. A multi-center collaborative double-blind study on clinical evaluation of AYUSH-SL in patients receiving mass drug administration for management of chronic filarial lymphedema—study protocol. J Res Ayurvedic Sci [serial online] 2021 [cited 2023 Mar 25];5:139-47. Available from: http://www.jrasccras.com/text.asp?2021/5/3/139/340291

Trial registration: CTRI/2019/08/020498.

Background

Lymphatic filariasis (LF) is a disfiguring disease caused by filarial parasites, viz., Wuchereria bancrofti, Brugia timori, and B. malayi transmitted by Aedes Anopheles, Culex, and Mansonia mosquitoes.^[1] Transmission in 90% of the cases in Asia are due to W. bancrofti.^[2] LF is the foremost cause of disfigurement and the second major cause of disability globally.^[3] Seventy million people are affected worldwide, whereas approximately 40 million people globally suffer predominantly from lymphedema, elephantiasis, and hydrocele.^[4] Globally 25 million people are estimated to be affected by hydrocele, and approximately 15 million people suffer from lymphedema.^[5] An estimated 859 million people from 50 countries across the world are threatened and require preventive chemotherapy.^[2] In India, indigenous cases have been reported from about 257 districts in 21 states/union territories where about 650 million populations are at risk and active transmission persists in 160 districts.^[6] The infection of the filarial parasites in the endemic areas mostly occurs in the early childhood, and clinical signs appear much later in the life.^[2] It can be acute, asymptomatic, or chronic infection. Acute condition is usually associated with fever, chills, swelling, and local pain. In chronic conditions, in response to the filarial parasite, lymphatic dysfunction causes severe morbidity, which includes lymphedema and hydrocele with adenolymphangitis (ADL). Lymphatic dysfunction may last for some years during which the infected person may still transmit the disease.^[7] Lymphedema and its advanced stage elephantiasis primarily occur in lower extremity and are quite common in women.^[2] In 2000, the World Health Organization (WHO) launched Global Programme to Eliminate Lymphatic Filariasis (GPELF) with an aim to eliminate it globally by 2020.^[8] India envisaged to eliminate the disease by the year 2021.^[9] India’s strategy for tackling LF follows the WHO’s recommended preventive chemotherapy involving a mass drug administration (MDA) program in endemic zones and providing Morbidity Management and Disability Prevention (MMDP) services to affected individuals.^[10]

As per Ayurvedic literatures, the disease is called as Shlipada. Etymologically, Shlipada in Sanskrit is derived from “Shli” (stone) and “Pada” (foot), which refers to stony limb or foot of an elephant.^[11] Ayurveda literature includes detailed description of classification,^[12] epidemiology, clinical features,^[13],[14] etiopathogenesis, and prognosis^[15] of Shlipada. As per Ayurvedic classics, the disease is prevalent in Anupdesha (marshy lands), which is a suitable environment for growth of vectors. Such climate causes vitiation of the Tridoshas (Vata, Pitta, and Kapha) with Kapha Dosha predominance, which further plays an important role in the progression of disease. According to Ayurveda, Shlipada is of three types. Vataja Shlipada is characterized by blackish swelling, rough and dry skin surface, pain that is irregular in character, and fever. Pittaja Shlipada is characterized by yellowish soft swelling with burning sensation and fever. Kaphaja Shlipada is characterized by unctuous, whitish, and pale hard swelling with mild pain.^[2],[8] In view of predominance of Kapha Dosha, the line of treatment of Shlipada includes therapeutic measures such as palliative therapy for Kapha Dosha, Langhana (fasting), application of pastes such as Sarshapa (mustard) Lepa, Sweda Karma (fomentation), Virechana (purgation), Raktamokshana (bloodletting), and Ushnopachara (heat therapy).^{[9],[10],[16]}

Rationale for the study

India has considerably progressed towards elimination of LF; yet, several challenges persist such as drug resistance risk and recrudescence of disease into the areas with interrupted transmission due to migration of the infected people.^[17] Preventive therapy plays a major role in efforts to control the disease by eradicating microfilariae.^[2] Measures such as surgery may be required to treat individuals suffering from LF. However, after stopping MDA in endemic areas, the morbidity management may become challenging as infected patients still have to endure the disease while living in community. In other words, it can be interpreted that in the present scenario there is no effective drug to control ADL, progression of filariasis, and lymphedema. Several herbal and herbo-mineral drugs are mentioned in the classical texts of Ayurveda for the management of Shlipada, which needs to be studied and validated for their efficacy in such conditions.

