An ethnopharmacological survey and comparative analysis of plants from the Sudhnoti District, Azad Jammu and Kashmir, Pakistan

Background This is the first comprehensive report on the traditional and novel uses of medicinal plants practiced by the indigenous communities of the Sudhnoti district of Azad Jammu and Kashmir (AJK), Pakistan. The area is rich in folklore and indigenous medicinal knowledge due to a unique tribal composition and socioeconomic conditions. This study aimed to document traditional knowledge of native plant use by the local communities, particularly those used for therapeutic purposes. Methods Field surveys were conducted from September 2015 to March 2017. Interviews with 125 local inhabitants of different tribes, age groups, genders, and occupations were conducted using structured and semi-structured questions along with group discussions. Data gathered on plant uses, local names, and modes of application of each plant species were organized in tables. Ethnobotanical indices such as use value (UV) and cultural significance index (CSI) were used to produce quantitative information on the plant use category, frequency, and cultural preference of species. Reports on therapeutic uses of medicinal plants were compared with previous studies. Results In all, 88 plant species from 45 families were reported, out of which 67 (77%) were used in ethnomedical applications. Asteraceae, Rosaceae, Fabaceae, and Lamiaceae were the dominant families. Berberis lycium was the most valued plant species, followed by Zanthoxylum armatum and Taraxacum officinale. Mentha arvensis had the highest cultural significance, followed by Mentha longifolia, Punica granatum, and Zanthoxylum armatum. Leaves were the most preferred plant parts in the preparation of medicine exclusively or mixed with other parts. The most frequently used process of crude preparation of medicinal plants was cooking. Oral intake was the predominant route of administration. Conclusions Our comparative analysis confirmed that most of the plants documented have uses that match those previously reported for the region and other parts of the world, with the exception of novel medicinal uses for 11 plant species, including Verbascum thapsus for earache, Elaeagnus umbellata for hepatitis, Achillea millefolium for oral care, Dicliptera roxburghiana to prevent sunstroke in cattle, Rumex hastatus for allergy antidote, Pyrus pashia for hepatitis, and Nerium oleander for diabetes. Supplementary Information The online version contains supplementary material available at 10.1186/s13002-021-00435-2.

use descriptions and comparative accounts can help rationalize any possible complexity arising from indices [7]. According to a study conducted in the Sudhnoti District, local communities of the area are highly dependent on plants for their therapeutic uses; therefore, documentation of their medicinal knowledge, as well as pharmacological validation thereof, is highly encouraged [8].
Although ethnobotanical studies have been conducted on a large scale in the surrounding areas of AJK [9][10][11][12], ethnomedicinal knowledge in the Sudhnoti District remains poorly studied. To the best of our knowledge, a single attempt was made that focused primarily on medicinal plants without acknowledging novel therapeutic uses of medicinal plants [9]. The aim of our study was two-fold: (1) to document the ethnobotanical knowledge of the area and (2) to compare it with other studies conducted in the surrounding regions and other parts of the world at large in efforts to identify novel medicinal plant uses that could merit further pharmacological evaluation.

Study area
The Sudhnoti District, AJ&K, study area lies in coordinates ranging from 33°.40′-33°50′ N latitude and 73.40-73°50′ E longitude. The altitude range is approximately 600 to 2100 m.a.s.l. (Fig. 1). The study area is comprised of hills and mountains along with small valleys and plains, spanning a total area of about 5695 km 2 . During the summer and winter seasons, the temperature ranges between 20 to 35°C and 5°C and 20°C, respectively.

Ethnographic composition
The study area has a distinct tribal composition with a majority of the population belonging to Sudhan tribe, followed by Awan, Syed, Gujjar, Qureshi, and Kashmiri ethnic groups. Nearly, the entire population lives in a rural setting with a strong religious influence. Hence, gender discrepancies still exist in the health and education sectors and the acquisition of traditional knowledge. When researchers approach a typical rural household, the elder male is most likely the source of traditional knowledge, especially when the researcher is male. In small towns, including the district headquarters, there is an appreciable number of educated youths possessing common knowledge of plant uses. However, the bulk of traditional folklore is still conserved with the illiterate elderly adults, living in the remote highlands and riverside pastures. Besides small-scale conventional farming and livestock rearing, most of the population survives directly on biodiversity and work on daily wages for sustenance. Pahari is the most commonly spoken language in the area, with different dialects spoken. Only a few informants from the Gujjar tribe prefer to speak Gojri while discussing traditional uses of plants; the same is true for elderly women from the Kashmiri tribe. The conventional healthcare infrastructure comprises a district headquarter hospital with a dozen doctors and less than ten basic health units in the entire district. On average, there is one physician per 7000 citizens in the district. Access to physicians is even more restricted for the people living in remote areas as there is a very underdeveloped road network in the study area.

