Wetland biodiversity of Ramaroshan Lake complex: a need for conservation

: The Ramaroshan Lake Complex, situated in the mid-hills of Sudurpaschim Province, is renowned for its scenic beauty, yet there is a notable dearth of information regarding its biodiversity and ecological status. This study represents the first systematic examination of seasonal variations in water quality parameters and biodiversity encompassing aquatic macroinvertebrates, fishes, birds, herpetofauna, mammals, and macrophytes, as well as the surrounding vegetation within the complex, spanning the winters and summers of 2018 and 2019. Among the twenty water quality parameters investigated, thirteen displayed significant seasonal differences across the lakes (p <0.05), with Batula and Ramaroshan lakes exhibiting elevated nutrient levels. Lamadaya Lake stood out with a highly diverse macroinvertebrate community compared to other lakes, while overall, the study recorded 45 aquatic macroinvertebrate families, three fish species, 79 bird species, 12 herpetofauna species, 12 mammal species, and 26 macrophyte species within the complex. Additionally, the surrounding vegetation comprised 193 distinct plant species. Notably, the complex currently hosts 14 IUCN Red List species, including Near Threatened (5), Vulnerable (5), Critically Endangered (1), and Endangered (3) species, as well as five migratory wetland bird species, underscoring its significance for wildlife conservation. Given the diverse and cross-cutting nature of wetlands, the development of science-based policies and coordinated efforts among central, provincial, and local governments are essential for the preservation and sustainable management of these vital ecosystems.


INTRODUCTION
Over 5% of Nepal's land surface area is covered by natural and man-made wetlands, of which nearly 97% are contributed by rivers and irrigated paddy fields, and only 3% of the wetlands belong to marshy lands and lakes, including reservoirs and ponds (DoFD 2012).These lakes are disproportionally distributed across the varying altitudes: 51% of the lakes are situated in the high mountains above 3,000 m, 42% are located in the Tarai below 500 m, and only 7% of lakes are located in the mid-hills between 500 m and 5,000 m (Bhuju et al. 2010).Wetland ecosystems provide critical habitats to a wide range of wildlife, support livelihoods, regulate ecosystem functions, and are a source of renewable energy (Zedler & Kercher 2005;Baral 2009;Shah et al. 2011;Lamsal et al. 2014;Regmi et al. 2021a;Shrestha et al. 2021).These wetlands support critical habitats for globally threatened species (BLI 2010).Many endemic species, including two mammals (ASM 2018), one bird (BLI 2020), 10 reptiles (Uetz et al. 2018), 11 amphibians (Web 2018), 15 fish (Eschmeyer 2015), and eight flowering species, are endemic to the wetlands of Nepal.Similarly, the majority of the wetlands have socioeconomic and cultural values, and riparian communities are highly dependent on wetland products (Khatri et al. 2010;Lamsal et al. 2014).
Due to the high significance of wetlands for wildlife and society, they need to be preserved and maintained.The Convention on Wetlands of International Importance is an international treaty that was signed in the city of Ramsar, Iran, on 02 February 1971, for protecting and maintaining the wetlands of ecological, botanical, limnological, hydrological, and zoological significance across the globe and designate as Ramsar Sites.The Koshi Tappu wetland was the first Ramsar Site in Nepal, designated in 1979, due to its importance for migratory birds.Since then, the government of Nepal has successfully designated a total of 10 wetlands of international importance.Among the 10 Ramsar Sites, eight are situated either in the high mountains or lowland Tarai, while only two are located in the midhills though the region covers over 40% of the total land surface.
Most of the wetlands, particularly in lowland Tarai, are highly threatened due to the high dependency of people on wetland products to sustain their livelihood (Sah & Heinen 2001), while wetlands situated in the inaccessible areas of mid-hills and high-mountain areas are nearly free from human pressures, hence serving as biodiversity reservoirs for many native and/or endemic species.
Ramaroshan Lake complex, located in a unique geographic location in the mid-hills of Sudurpaschim Province of Nepal, may serve as a critical habitat for wide ranges of wildlife (DoF 2017).The lake complex is one of the major habitats of Nepal's national bird, the Himalayan Monal Lophophorus impejanus, and a new record of a breeding site of a wetland-dependent migratory species, the Mallard Anas platyrhynchos (Aditiya Pal pers.comm.June 2019).The inlets and outlets of the lake complex are also important habitats for a globally 'Near Threatened' species, Epiophlebia laidlawi (Nesemann et al. 2011;Shah et al. 2012;Deep Narayan Shah pers. comm. June 2019).Moreover, the lake complex is the source of the Kailash River, which sustains hundreds of thousands of downstream communities in the province.Many river systems of the province have been recently explored for their biodiversity across disturbance scales (Shah et al. 2020a), spatial scale along the longitudinal gradient (Shah et al. 2020b), stressor types (Sharma & Shah 2020), and microhabitats (Bhandari et al. 2018), but the lake complex has not yet been studied from the wider aspects of wetland biodiversity except for water quality and bathymetry (Chalaune et al. 2020).Therefore, a detailed scientific study of the wetland complex was felt necessary.The present study was carried out to assess and document the water quality and the extent and distribution of wetland floral and faunal diversity in the lake complex.