Based on the experiences of Ayurveda experts who have found this drug effective in manifested cases of filariasis, the Central Council for Research in Ayurvedic Sciences (CCRAS), Ministry of Ayush, Government of India has developed an Ayurvedic herbal coded tablet “AYUSH-SL” for the management of FL. The standardization and safety/toxicity studies of this herbal formulation showed microfilaricidal activity against adult worms both in vitro and in vivo.^{[18],[19],[20]} However, clinical efficacy of AYUSH-SL is yet to be proven. As a measure of safety, it has been planned to study AYUSH-SL as add-on therapy along with MDA for the management of chronic FL.

Study objectives

The study is primarily designed with an objective to determine the efficacy of AYUSH-SL in reducing the symptoms of FL in patients who are receiving MDA treatment. The secondary objectives of the study are to evaluate clinical safety, improvement in quality of life (QoL), and percentage decrease in the recurrence of acute episodes of symptoms of LF.

Study design

The study is a prospective, double blind, parallel-group, placebo-controlled, multi-centric clinical interventional study at the outpatient department (OPD) clinic and community-based level. The duration of the study is 2.5 years. The trial is registered at CTRI CTRI/2019/08/020498.

Methods: Participants, interventions, and outcomes

Study setting

The study will be conducted at three participating centers, viz., Calcutta School of Tropical Medicine, Kolkata in collaboration with Central Ayurveda Research Institute for Drug Development (CARIDD), Kolkata; Central Ayurveda Research Institute for Hepatobiliary Disorders (CARIHD), Bhubaneswar; and Regional Ayurveda Research Institute for Skin Disorders (RARISD), Vijayawada.

Participant eligibility criteria for enrollment

Diagnosed cases of filariasis [as evidenced by a positive immune-chromatographic card test (ICT) or enzyme-linked immunosorbent assay (ELISA) or MF test report] presenting with features of LF, who are receiving MDA, will be considered as potential participants. Individuals eligible as per screening criteria and willing to give written inform consent to participate in the study will be included as participants.

Inclusion criteria

The inclusion criteria of study participants include individuals of either sex within the age group 18–65 years, with a clinical diagnosis of filariasis (with positive ICT or ELISA or MF with documentary evidence), diagnosed with filarial lymphedema for at least 1 year, manifested cases of filariasis with lymphedema of Grade I and II (lower limbs), and having history of taking the MDA within the last 5 months (but not within the last 1 week), and willingness to give informed consent.

Exclusion criteria

The exclusion criteria for the study include diagnosis of filarial lymphedema with more than 5 years duration as evidenced by supporting documents (prescription, laboratory diagnosis report, clinical profile, etc.); presence of deformity (nodular deformity, thorny deformity, anthill-like deformity); edema of any other origin (myxedema, etc.); having history of continued medication for the treatment of any chronic disease such as epilepsy, chronic liver disease, chronic kidney disease, ischemic heart disease, and congestive cardiac failure; uncontrolled diabetes mellitus (HbA1c >7.5); patients on anti-hypertensive drugs such as diuretics; patients on tetracycline therapy; Hb levels less than or equal to 9 g/dL; pregnancy or lactating women; acute cases of dermato-lymphangio adenitis (ADLA) without any features of FL; and patients with history of alcoholism/drug dependence.

The participant may be withdrawn from the trial if he/she develops any serious adverse reaction (necessitating hospitalization) or if there is non-compliance of the treatment regimen (minimum 80% compliance is essential to continue in the study). The decision to withdraw a participant from the trial would be taken only by the investigators, who will then set out a detailed justification and indicate the line of further management according to need. The same will be informed to the sponsor and the Ethics Committee within 2 working days.

Study intervention

The study intervention has been designed to be given in two groups. The first group will receive MDA plus a coded Ayurveda drug AYUSH-SL in tablet form (each tablet containing 500 mg of the active drug and 2 tablets twice a day will be given after breakfast/food, at morning and evening with water). The other group will receive MDA plus matching placebo. The duration of therapy will be of 12 weeks, and follow-up assessment will be done at the 24th week. The trial drug AYUSH-SL/matching placebo will be supplied by CCRAS, New Delhi and procured from GMP approved pharmacy.