Sampling and data collection
Six different sites in the Sudhnoti District were selected based on altitude and demographic composition for sampling and data collection, namely, Azad Pattan, Baral, Baloch, Palandri, Gorah, and Trarkhal (Fig. 1). Prior to data collection, a preliminary survey was conducted in which local administration and community stakeholders were engaged for prior consent. Multiple field surveys were conducted from September 2015 to March 2017. Prior informed consent was always verbally obtained before conducting interviews, and the ethical standards of the Society for Economic Botany and International Society of Ethnobiology were followed [13]. Open-ended interviews and structured questions followed field surveys and transect walks either at common places where local people gather (e.g., local bazaars) or at the residences of informants. The first author conducted all interviews in the local language (Pahari) and subsequently documented the interviews in Urdu and English. Local people with biomedical and geographical knowledge were also involved whenever required for better communication, such as understanding disease symptoms. Local healers known as "Pansari," and nomadic healers locally known as "Sunyasi," were interviewed with temporarily mounted plant samples.
During fieldwork, 88 plant specimens were collected, preserved, identified, and indexed. A total of 121 informants of 30-85 years old were interviewed ( Table 1). The gender division of infomants was 55% male and 45% women. Collected plant specimens were dried, pressed, and mounted on herbarium sheets. The identification of plants was carried out with the assistance of Dr. Rehmatullah Qureshi at PMAS Arid Agriculture University. The Flora of Pakistan [14] was used as a standard for identification and nomenclature. Voucher specimens were submitted to the herbarium of PMAS Arid Agriculture University, and accession numbers were acquired. Botanical nomenclature followed the most updated accepted names included (http://www. theplantlist.org) and family designations followed the Angiosperm Phylogeny Group IV assignments [15]. Digital images of the collected herbarium specimens are available in Additional file 1.

Use value (UV)
Use value, a primary measure of all reported uses of a plant species cited by an informant was calculated by the following formula where U i is the number of uses revealed by each informant for a given species and N is the total number of informants.

Cultural significance index (CSI)
CSI is an anthropological index developed by [16] and modified by [17]. CSI calculates the importance of plant taxon through a researcher-generated weight ranking of multiple factors. A two-point scale for the variables such as use frequency (2 = species effectively used, 1 = not the preferred use), management, and preference were used. To reduce the sensitivity of the method to sampling intensity, a correction factor is also incorporated as in the consensus method [17,18].
Here, "i" refers to the species management, "e" is the preference of use of a plant species by informant for a specific purpose, and "c" is the frequency of use reported.

Comparative analysis
Primary ethnobotanical uses of all plant species were compared to regional studies containing "ethnobotany," "medicinal plants," "folklore," or related search terms using different databases [19]. Dissimilar uses cited by other researchers were reviewed and cited. To avoid repetition of similar uses reported in multiple articles for a particular species, the one chosen for citation was the one conducted nearest to the immediate study region.

Novelty assessment
A total of 13 plant species were identified as being reported for novel uses from the region. These plant species were subjected to a robust comparative review using literature databases, i.e., PubMed, Scifinder, and NAPR ALERT, to develop a broader picture of their previously reported pharmacological activities and cross-examining the novelty of uses reported in the present study. Novel uses are highlighted in bold in Table 2.

Diversity of medicinal plants
During the first phase of this study, 88 plant species from 47 families were tabulated after taxonomic identification and scrutiny. Asteraceae was the dominant family with eight plant species, followed by Rosaceae (6), Fabaceae (6), and Lamiaceae (5). Other significant families were Poaceae and Acanthaceae, each with four plant species (Fig. 2). All other plant species represented 45 angiosperm families. The data show that angiosperms contribute to 97% of the collected plant species, followed by ferns (3%).

Medicinal taxa cited by different ethnic groups
The informants belonging to different ethnic origins were grouped into three major categories: Sudhan, Gujjar, and Others (Kashmiri, Syed, Quereshit, etc.). Sudhans, owing to their largest share in population, cited the highest number of plants (48), Gujjars cited 21, and others reported 16. There was also an overlap of taxa between ethnic groups: six between Sudhan and Gujjars and five between Sudhan and Others. Only four plants were commonly cited by all three groups (Fig. 3).