Study area
The study was carried out in the four lakes of the Ramaroshan Lake complex, i.e., Ramaroshan, Batula, Jingale, and Lamadaya lakes (Figure 1; Image 1).The lake complex lies in Ramaroshan Rural Municipality in the Achham District of Sudurpaschim Province in Nepal.The rural municipality has 4,832 total households with a total population of 23,600, including 11,092 males and 12,508 females, respectively (CBS 2021).Ramaroshan is a proposed protected forest in Nepal that covers an area of 3051.29 ha for the conservation of its unique wetland ecosystem and biodiversity (DoF 2017).The ecosystems of the Ramaroshan protected forest consist of dense forest (96.95%), grassland (1.50%), lakes (1.09%), and rivers and streams (0.46%) (DFO 2019).The lake complex is the union of 12 lakes that cover an area of 30 ha (1.09%), but water remains throughout the year only in four lakes (Ramaroshan, Batula, Lamadaya, and Jingale).

Sampling sites
The sampling sites were distributed in all the study lakes namely Ramaroshan, Batula, Jingale, and Lamadaya (Table 1).Jingale is the largest lake among the four lakes studied.In each lake, three littoral sections were selected for the sampling of aquatic macroinvertebrates and measurements of water quality parameters.The study was conducted during the winter (November-February) and summer (May--June) seasons of 2018 and 2019.)

Water quality parameters
Water quality parameters such as pH, water temperature, turbidity, dissolved oxygen (DO), electrical conductivity (EC), and total dissolved solids (TDS) were measured at three sites in each lake using a Hanna multi-parameter probe (Model: HI9829) and turbidity meter.Composite water samples were collected for the determination of total hardness, calcium hardness, magnesium hardness, total alkalinity, chloride, free carbon dioxide (CO 2 ), calcium cations (Ca 2+ ), magnesium cations (Mg 2+ ), sodium cations (Na + ), sulphate anions (SO 4

Aquatic macroinvertebrates
Aquatic macroinvertebrates were sampled from littoral sections of the lakes following Shah et al. (2015).The samples were collected from three littoral sections of each lake studied.In total, 15 macroinvertebrate samples, including one sample each from the inlet and outlet of the lake complex were collected during field visits.The samples were taken using a standard circular metallic framed hand net of mesh size 500 µm and preserved on site in 95% ethanol for further laboratory

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processing.The samples were processed at the Aquatic Ecology Centre (AEC) at Kathmandu University (KU).The macroinvertebrates were identified at the family level (Nesemann et al. 2007(Nesemann et al. , 2011;;Shah et al. 2015Shah et al. , 2020)).The identified samples were preserved in 90% ethanol and stored at AEC, KU.

Fish
The passive entanglement gear technique was used for fish sampling.The fish samples were collected through gill nets placed at different parts of the lake.
Three lake sections -left bank, right bank, and center-were selected in each lake for fish sampling.At each site, two-gill nets were placed and removed every two hours.All captured individuals were taken to a nearby dry place, identified to species level (Shrestha 2019), measured, photographed, and then released back into their original habitats.Specimens that could not be identified in the field were fixed in 10% formalin for 24 hours and subsequently preserved in 70% ethanol.Voucher specimens were deposited at the National Fishery Research Centre, Godavari, Lalitpur, Nepal.