Outcome

The participants will be evaluated for clinical assessment based on the following outcome measures:

Primary outcome measure

The primary outcome measure includes as follows:

• Reduction of edema (at baseline, 4th week, 12th week, and 24th week)*
- ˚ Lower limb measurements (in cm) (using a standard measuring tape 150 cm)
- ▪ Limb: from anterior superior iliac spine to ground at 90°
- ▪ Foot: from medial side (midpoint of heel to the tip of great toe)
- ▪ Foot: from lateral side (from midpoint of heel around lateral border of foot touching the tips of toes to the tip of great toe)
- ▪ Maximum circumference of calf (on standing)
- ▪ Maximum circumference of mid-thigh (on standing)
- ▪ Maximum circumference of patellar (at mid-level)
- ▪ Circumference of ankle (joining the medial and lateral malleolar protuberances)
- ▪ Circumference of mid-foot (joining the mid-point of the arch on the medial margin of the foot and the base of the fifth metatarsal)
- ▪ Circumference of foot joining the heads of the metatarsals
- ˚ Volume displacement measurement (in mL)
- ▪ Water displacement in a filled bucket (by dipping the limb)

*Percentage reduction from the baseline value will be computed for analysis.

Secondary outcome measure

The secondary outcome measures are improvement in QoL assessed using Lymphatic Filariasis Specific Quality of Life Questionnaire having domains such as mobility, self-care, usual activities, disease burden, pain/discomfort, psychological health and social participation, and prevention of recurrence of acute episodes through presence and severity of episodes in the past 4 weeks (at baseline, 4th week, 12th week, 24th week).

Participant timeline

Patients’ information including medication history will be obtained from the health center and the treating physician following getting their consent for the same. After obtaining informed consent, only those patients who fulfilled inclusive criteria and also agreed to participate will be screened based on laboratory investigations. The investigation includes immunochromatographic (ICT) filariasis test, urine routine and microscopic examination (specific for chyluria), hemogram (including total count (TC), differential count (DC), erythrocyte sedimentation rate (ESR), haemoglobin (Hb) %), absolute eosinophil count, liver function test (LFT) (including serum glutamic-oxalacetic transaminase (SGOT), serum glutamic-pyruvic transaminase (SGPT), serum bilirubin, plasma protein, albumin, total protein, serum alkaline phosphatase), kidney function test (KFT) (serum creatinine and urea), lipid profile (serum cholesterol, serum triglycerides, low density lipoprotein (LDL), high density lipoprotein (HDL), and very low density lipoprotein (VLDL)), blood sugar (random) and glycosylated Hb, thyroid profile, and electrocardiography.

Enrollment of participants will be done at Filaria Clinics of Ayurveda Hospitals under supervision of an Ayurvedic physician. [In Kolkata, patients would be recruited at Filaria Clinic/Clinical Pharmacology Clinic, CSTM, Kolkata, India.]. The reason for drop-outs, if any, will be recorded during the trial.

Period of treatment and subject compliance monitoring

Initially, a washout period of 2 weeks would be given to the participants who are already receiving any medicines for treatment apart from MDA. Participants compliance will be monitored by the Drug Compliance Report Form through regular follow-up of the participants. The visits of participants would be at 0 day (baseline), 1 week, 4th week, and 12th week, and post-treatment follow-up will be done at 24th week.

At baseline, demographics details and medical history will be recorded. Clinical examination and laboratory investigations will also be done. Assessment through Ayurveda parameters, assessment of LF, QoL questionnaire, measurement of lymphedema by limb circumference measurement, and issuance of drugs will be noted. At 7th day, clinical examination and laboratory investigations will be done. Concomitant medication, rescue medication, assessment of adverse drug reactions/adverse events, issuance of drugs, and filling up of drug compliance report form will be carried out during the visit on 7th day. Similar examinations will be done and observations will be noted at 4th and 12th week [Figure 1].

Figure 1: Study flowchart

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Follow-up check-up without treatment

Follow-up check-up without treatment will be carried out at the end of 24th week. All investigations which will be done during initial examination will be again performed during the follow-up period [Table 1].

Table 1: Study schedule

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Safety assessment

The participants will be monitored for clinical safety, and laboratory investigations will be carried out at each follow-up [Table 2]. Adverse event, if any observed during treatment or follow-up, will be documented and it will be managed accordingly. The same will be reported to the Institutional Ethics Committee (IEC) and the sponsor(s) by the investigator at the earliest.

Table 2: Assessment parameters

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Sample size

For this study, purposive sampling is selected as the disease burden is more in discrete areas. Participants will be selected from the endemic areas from the districts in the neighborhood of the study centers.