Ethnobotanical uses and therapeutic value
Out of the total 88 plant species, 78% of plants were used for therapeutic purposes, followed by food (18%), and wood (8%). Plant species having aesthetic or religious value are grouped together (Fig. 4). The used categories are highly overlapping, as different plant parts of certain species were reported for different uses by different community sections simultaneously.

Parts used for medicinal purposes
Among the plant species used for medicinal purposes, leaves were the most commonly used plants parts (37%), followed by aerial parts (23%), and fruits (18%). However, in the case of medicinal herbs, the use of aerial parts and the whole plant is dominant (Fig. 5). The use of bark and roots is culturally discouraged due to the local tendency toward resource conservation.

Method of preparation of folk recipes
The largest share of plant species (20%) had a reported use as cooked vegetables. However, the share of plants used as infusions (21%) and decoctions/teas (13%) together was 34% of all the species (Fig. 6). Interestingly, there is a notable trend of multi-plant formulations devised by semiprofessional herbalists and traditional practitioners. In these cases, the powder of more than one plant or plant part is orally administered with water in a formula known as "Phakki." There are some instances with such recipes where overdose and malpractice resulted in adverse drug reactions [66].

Route of administration
With the ancient mindset that human ailments result from an abnormality in the body's internal system, most of the botanical preparations are taken orally (85%). External application is restricted to pain management, bone fractures, wounds, and skin infections. A couple of plants were reported for nasal inhalation and as drops for the eyes or ears. The use of certain plant species as chewing sticks (Zanthoxylum armatum DC, Rutaceae, and Nerium oleander L., Apocynaceae) is also considered internal here for ease of classification.

Use value
A total of 20 plant species were identified as the most valuable in the area (

CSI
Cultural significance of a plant species is markedly affected by preference, management, and frequency of use by local inhabitants. Mentha arvensis, with a value of 9.9, has the highest cultural significance followed by Mentha longifolia (9.7), Berberis lyceum (9.6), Justicia adhatoda (8.8), and Zanthxylum armatum (6.7). Other species with cultural significance are Punica granatum, Artemisia absinthium, and Swertia chiraita ( Table 2).

Novelty analysis
The current study's fundamental objective is to compare the findings of the field survey and data analysis with previously reported uses from the region. Comparative analysis identified 11 plant species with specific uses reported from the area for the first time, shown in bold in Table 2. For example, Elaeagnus umbellata was reported for liver ailments, Gentiana olivieri for genital infection, and Pteris cretica for antifungal activity. Pharmacological profiling of plant species was developed with a literature review (Table 2).