Bird survey
Bird surveys were conducted using the open-width point count method along transects near the lake's shoreline, following the protocol outlined by Bibby et al. (2000).Additionally, we employed the area search method during the field study, as described by Slater (1994), Dieni andJones (2002), and Adhikari et al. (2022).The length of each transect was determined based on the specific characteristics of the habitat and landscape, following principles outlined by Hanowski et al. (1990).Within each transect, we established a minimum of five vantage points at 100-meter intervals, where we used binoculars (Nikon 20x50) to count bird species.At each point, we conducted five-minute counts of bird species.We documented all observed species, aided by both visual and auditory observations, including relevant habitat and environmental variables.To ensure data accuracy, each transect was surveyed by two observers.Subsequently, we combined the recorded bird species lists from various vantage points within each transect.Survey periods included mornings from 600 h to 1200 h and evenings from 1500 h to 1800 h.Bird species were identified using the field guidebook for birds of Nepal authored by Grimmett et al. (2016a, b).

Herpetofauna
Both amphibians and reptiles were surveyed using nocturnal and diurnal and transects respectively in a time-constrained visual encounter survey (Khatiwada 2012;Khatiwada et al. 2016;Khatiwada et al. 2019).Transects were searched by four people for two hours using torches, walking at a slow pace at night (700 h -900 h) and during the day (1000 h-1300 h).The number of species and individuals encountered in each transect was recorded along with all habitat and environmental variables.Apart from nocturnal and diurnal transects, opportunistic random surveys were also carried out to document the occurrence of herpetofauna species in the area.All individuals encountered were captured and stored in a 15 L plastic bucket with small holes in the lid.Some uncaptured individuals were also counted.All captured individuals were taken to a nearby dry place where the animals were measured and identified at the species level based on guide books: Schleich & Kästle (2002) and Shah & Tiwari (2004), and then released back into their original habitats.Male frogs were identified based on secondary sexual characteristics in the presence of black pigment on the throat (vocal sac) and nuptial pads, and females by the enlargement of the coelomic cavity in gravid individuals.Specimens that were difficult to identify based on morphological traits in the field were euthanized in a chlorobutanol solution, fixed in formalin for 24 hours, and subsequently preserved in 75% ethanol.The morphological parameters (e.g., body length, fin length, and eye diameter) were measured and compared with identification keys.The species nomenclature follows Frost (2019).Voucher specimens were stored at the Central Department of Zoology, Tribhuvan University, Kathmandu, Nepal.

Mammal survey
Five systematic transects (varying from 0.42 to 1.5 km) were laid in the riparian areas.The transect line was searched by 2-3 people, and all the animals sighted and indirect signs of mammals such as scats, pellets, droppings, dung, pugmarks, scrapings, carcasses, quills, and burrows were recorded.Apart from these, an opportunistic survey was also conducted around the lake to record mammals.

Macrophytes and vegetation survey
Macrophytes and vegetation were surveyed in and around the lake to enumerate the checklist of plant species.Surveying along line transects starting from shore to the lake interior is a flexible method to document macrophytes in small lakes (Titus 1993).We used two 25 m long-line transects from Lake Shore to the centre and noted the macrophytes at different distances.A floating tube was used to swim, and a rake was used to collect submerged macrophytes.
A vegetation survey in the surrounding forests (about 100 m from the lake shore) was carried out to prepare the checklist of plants occurring in the lake complex area.Transect walks along the trails and through the forest were performed to collect plant specimens.
Collected specimens were identified on-site, while unidentified specimens were preserved following standard herbarium methods (Bridson & Forman 1999).Herbarium specimens prepared for further identification were deposited at the National Herbarium and Plant Laboratory in Kathmandu.Plants were identified using relevant identification keys (Polunin & Stainton 1984;Grierson & Long 1983, 2001).

Data analysis
The Nepal Lake Biotic Index (NLBI) for lakes and the Biotic Index (Shah et al. 2020c) for running waters (inlet and outlet) were calculated by assigning tolerance scores to macroinvertebrates identified at the family level (Shah et al. 2011(Shah et al. , 2020c)).In these methods, the index value is the sum of the tolerance scores divided by the number of scored taxa for a site, which then J TT translates to the lake water quality class (LWQC) for indicating the degree of degradation).
Non-metric Multidimensional Scaling (NMDS): Nonmetric multidimensional scaling (NMDS) was performed to cluster sites based on macroinvertebrate abundance data.Prior to analysis, macroinvertebrate abundance data were transformed to log (x+1).The Bray-Curtis distance measure was employed in NMDS, and the analysis was conducted using the R package (R Core Team 2019).