The total sample size (n) is 180, with 90 patients to be enrolled in each group considering a difference of 1.2 cm in limb circumference after treatment between the two groups to be clinically relevant, standard deviation of 2.5 cm based on the results of the previous studies, 95% confidence level (α = 0.05), and 80% power. This includes the margin for attrition due to drop-outs at the rate of 25%. There would, however, be competitive enrollment at the three participating centers.

Allocation and blinding procedure

Participants will be randomly allocated in the study with an allocation ratio of 1:1. To avoid recurrence of enrollment of the same participant, each participant will be assigned with a unique number to identify them. By using statistical methods, a neutral person (statistician) will make a key for the two arms. According to the key, the individual drug/placebo pack/trial drug will be prepared and coded on the drug pack. Individual treatment code will be provided by CCRAS with indication of the treatment allocation for each randomized patient. One set of envelopes will be held by CCRAS and the investigator will keep a second set at the hospital in a safe accessible place.

Data collection

All the information pertaining to the clinical trial will be appropriately handled and carefully documented as well as meticulously stored at the study centers for precise interpretations and verifications. The filled E-format will be sent to the CCRAS headquarters on a regular basis.

Training of project personnel

The investigator of the participating centers would be trained to ensure standardized methods of data collection.

Data management

The records and data gathered during the study will be saved in password-protected folders on a secure computer and also in external data storage device. Only the investigator will have access to the saved data.

Statistical analysis plan

Data obtained after complete follow-up of participants will be pooled and analyzed statistically through Statistical Package for Social Sciences (SPSS) 15.0 version using appropriate statistical methods. Analysis of the data will be done by Council’s Statistical Section. A Joint Technical report will be published by CCRAS and manuscript will be published in peer-reviewed index journal.

Study monitoring

On completion of the trial period of 12 weeks by at least 25% of the participants, the interim analysis will be done as per requirement. Progress of the trial will be monitored by the project monitoring committee through review meeting and site visits as per requirement to ensure strict adherence to the trial protocol and correct completion of the forms and to discuss any problems being faced by the research staff at the participating site.

Drug management and accountability

The study drug will be prepared at GMP-Certified Pharmaceutical Company, following guidelines laid down in Ayurvedic Pharmacopeia of India. Proper labeling including manufacturing date, expiry date, etc. will be ensured. Testing for safety parameters such as heavy metals, aflatoxin, and pesticide residues will be carried out. The study drug will be kept in a secure place and will be supplied to the patients under the observation and responsibility of the investigators. A record of the study drugs dispensed will be kept to avoid any discrepancies between amounts dispensed and remaining/returned.

Drug compliance

Treatment cards will be issued to the registered participants wherein the entire treatment regimen will be written. The participants having equal to or more than 80% compliance will be continued in the study. The compliance will be assessed at every visit by assessing the approximate number of capsules intake by the participants and the returned empty capsules foils.

Prior and concomitant medication

The participants will also be instructed not to take any drugs other than the trial drugs for any disease. The investigator will instruct the participants to consult them for any other signs and symptoms or if anything unusual occurred. Any other medication(s) taken by the participants will be recorded by the investigator.

Rescue medication/quick-acting medication/fast-acting medication

The use of rescue medication for alleviating any emergency condition as per the wisdom/discretion of the investigator has been permitted in the study. It will be documented in detail in the case report form.

Ethics and dissemination

The trial will be conducted in accordance with ethical principles that have their origin from the Declaration of Helsinki for biomedical research and ICMR ethical guidelines involving human participants, Good Clinical Practice (GCP) guidelines for Ayurveda, Siddha, and Unani drugs. The trial protocol and participant information sheet along with consent form will be submitted to the IEC before initiation of the trial, and written approval of the same will be obtained. Approval for protocol amendments (if any) will also be obtained from IEC as per usual procedure.

Protocol amendment

The protocol of the trial will be followed and deviations, if any, will be made only if alleviation of an immediate hazard to trial participants is required. All the deviations from the protocol such as any changes to interventions, examinations, data collection, and method of analysis will be reported to sponsors and IEC with statements of exact reason for the deviation at the earliest.

Participant information and consent form

The investigator will obtain a voluntarily signed and witnessed informed consent from all the participants before initiating the trial. The participants will be informed about the trial in verbal form and written in easily understandable and readable form before initiating the trial. The participant will be informed about the aims and objectives, methodology, any discomfort, and expected benefit of the trial. The participation in the trial will be voluntary, and the participants can leave the trial at any time as per their wish.

