Biodiversity, distributions and isolation of microplastics pollution in finfish species in the Panjkora River at Lower and Upper Dir districts of Khyber Pakhtunkhwa province of Pakistan

The basic aim of this study was aimed to determine the ichthyofaunal diversity of River Panjkora in both upper and lower Dir districts in Khyber Pakhtunkhwa province of Pakistan.Fish samples were collected by using fishnets from March to September 2020. A total of 724 specimens were collected and classified into 5 families, 14 genera, and 18 species. The overall results revealed that most fish fauna of river Panjkora contains 8 species of family Cyprinidae (56.49%) followed by 4 species of Nemacheilidae (24.44%), 2 species of Channidae (10.63%), and Sisoridae (7.04%), and 1 species of Mastacembelidae (1.38%), respectively. Among all kinds of fish species, Schizothorax plagiostomus (16.57%) was highly dominated and followed by Carassius auratus (11.87%) and Racoma labiata (9.66%) and were reported as highly abundant, especially during April, May, and June. The least abundant species were Glyptothorax punjabensis, Glyptothorax sufii, and Mastacembelus armatus , that constituting 2.48%, 2.20%, and 1.38% of the total fish samples. The Overall Simpson’s diversity (1-D= 0.919) and Simpson’s Reciprocal index values (1/D= 12.3876), and Shannon’s index (H= 2.68) were indicating that river Panjkora contains a quite rich and diverse group of fish species. The highest microplastics observed in site 7 compared to other study area. Conservation steps should be taken as a top priority to protect and conserve the marine environment and natural heritage from further loss, extinction and stop or minimize losses incurred through irresponsible fishery practices


Biodiversity, distributions and isolation of microplastics pollution in finfish species in the Panjkora River at Lower and Upper Dir districts of Khyber Pakhtunkhwa province of Pakistan
Biodiversidade, distribuição e isolamento de poluições por microplásticos em espécies de peixes no rio Panjkora nos distritos de Lower e Upper Dir da província de Khyber Pakhtunkhwa do Paquistão on Fisheries of the University of Karachi and University of Malakand KPK Pakistan

Study area
District Dir is located in the Khyber Pakhtunkhwa province of Pakistan, at 34º, 37′ to 35º, 07′ North and 71º, 31′ to 72º, 14′ East, and found at 823 meters (2700 feet) above the sea level. Kohistan of Upper Dir is the actual origin of River Panjkora. This river flows southward and almost passes through the middle of both the Lower and Upper Dir districts, and meets the Swat River in Sharbatti, District Malakand, behind Totakan (Bosaaq Bridge). The Panjkora River was called for its five major tributaries, which meet it at four locations: Chukiatan for Barawal Stream, Sheringal for Gwaldi Stream, Akhagram for Dobando Stream, and both Nurhund and Usherai Dara Streams for Nurhund Stream (Figure 1).

Fish collection
In this study, collection of fish sampling were made at seven locations in River Panjkora during the study period from March to September 2020 ( Figure 1). Fishes were caught with gill nets, cast nets and hooks, and preserved in 10% formalin solution.

Measurements and weighting
All measurements and weight of fishes were made by using the meter scale and Venire calliper. The total length (TL) of fishes was resolute by using scale and china tape. Body weight was noted with the help of an electric balance. Lateral-line scales were counted from the anterior to the posterior region scales. Transversal scales were counted as scale rows found above the lateral line and below the lateral line separately

Fish identification
All fishes identified were made by using standard taxonomic keys provided by Mirza (1990) namely "Pakistan ki Taza Pani ki Machliyan", Freshwater Fishes of the Indian Region (Jayaram, 1999) and Inland fishes of India and adjacent countries by Talwar and Jhingran (1991) (Figure 2).

Microplastic isolation
Microplastic isolated from the selected fishes by following the methods of Hassan et al. (2023).

Results and Discussion
A total of 724 specimens contain 5 families and 18 species as shown in the Table 1. Cyprinidae was the most dominant 56.49% (n=409/727) followed by