Shannon diversity index (H)
The Shannon diversity index (H) was used to assess species diversity within a community (Shannon 1948 Where p i is the proportion (n/N) of individuals of one particular species found (n) divided by the total number of individuals found (N), ln is the natural log, Σ is the sum of the calculations,

Community Diversity Measurement -Simpson Index (D):
The Simpson index was determined to measure community diversity in relation to habitats (Simpson 1949).
Where n is the number of individuals of one particular species, N is the total number of individuals found.Σ is the sum of the calculations.D values range between 0 and 1.

Evenness and Equitability:
Evenness (e) was used to determine the distribution of individuals of a taxon in a community.It is constrained between 0 and 1.0: Evenness = H'/Hmax Where H' is the Shannon diversity index H max is the maximum taxon recorded at a site.
Jacob's Equitability index: Jacob's Equitability (J) was used to measure the evenness with which individuals are divided among the taxa present.

Equitability (J) = H'/lnS
Where H' is Shannon's index of diversity, S is the number of taxa Fisher's Diversity Index: Fisher's index describes the mathematical relationship between the number of species and the number of individuals in those species (Fisher & Yates, 1943).The Fisher diversity index is defined implicitly by the formula below: Fisher's diversity index Where n is the number of individuals and a is Fisher's alpha).

Biodiversity survey Aquatic macroinvertebrates
A total of 45 families belonging to 14 orders of macroinvertebrates were recorded in the study lakes including the inlet and outlet of the Ramaroshan Lake complex (Supplementary 1).Diptera was the most dominant and diverse order followed by Odonata and Mollusca in the lakes, while Trichoptera was the most dominant and diverse order followed by Diptera and Ephemeroptera in running waterbodies (inlet and outlet streams) of the lakes (Figure 2).Plecoptera was found only in Lamadaya Lake and running water bodies.Among lakes, Lamadaya was found to be highly diverse in terms of taxa composition, while Ramaroshan was the least diverse.Family richness ranged from 10 to 25 in the lakes, and 14 to 30 in running waterbodies.Family

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richness was low for the winter season compared to the summer season in the lakes and running water bodies (Figure 2) The lakes were categorized into a 'fair' LWQC for both seasons while the water quality class for running water bodies was categorized into a 'good' status for winter and a 'fair' status for the summer season in the outlet (Figure 3).
Non-metric multidimensional scaling (NMDS) disentangled sites into three clusters-Cluster 1 representing sites of running water bodies, Cluster 2 for sites of Lamadaya, and Cluster 3 for sites of the remaining three lakes (Figure 4).

Fish
Altogether, three species of fish, namely Schizothorax nepalensis, S. richardsonii, and Garra gotyla, belonging to the Cyprinidae family, were recorded in the lakes of Ramaroshan Lake Complex.Among these reported species, S. nepalensis listed as Critically Endangered, which is endemic to northwest Nepal (Regmi et al., 2021b), and S. richardsonii which is common to major river systems (Koshi, Gandaki, and Karnali), is listed as a vulnerable category in the IUCN Red List.

Bird survey
In total, 1018 individuals (winter = 611 and summer = 423) of birds from 79 species belonging to 33 families and 15 orders were documented in the lake complex

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There was no significant variation in the Shannon diversity index, Species dominance index, and Simpson index of diversity for birds between winter and summer seasons (Table 3; p = 0.79).The species' evenness of birds (0.82) and Jacob's coefficient of equality (0.95) were lower in winter than in the summer season (evenness = 0. 80, Jacob's coefficient of equality = 0.95).

Herpetofauna
Within the lake complex, a comprehensive survey documented a total of 121 amphibians, representing 7 distinct species distributed across 5 families.Notably, Nanorana legibii dominated the population at 50.4%, followed by Duttaphrynus himalayanus at 32.2% and Hoplobatrachus tigerinus at 5.8% (Figure 7).Two endemic amphibian species, Nanorana minica and Amolops marmoratus, were also identified at the study sites (Table 4).It is worth highlighting that both Liebiegi's Paa Frog Nanorana legibii and Small Paa Frog Nanorana minica are categorized as globally Vulnerable on the IUCN Red List.Furthermore, the Indian Bull Frog Hoplobatrachus tigerinus, classified as globally Near Threatened by the IUCN in 2021, was also observed within the lake complex.
A total of five species of reptiles were recorded during the field survey.Among them, Laudakia tuberculata (48%) was the most abundant species in the study area, followed by Calotes versicolor (25%), and Eutropis carinata (21.4%), respectively (Table 4).