Discussion

The WHO encourages the countries to take measure for eliminating LF. India alone accounts for 40% of the world’s burden of LF.^[21] Lymphedema due to LF is the second major cause of disability globally, which accounts for approximately 2 million disability-adjusted life years (DALYs) lost.^[4] Lymphedema when advances then becomes irreversible due to initiation of dermis fibrosis and certain reactive changes occurring in the subcutis and epidermis. Management of morbidity and disability prevention requires broad strategies involving secondary and tertiary prevention measures. The secondary prevention includes basic hygiene such as skin care, exercise, prevention of acute attacks, and advancement of lymphedema to elephantiasis, whereas tertiary prevention includes psychological as well as socioeconomic support to the people with disability.^[14] People suffering with lymphedema require care throughout life for preventing the advancement of the disease to severe stages and management of the disease. The treatment accessible for lymphedema patients is presently quite arduous in comparison to the satisfactory response due to which patients show less concordance to the multifaceted schedules. New drugs which will enable reversal of lymphedema may be beneficial and essential in view of Sustainable Development Goal 3.8, which aims for safe and effective affordable essential health care for all.^[4]

Ayurveda medicines are immerging as a hopeful measure not only to cure range of diseases but also as preventive medicine which can improve QoL. Ayurveda texts such as Bhaishajya Ratnavali, Vrindamadhava, Yogaratnakara, Bhavaprakasha have indicated many herbal and herbo-mineral drugs and formulations for the management of LF.^{[22],[23],[24]} Some previous researches in acute cases of filariasis and in microfilaremia have proved the effect of coded Ayurveda drug AYUSH-64.^[25] According to another study, integrative treatment combining Ayurveda, Yoga, and allopathic medicines significantly improved the QoL of lymphedema patients. The treatment was quite effective in all grades of lymphedema of any duration of illness.^[26]

LF patients due to associated disfigurations and disability have to face social stigmas that result in economic and psychological stresses.^[27] Chronic clinical manifestations as seen in LF-infected patients such as lymphedema, elephantiasis, and hydrocele are not treated by MDA.^[28] Effective management of lymphedema is the need of the hour for preventing disease progression and episodes of ADL. The clinical trial designed with the objectives to determine the efficacy of coded Ayurveda drug AYUSH-SL in reducing LF in patients receiving MDA treatment, improvement in QoL, and percentage decrease in recurrence of acute episodes along with clinical safety may generate the evidences that will validate Ayurveda treatment approaches for the management of the disease condition.

Trial status

The trial has been initiated.

Acknowledgments

The authors extend their gratitude to Director General CCRAS, Ministry of Ayush, Government of India for providing financial support and approval for conducting the trial. The authors also thank the study participants for their active participation in the trial.

Financial support and sponsorship

Financial support for the study is under Intra-mural Collaborative Research Scheme of Central Council for Research in Ayurvedic Sciences (an autonomous organization under Ministry of Ayush, Government of India).

Conflicts of interest

There are no conflicts of interest.