Discussion
The study area has diverse vegetation where 47 plant families represented a field collection of 88 plants. This high level of biodiversity was also reported by a previous study conducted in the area [7]. Altitude variation is one of the factors affecting floral richness in the area, as the study area of a mere few thousand km 2 represents subtropical (altitude 600 to 2100 m.a.s.l.) to alpine climate conditions. There are rich traditional medicinal practices in the area where 68% of plant species were used for therapeutic purposes. This indicates the presence of a large number of medicinal plants in the local flora and diverse traditional knowledge. Major ethnic groups such as Sudhan and Gujjar prefer to treat their daily ailments with plant-based remedies. The diversity in traditional knowledge is linked to the fact that both of the major tribes have a strikingly different tribal descent where Sudhans originally migrated from Afghanstan still hold strong ties with Pashtoon folklore. On the other hand, Gujjars, originally migrated from central Asia and northeastern Europe, have a legacy of a nomadic lifestyle and remain settled in alpine Himalayan pastures since hundreds of years. Moreover, District Sudhnoti is one of the most remote districts of Azad Kashmir, having an inferior transport system and road infrastructure. This disadvantage in communication has a conservatory impact on traditional knowledge as an influx of people with other cultural backgrounds is occurs only occasionally. The majority of the study area population lives in far flung rural areas and manage primary health care needs from traditional means. As easy access to hospitals and allopathic medicines is restricted to a mere few thousand people living around the district headquarters of Sudhnoti and a couple of small towns.
Another reason for the strong belief of the local populace on plant-based remedies is due to the influence of traditional healers, who are often adept in the spiritual component of healing. Generally, healers are wellequipped with the concept of Dam Darood (insufflation) and the mythical practices and prayers accompanying adminstration of herbal formulations. Consequently, they prescribe herbal remedies aided by insufflation accompanied by a period of ardent prayers for that particular ailment. For example, elderly women breathe on the roots of Gentiana oleiveri and advise it to be used for children with genital infections.
Locals use some plant species due to their medicinal or nutraceutical significance mentioned in Holy scriptures such as Quran and Hadith. For example, Ficus is used as an expectorant and Ziziphus as a pain reliever because they are mentioned in Holy texts as sacred plants that can cure a range of human ailments. The combination of traditional and spirtual healing is also reported from other parts of the world as well [67,68].
This study identifies leaves as the most widely used plant part for medicinal purposes. Our findings are in agreement with surveys conducted previously in surrounding areas in the region [32]. The reason for the extensive use of leaves could be their ease of access and having reserves of active metabolites [69,70]. Here, we found that the preparation of plant-based remedies in infusions and decoctions is the most common form (37%). Technically, an infusion is different from a decoction in a sense that the former is a liquid preparation in which boiling water is poured on the plant material while the latter is the boiling of plant material in the water [71]. Our findings are supported by previous studies conducted in the Poonch region [32].
Use-value is a basic index that accounts for all possible uses of a plant species. A plant with a high use-value index is often more commonly found and frequently used; examples include Berberis lycium, Zanthoxylum armatum, and Taraxacum officinale. These findings are generally in agreement with previous studies conducted in the are. For example, Berberis lycium has previously been reported as the most valued plant species in the area [8]. Herein, we confirm these findings. B. lycium was also reported as a high use value plant from Kel village of Neelum Valley, Azad Kashmir, Pakistan, where the local communities used a paste and poultice made from this plant for the treatment of jaundice and diarrhea [72]. Higher use-values often contradict the novelty of plant uses, as the most commonly found plant is more frequently cited and valued by a large number of informants than scarcely found novel plants.
Our methodology discouraged the use of reports on plant species involving agronomic or commercial usevalue; this preserved the indices reflection of traditional ethnobotanical knowledge. The cultural significance of a plant considers preference for it, use frequency, and management [18]. Mentha arvensis has the highest CSI value despite having fewer citations due to restricted occurrence along the waterways of high temperate areas. Indeed, local people prefer to use and manage Mentha arvensis for medicinal and food purposes.
In recent years, plant use information has been compared by using statistical tools such as the Jaccard index. Relevance of these tools is, however, a topic of debate for ethnobotanical studies. It was noted that if plant use information documented from a particular area is subjected to a robust review and comparison with previously published reports on that taxon, its novelty is largely established [73].
The present study reports novel medicinal uses of 11 plant species (Table 2, in bold). These plant species were subjected to a robust pharmacological review from other parts of the world (Table 3). In our study, for example, A. millefolium is reported for gum soreness and kidney stones. The pharmacological review of this plant shows its application as an antispasmodic, diuretic, urinary antiseptic, and anticancer therapy [74]. Dicliptera roxburghiana is given to cattle for the amelioration of "Takko," an ailment characterized by severe thirst, dehydration, and lethargy. One of the causes of "Takko" as per the general perception of Gujjar (nomadic) tribesmen is the prolonged exposure of cattle to sunlight. Another novel use is the oral intake of Elaegnus umbellata fruit for hepatitis B. Informants in our study considered hepatic disorder as a sign of heating up of the liver and stomach. Therefore, they take fruits of E. umbellata as a cooling agent that potentially has some antiinflammatory or antiviral effects. The plant is otherwise known for pharmacological activities such as antimicrobial [75], phytotoxic [76], antioxidant, and antiproliferative effects [77]. Ethnomedicinally, it has been reported to treat cardiac problems, bacterial infections, and as an astringent in Malaysia and Pakistan [39].