Mammals
This study documented a total of 12 mammal species.Notably, four of these species enjoy legal protection under the DNPWC Act of 1973, enforced by the government of Nepal.These protected species include the Leopard Panthera pardus, the Red Panda Ailurus fulgens, the Asiatic Black Bear Ursus thibetanus, and the Himalayan Goral Naemorhedus goral.The Red Panda is of particular concern as it holds the classification as being 'Endangered' according to the IUCN Red List.Similarly, the Himalayan Black Bear and Leopard are categorized as 'Vulnerable' under the IUCN Red List, while the Assam Macaque and Himalayan Goral fall within the 'Near Threatened' category (Table 5).

Macrophytes and Vegetation Survey
In total, the lakes of the Ramaroshan Lake complex harbored 25 species, encompassing 14 families of macrophytes (Table 6).Predominantly, Scirpus compressus, Scirpus sinensis, and Polygonum hydropier thrived as major emergent plants along the shores and in marshy areas.Among submerged vegetation, Ceratophyllum demersum and Potamogeton nutans prevailed.The complex featured Nelumbo nucifera as the sole-rooted floating macrophyte species.Additionally, the region supported two wetlanddependent plants, Allium waalichina and Ophioglossum nudicaule, esteemed for their medicinal attributes and utilized as vegetables by the local populace.
Expanding the scope, the Ramaroshan Lake complex area showcased an impressive biodiversity of 167 plant species, spanning 70 families (Supplementary 3).

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Notably, Rosaceae stood out as the largest family with 17 species, followed closely by Asteraceae with 14 species and Poaceae with 13 species.Taxus wallichiana, a valuable medicinal plant, flourished abundantly within the complex area.It's worth mentioning that this species holds a spot on the IUCN Red List as endangered and is also listed in CITES Appendix III.

DISCUSSION
Ramaroshan Lake complex is known for its unique landscape and high biodiversity.The lake complex provides forest resources such as fodder for livestock and bamboo and firewood for household consumption in adjacent communities.The complex is an excellent area for livestock grazing.Local inhabitants obtain water for drinking purposes, domestic consumption, and irrigation.Like other wetlands in Nepal, the lake complex is also affected by land encroachment.In 30 years, the wetland area of the complex has shrunk by 16% due to land use and land cover changes (Paudel et al. 2022).

Water quality status
Ramaroshan Lake complex, being situated in the remote mid-hills of the country, has water quality parameter values for all four lakes within the permissible limit for the winter and summer seasons (see Table 2).Dissolved oxygen (DO) for all four lakes was greater than 5 mg/L, indicating good status for maintaining higher forms of aquatic life in water (Bozorg-Haddad et al. 2021).DO was highest in Lamadaya lake (6.35 mg/L) and lowest in Batula lake (5.26 mg/L).Similar DO values were also reported from the lake complex for the winter season in 2020 (Chalaune et al. 2020) and were comparable with the DO values recorded in other lakes of the region (Gurung et al. 2018).DO greater than 4 mg/L is suitable for bathing, aquaculture, and irrigation (Bozorg-Haddad et al. 2021).
Lamadaya Lake unlike other lakes, had many physical parameters that were significantly different.This might be due to its smaller size, being relatively shallow, and being surrounded by dense forest.Hydrological fluctuation between seasons might have played a major role in making it different (Regmi et al. 2021a).Similarly, Ramaroshan Lake and Batula Lake being situated in the lower region of the lake complex, the nutrient parameters such as nitrate and phosphate; hardness, and alkalinity were found to be high compared to Jingale and Lamadaya lakes.

Biodiversity
Ramaroshan Lake complex is situated in the temperate zone, low species richness can be expected in comparison to lowland Tarai because species richness declines with increasing elevation in the Himalayas (Shah et al. 2015;Basnet et al. 2016;Araneda et al. 2018).

Aquatic macroinvertebrates
Water quality is a crucial parameter that determines biotic community composition in lake environments.We observed significant changes in water quality parameters across the lakes (Table 2), and this could be a key factor for the differences in the composition of macroinvertebrates between Lamadaya and the other lakes (Figure 3, 4).Warm water-adapted macroinvertebrates of insect orders Odonata, Coleoptera, and Hemiptera, together with annelid worms and Mollusca were diverse and abundant in lakes during the summer season (Figure 3).These findings are similar to the findings for tropical lakes (Shah et al. 2011;Shrestha et al. 2021).Diverse macroinvertebrates were recorded in Lamadaya Lake which might be due to the occurrence of mosaic habitats mainly comprised of soft substrates like leaf litters, twigs, and macrophytes.Soft substrates not only provide suitable habitats for macroinvertebrate colonization but also prevent predation (sensu Shah et al. 2011).