Introduction
One billion people globally consume fish, which is one of the major sources of high-quality protein and are used to develop nation economy Ahmad et al., 2020). Fish is the cheapest source of high-quality protein fatty acids and antioxidants that can protect our body against certain diseases like cancer. Fish are susceptible to ecological modifications, and their diversity in a broad spectrum of differential tolerance reflects the health status of the ecosystem (Chowdhury et al., 2019;Hassan et al., 2020a;Sabbir et al., 2020;Hussain et al., 2021). Changes in ecosystems of the food tropical level implanted remarkable effects on the abundance and distribution of fish. Pakistan has rich freshwater fish fauna, which is represented by 193 fish species (Rafique and Khan, 2012;Hassan et al., 2023). More than 180 species had been reported in the Indus River in Pakistan (Mirza and Mirza, 2014). Many researchers had made valuable contributions in the study of the fish fauna of the Khyber Pakhtunkhwa, including Ahmad and Mirza (1963), Khan (1983), Butt (1986), Javed et al. (1996), Mirza (2007), Ahmad and Hasan (2011), Hasan et al. (2013, 2015, Saeed et al. (2013), Ishaq et al. (2014), Muhammad et al. (2014) and Akhtar et al. (2014), respectively. However, the broad range of biodiversity and distribution patterns of fishes found in the Panjkora, river, and its tributaries require more investigation. Fish fauna of the Panjkora river contains a high proportion of endemic species were belong to three genera includes i.e., Triplophysa, Schistura, and Glyptothorax. All these species are mainly cold water fishes and confined only in headwater hilly streams and rivers, and isolated from warm water fish populations stretches between them. The growing decline in fish biodiversity has now shocked the world conservation community. Roughly about 20% of the world's freshwater fishes are currently in endangered or extinct stages (IUCN, 1989;Hassan et al., 2020b). While the identification and characterization of existing species had been considered as an essential part of their conservation and needs more attention in the scientific literature (Hassan et al., 2020a;Romanova et al., 2018). Fish species composition and their relative abundance might be changed due to the seasonal variations (Jisr et al., 2018;Abro et al., 2020;Islam et al., 2020). The present study was aimed to know the current status of the fish fauna of River Panjkora at Districts Dir (Upper and Lower), Khyber Pakhtunkhwa, Pakistan.

Figure 1. River Panjkora District Dir (Upper and Lower) Pakistan digits refer to the sampled localities (see
The most abundant species of family Megachilidae was Schistura alepidota 8.28% and found in one site. Family sisoridae was represented by three fish species, Glyptothorax punjabensis, Glyptothorax. sufii and Nangra robusta with diversity value (D=0.004493 and H=0.3438). While the most abundant species of family Sisoridae was Nangra robusta having 5.93% of the total catch and distributed in three sites. Family Channidae includes two fish species, i.e., Channa gachua and Channa punctate that contributed 7.04% of total fish caught and with total diversity value (D=0.002464 and H=0.2346). The most abundant species of family channidae was Channa gachua having 4.14% of the total catch and was found in two zones. Family Mastacembelidae was represented by only single species i.e., Mastacembelus armatus that contributed 1.38% of the total fish caught and having total diversity value (D=0.0001719 and H=0.05915) and found in only one side of the study area Fishes caught during thisstudy were showing theirabundance in the month of May with highest diversity value (D= 0.323777 and H=0.94565), and less abundant in September with lowest diversity value (D= 0.022855 and H=0.06675). The overall Simpson's diversity (1-D= 0.919) and Simpson's Reciprocal index values (1/D= 12.3876), and (Shannon's index = 2.68) were indicating that River Panjkora is quite rich and contain diverse groups of fish species.. Thus, this study also reveleved that fish diversity of River Panjkora was high in the month of May with 14 fish species and 35% abundance, followed by June with 11 fish species and 19% abundance, July with 12 fish species and 15% abundance, April with10 fish species and 12% abundance, March with6 fish species and 8% abundance, and August and September months were with least diversity (6 to 8 fish species and 7% and 2% abundance as shown in the Table 2 and 3, respectively. In this study, about 5 families, 14 genera and 18 fish species of the and were reported in Panjkora River,, which was less than Islam and Siddiqi (1971), who also had been reported eight families, 16 genera and 21 fish species from river Jhelum. On the other hand, Mirza and Sandhu (2007) had described about 189 species from the Indus River; while Rafique et al. (2003) , vol. 42, no. 3, pp. 285-289. BUTT, J.A., 1986. Fish andFisheries of KPK, Pakistan. Biologia, no. special, suppl. 1986, pp. 21-  In this study, a total of 120 fish individuals collected from the each species. Microplastics were found in 12 of the 120 examined fish.A totals of 10 microplastic items were recovered from the 100 individuals: 4 microplastics from the gastrointestinal tract (25%), 6 items from the gills (50%) and 4 from the muscle (25%). Based on microplastic size, all the species had more fibers in the size range 100-400 mm in the gastrointestinal tract and 98-140 mm in gills. In the dorsal muscle, in the size range 80-600 mm (Lusher et al., 2013;Ory et al., 2018) The highest microplastics (90%) observed in site 7 and (10%) microplastics found in site 4 and 5. No microplastis were found in sites 1, 2 and 3. According to the recent study the site 7 is contaminated site because medical and general store discarded a lot of polthene bags in the river. The plastics have the chemical composition from which they are made (Jabeen et al., 2017;Hassan et al., 2023). High incidences of environmental pollution and polymerization processes causes contamination reach the food chain, dangerous long-term carcinogenicity. Impact on fish, animals and humans due to the release of chlorofluorocarbons (cfcs), formaldehyde, lead, mercury and cadmium. The accumulation of polyethylene bags affected on survival and growth of fishes, like habitat conditions, development pattern, early life, feeding, fish fatness, degree of stomach completeness, gonad middle age, physical condition, overall health, gastrointestinal tract, dorsal muscle, lipid oxidative damage in the brain gills and increased brain AChE activity in fish containing microplastics similar finding (Hester-Ronald and Harrison, 2011;Hassan et al., 2023).