Gentiana olivieri is another plant reported here for a novel use that reduces genital infections and inflammation in male children. The symptoms of this condition include inflammation of the testicles followed by itching and soreness. Fresh flowers are directly squeezed into the mouth in the form of drops as an oral dosage. Additionally, the dried root segments are enveloped in a piece of cloth and advised to be worn around the neck for spiritual healing effects. Previously, it has been reported for hepatoprotective activities in Turkish folk medicine [96]. Pharmacological applications against epilepsy [79], for radical scavenging [80], and as immunomodulatory and hepatoprotective agent [81] have also been reported.
Nerium oleander, a well-known plant for its poisonous effects due to the presence of cardiac glycosides, was reported for diabetes and oral hygiene. In the traditional treatment of diabetes, leaf powder is mixed with the powder of Stewia species to neutralize its toxicity and improve the taste. A young shoot is used as a chewing stick (Miswak) to cure gum infections. Previously, N. oleander has been reported for stomach and earache [22]. Anticancer and antimicrobial activities have been reported for this species [88].
Pteris cretica, a fern, is locally used to clean the milk utensils. It is considered useful for reducing foul odors, but it is also believed that the plant species is a cleansing agent purging milk utensils in a religious context. It is important to note that milk (particularly cow's milk) is considered a holy food item in Hindu culture. Hindus have long inhabited the study area, and therefore, imprints of traditional Indian culture are still evident. Previously, this species has been reported for wound healing applications [10], and it exhibits antimicrobial, antioxidant [89], anti-inflammatory, and anticancer activities [97].
Pyrus pashia is one of many wild fruit yielding species of Rosaceae family. Young apices (with leaflets) of the plant are ground, and the aqueous extract is taken orally for hepatitis B and C, colloquially known as "black jaundice." In previous studies, the plant was reported for the treatment of baldness [10]. In vitro bioactivities, such as antimicrobial and antioxidant, have also been reported [89].
Sarcocca saligna locally known as "naidroon" is another species reported here for applications in treating toothache and fevers. Roots of the plant are crushed in water, and the filtered extract is used in the form of drops onto the affected teeth. In fever, particularly one that of enteric origin (Typhoid), the extract is diluted and taken orally. Previously the plant has been reported for treating hypertension [36]. Pharmacological reports include antimicrobial, antispasmodic [90], cardio suppressant, vasodilatory [91], and hyperglycemic effects [98]. A leaf infusion of Rumax hastatus is reported here for liver ailments, particularly jaundice, owing to its cooling effect. Local healers consider jaundice a sign of an "internal heating" and a forthcoming indication of chronic hepatic disorders. Therefore, R. hastatus is taken as a chilling agent that "cools down" the body and helps the liver to alleviate the damage. The plant is also reported as an antidote for allergic reactions to weeds such as Parthenium hysterophorus. Here, fresh leaves of R. hastatus are reported to be rubbed on the affected body parts. R. hastatus has been previously reported for antirheumatic activity, as a tonic, laxative [55], and as an antioxidant, antitumor, and antimicrobial agent [56,57].
Leaves of Verbascum thapsus are reported here for healing physical trauma; briefly, the fresh leaf is coated with salt sprinkles and wrapped around the injury site to as an analgesic and to heal contusions. V. thapsus is commonly known as a wild form of tobacco and smoked at times with a recreational sense. It has also been reported in the region for asthma, sore throat, and inflammation [65]. It is also reported to have anti-lice and flea activity and used for earache [99]. Pharmacological evaluation has consistently established antiviral activities of V. thapsus [92][93][94]. The plant is also reported for anticancer activity [95].
On the other hand, uses reported from other parts of the world are similar to what we report here, such as the antimalarial use of Melia azedarach [45,46], indicating a global consensus. This phenomenon allows for different cultures to learn from each other about further medicinal uses of their plants and reveals to researchers more bioactivities that some phytochemicals may exerte. Lastly, we conclude that the new traditional ethnopharmacological applications reported here for 11 plant species should be pursued within the framework of ethnobotanical drug discovery. Significant antibacterial activity of plant species including Z. armatum, A. capillus venaris, M. annua, and A. absintium has already been reported by our research group [100].

Conclusions
This is the first detailed ethnobotanical study conducted in District Sudhnoti of Azad Kashmir. A total of 88 plants belonging to 47 families were reported, verifying the high biodiversity of medicinal species in Sudhnoti and, therefore, the need to preserve it. Moreover, 11 plant species were found to have novel uses not reported elsewhere. Information concerning the ethnomedicinal uses of plants, specific plant parts, and application methods is critical to both the preservation of traditional knowledge and provides a basis for future drug discovery activities. In this context, the current study enriches our knowledge basis of the medicinal plant potential and traditional knowledge of District Sudhnoti. Future ethnobotanical and phytochemical research will be fundamental to edxploring the pharmacological potential of these and other botanical traditions.