Fishes
The lake complex highlights its importance as a critical habitat for critically endangered fish species: Snow Trout Schizothorax nepalensis and vulnerable fish species (Schizothorax richardsonii).

Birds
The Ramaroshan Lake Complex plays a pivotal role in providing a crucial habitat for bird diversity, as evident from Supplementary 2. A notable highlight is the documented presence of the globally endangered vulture species Neophron percnopterus, the Egyptian Vulture, underscoring the complex's significance.This mountainous ecosystem serves as a sanctuary for J TT globally threatened vulture species, with a majority (7 out of 9) choosing mountain cliffs and towering trees for nesting (DeCandido et al. 2012).Notably, this study reveals that 8.9% of the bird species documented in Nepal, amounting to 891 species according to DNPWC and BCN 2019, find a habitat in the Ramaroshan Lake complex.In Nepal, approximately 200 wetland bird species have been recorded (BCN 2018), with 15 of them (7.5%) also making their presence known in the Ramaroshan Lake complex.It's worth noting that the winter season sees a decline in the sighting of wetland birds, likely attributed to the sub-zero temperatures during this period (DFO 2019).

Herpetofauna
Ramaroshan Lake complex is rich in herpetofauna diversity.A total of 7 species of frogs and 5 species of reptiles were recorded during this study, which is 12.5% (out of 56) and 4.5% (out of 117 species) of the total species recorded from Nepal, respectively (Shah & Tiwari 2004a).High altitude supports a low number of herpetofauna as they are cold-blooded animals (Khatiwada et al. 2019).
Among the recorded amphibian species, Liebiegi's Paa Frog Nanorana legibii and Small Paa Frog Nanorana minica are listed under the globally vulnerable category, while the Indian Bullfrog Hoplobatrachus tigerinus is listed as a globally near threatened species.Studies have shown that frogs are an important source of livelihood for many people (Khatiwada & Haugaasen 2015) and remain an integral part of local medicinal heritage (Mohneke et al. 2011;Lynch et al. 2023).Amphibians and reptiles have long been used by humans as food and medicine (Gonwouo & Rödel 2008;Khatiwada & Haugaasen 2015).Local people in the Ramaroshan areas use Paha frogs (Nanorana and Amolops species) as food and medicines.Over-collection of the species may lead to local extinctions or severe population declines.As many amphibian species predictably aggregate for reproduction or hibernation, this makes them particularly vulnerable to intensive collecting efforts.Recent studies have indicated that commercial or subsistence harvesting has contributed to a decline in many reptile species (Webb et al. 2002).Khatiwada & Haugaasen (2015) revealed that Paa and Amolops are the most exploited frog species by the local people for food and medicinal purposes in mountainous parts of Nepal.This heavy exploitation may also lead to local or global declines and even extinctions through unsustainable collection (Warkentin et al. 2009).

Mammals
The mammals in the Ramaroshan lakes area were reported based on signs and direct sightings, and 12 species of mammals were reported during the field study.Among the reported species, one is endangered, two are vulnerable, and two are near threatened mammals, according to the IUCN Red List.Nepal supports 212 species of mammals (Amin et al. 2018), but this small area alone supports 12 species of mammals thus highlighting the importance of the Ramaroshan Lake complex for mammal conservation.

Macrophytes and Vegetation
This study significantly expanded upon the existing knowledge of the area's flora by documenting a total of 26 macrophyte species, thus surpassing the previously reported count of 10 wetland species (Paudel & Pandey 2016).Furthermore, compared to an earlier study (DoF 2017) that documented only 124 plant species within the lake complex (see Supplementary 3), our research uncovered additional plant species.It's worth noting that there is a limited body of research assessing macrophytes in the mid-hills (Basnet et al. 2016), Churia, and Tarai regions of Nepal (Regmi et al. 2021a).For instance, Basnet et al. (2016) identified fewer than 10 macrophytes in Rara Lake, located in the High Mountain region.In contrast, the wetlands of the Tarai-Plain, as highlighted by Regmi et al. (2021a) and Burlakoti & Karmacharya (2006), hosted over 50 macrophyte species.This observation suggests a pattern of increasing macrophyte species richness from the high mountain to the lowland Tarai regions.Despite its location in the mid-hill region, the Ramaroshan Lake Complex exhibited a modest richness of macrophytes.Additionally, the presence of terrestrial flora, including endangered species like Taxus wallichiana, contributes to the overall biodiversity of the lake ecosystem.It's important to note that our vegetation survey was exploratory, and further extensive sampling in both forests and lakes may reveal more plant species.