References

1.	Global programme to eliminate lymphatic filariasis: Progress report. 2018; WHO. Available from: https://www.who.int/publications/i/item/who-wer9441-457–472. Last accessed on March 2020.
2.	Lymphatic filariasis. WHO. Available from: https://www.who.int/news-room/fact-sheets/detail/lymphatic-filariasis. Last accessed on March 2020.
3.	Lourens GB, Ferrell DK. Lymphatic filariasis. Nurs Clin North Am 2019;54:181-92.
4.	Specht S, Suma TK, Pedrique B, Hoerauf A. Elimination of lymphatic filariasis in South East Asia. BMJ 2019;364:k5198.
5.	Morbidity Management and Disability Prevention (MMDP). WHO. Available from: https://www.who.int/activities/building-capacity-of-national-programmes-to-implement-who-recommended-strategies/morbidity-management-and-disability-prevention. Last accessed on March 2020.
6.	Elimination of Lymphatic Filariasis, National Centre for Vector Borne Diseases Control, Directorate General of Health Services, Ministry of Health and Family Welfare, Government of India. Available from: https://nvbdcp.gov.in/index4.php?lang=1&level=0&linkid=405&lid=3681. Last accessed on March 2020.
7.	Lymphatic Filariasis (LF), National Vector Borne Disease Control Programme. Ministry of Health. Available from: https://nvbdcp.gov.in/index4.php?lang=1&level=0&linkid=451&lid=3728. Last accessed on March 2020.
8.	Global programme to eliminate lymphatic filariasis: Progress report. 2019. WHO. Available from: https://www.who.int/publications/i/item/who-wer9543. Last accessed on March 2020.
9.	National Vector Borne Disease Control Programme. Available from: https://nvbdcp.gov.in/index1.php?lang=1&level=1&sublinkid=5851&lid=3686. Last accessed on March 2020.
10.	Accelerated Plan for Elimination of Lymphatic Filariasis. 2018. Available from: https://nvbdcp.gov.in/WriteReadData/l892s/1031567531528881007.pdf. Last accessed on March 2020.
11.	Vagbhatta, Asthanga Hradayam Vidvanmnoranjini Hindi Commentary by Pt. Kashinath Shastri. 1st ed. Varanasi: Krishnadas Acadamy; 1994. p. 559.
12.	Agnivesha, Charak Samhita, Charak Chandrika Hindi Commentary by Brahmanand Tripathi, Reprint ed. Varanasi: Chaukhambha Subharati Prakashan; 2008. p. 467.
13.	Sushruta . Sushruta Samhita Nibandha Sangraha, Dalahan Commentary by Acharya Priyavat Sharma, Reprint ed. Varanasi: Chaukhambha Orientalia; 2009. p. 317-18.
14.	Sushruta . Sushruta Samhita, Nibandha Sangraha, Dalahan Commentary by Acharya Priyavat Sharma, Reprint ed. Varanasi: Chaukhambha Orientalia; 2009. p. 477-8.
15.	Madhavakar . Madhava Nidana, Madhukosha Commentary by Brahmanand Tripathi, Reprint ed. Varanasi: Chaukhambha Subharati Prakashan; 2007. p. 38-9.
16.	Bhavaprakash B. Sanskrit English Translation by Sri. Kanthamurthy K.R. Varanasi: Krishnadas Academy; 2002. p. 546-7.
17.	Hussain MA, Sitha AK, Swain S, Kadam S, Pati S. Mass drug administration for lymphatic filariasis elimination in a coastal state of India: A study on barriers to coverage and compliance. Infect Dis Poverty 2014;3:31.
18.	Anonymous. CCRAS Annual Report 2004–05. New Delhi: Ministry of Ayush, Government of India. ; 2005 p. 63.
19.	Anonymous. CCRAS Annual Report 2005–06. New Delhi: Ministry of Ayush, Government of India. ; 2006 p. 66.
20.	Anonymous. CCRAS Annual Report 2006–07. New Delhi: Ministry of Ayush, Government of India. ; 2007 p. 64.
21.	Neglected tropical diseases: Lymphatic filariasis. WHO SEARO. Available from: http://www.searo.who.int/entity/vector_borne_tropical_diseases/topics/lymphatic_filariasis/en. Last accessed on March 2020.
22.	Shri G. Bhaishajyaratnavali. In: Choudhary Anand K, editor. Shlipada Chikitsa. Vol. 2. Varanasi: Chowkhambha Sanskruta Bhavan Publication; 2006. p. 729-30. 733-4.
23.	Anonymous. Yogaratnakara, Chetty M, Babu S, editors. Shlipada Chikitsa. Vol. 2. Varanasi: Chowkhambha Sanskrit Series Publication; 2008. p. 878.
24.	Anonymous. Vrundamadhava or Siddha Yoga. Tevari Premvati, editor. Varanasi: Chowkhambha Visva Bharathi Academy Publication; 2006. p. 563-4.
25.	Naidu ML. Effect of Kuberaksha Patra Churna, Vriddhadaru Mula Churna and Kandughna Taila in Shlipada (manifested filariasis). Ayu 2012;33:102-9. [PUBMED] [Full text]
26.	Narahari SR, Bose KS, Aggithaya MG, Swamy GK, Ryan TJ, Unnikrishnan B, et al. Community level morbidity control of lymphoedema using self care and integrative treatment in two lymphatic filariasis endemic districts of South India: A non randomized interventional study. Trans R Soc Trop Med Hyg 2013;107:566-77.
27.	Kulkarni P, Thomas JJ, Dowerah J, Murthy MRN, Ravikumar K. Mass Drug Administration Programme against lymphatic filariasis—An evaluation of coverage and compliance in a Northern Karnataka district, India. Clin Epidemiol Glob Health 2020;87-90.
28.	Walsh V, Little K, Wiegand R, Rout J, Fox LM. Evaluating the burden of lymphedema due to lymphatic filariasis in 2005 in Khurda district, Odisha state, India. PLOS Negl Trop Dis 2016;10:e0004917.