Threats to the Ramaroshan Lake Complex
The lake complex is a tourist destination for local people in the district.However, the area is not as well visited by domestic or international tourists as other lakes in Nepal, such as Gosaikunda, Rara Taal, Pokhara Lake Clusters, etc., due to poor road and air connectivity despite its beautiful landscape.Therefore, minimum tourist influences and minimum activities can be seen.However local people visit the areas frequently for fodder collection, and they use the lake complex for J TT grazing their livestock.Some of the local people are often sighted poaching birds such as the Kalij Pheasant Lophura leucomelanos for meat consumption (Aditiya Pal pers.comm.June 2019).Local people harvest Paha frogs (Nanorana and Amolops species) in large quantities for food and medicinal purposes, which may affect the population of the species in the near future.Plastic pollution is increasing in the littoral sections of the lakes

Conservation value of Ramaroshan Lake Complex
The Ramaroshan Lake Complex and its surrounding catchment area are home to a multitude of species with significant conservation value.These include various aquatic macroinvertebrates such as the Relict Himalayan Dragonfly Epiophlebia laidlawi, fish species like Schizothorax nepalensis and Schizothorax richardsonii, bird species including the Egyptian Vulture Neophron percnopterus, Asian Woolly-necked Ciconia episcopus, Cheer Pheasant Catreus wallichii, and Northern Lapwing Vanellus vanellus, frog species such as Liebiegi's Paa Frog Nanorana legibii, Small Paa Frog Nanorana minica, and Indian Bull Frog Hoplobatrachus tigerinus, as well as mammal species including Royle's Pika Ochotona roylei, Assam Macaque Macaca assamensis, Himalayan Goral Naemorhedus goral, Leopard Panthera pardus, Himalayan Black Bear Ursus thibetanus, and Red Panda Ailurus fulgens.The lake systems are encompassed by pasturelands, expansive grasslands, and dense forests that further support a diverse range of wetland-dependent and forest birds.Given its unique geographical location, suitable wetland habitat, native biodiversity, and essential ecosystem services, the Ramaroshan Lake Complex meets the criteria for designation as wetlands of international importance (Ramsar Site).It is imperative that the Ramsar focal agency for Nepal actively pursue this designation.
With the country's adoption of a federal structure, there exist opportunities to integrate wetland management considerations by formulating regulatory frameworks at the central, provincial, and local levels.To ensure the sustainable management of these wetlands, it is crucial to engage and incorporate local communities into this regulatory framework.This approach will facilitate timely monitoring, restoration efforts, and the judicious utilisation of wetland resources.

Table 2 .
Mean and standard values of physico-chemical parameters for each study lake for the winter and summer seasons of year 2018 and 2019.Values indicated in bold digits are significant between seasons.The symbols (Asterisks) "*", "**" and "***" represent significance levels at 0.05, 0.01, and <0.001.

Figure 2 .
Figure 2. The family richness of aquatic macroinvertebrates was recorded in the winter and summer seasons of the year 2018 and 2019 in the study sites of the Ramaroshan Lake Complex.Inlets and outlets are running streams coming to the lake and leaving the lake.

Figure 3 .
Figure 3. Lake water quality classes of four study lakes and water quality classes for running water bodies.Yellow color indicates fair water quality while green indicates good water quality.

Figure 5 .
Figure 5. Number of bird species recorded with respect to order for winter and summer seasons in the Ramaroshan Lake Complex.

Figure 6 .
Figure 6.Number of bird species across habitat types recorded in and around Ramaroshan Lake Complex.

Figure 7 .
Figure 7.The relative percentage of amphibian species recorded in the Ramaroshan Lake Complex.

Bird species with their number of individuals observed in the Ramaroshan Lake Complex area. Abundance (%) refers to the total percentage contribution of each species to the total sample for both seasons. EN = Endangered, VU = Vulnerable, NT= Near threatened and LC= Least concerned.
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