Research article · 2026 · Full text
International Journal of Biology SciencesSingle and combined effects of microplastics and wastewater on kidney of Cyprinus carpio
Volume 8(3) · Pages 30–38 · DOI: 10.33545/26649926.2026.v8.i3a.592 ↗
Research areasMicroplastics and Plastic PollutionEnvironmental Toxicology and EcotoxicologyMarine Biology and Environmental Chemistry
Abstract
Author-writtenRapid pace of urbanisation and industrialisation has led to accumulation of different pollutants such as domestic sewage, agricultural runoff and industrial waste in water bodies. Additionally, microplastics (MPs) are problematic due to their toxicity, environmental persistence, abundance and ability to adsorb different contaminants. Among different aquatic organisms, fishes are considered as an exceptional indicator to study water quality and well-being of aquatic ecosystem due to their sensitivity to alterations in environmental conditions. In addition, they are an important protein source; however, toxicants in them could affect health of human beings consuming them. Therefore, the current study was designed to study the combined effects of low-density polyethylene (LDPE) MPs and wastewater on histomorphology of kidney of minor carp Cyprinus carpio. The results showed that significant alterations such as paracellular spaces, damaged hematopoietic tissue, detachment of renal tubule from basal lamina, damaged renal tubules, paracellular spaces, vacuolization, necrosis and exudate were found in kidneys of fingerlings belonging to treated groups. Moreover, Organ index of kidney also indicated significant (p≤0.05) rise in different treatment groups in comparison to control group. Overall, the results concluded that the combination of LDPE MPs and wastewater effects kidneys of C. carpio fingerlings in duration and dose-dependent manner with limited reversibility following exposure.
Summary
Source-checkedPlain-language reading of the full paper, prepared with AI and checked against the text. Each point links to the authors' own words.
Why it matters
- Urbanization and industrialization have caused various pollutants, including industrial waste and domestic sewage, to accumulate in water bodies. View evidence ↗
- Fish are highly sensitive to environmental changes, making them excellent bioindicators for assessing water quality and ecosystem health. View evidence ↗
- Microplastics (MPs) are a significant concern for aquatic life because they are toxic, persist in the environment, and can adsorb other contaminants. View evidence ↗
The question
- The study aimed to investigate how the combination of wastewater and low-density polyethylene (LDPE) microplastics affects the kidney structure of the minor carp, Cyprinus carpio. View evidence ↗
What they did
- Cyprinus carpio fingerlings from a fish farm in India were used and acclimatized in 100-liter glass tanks for 14 days. View evidence ↗
- The lethal concentration (LC50) was 64.47% for wastewater alone and 53.72% for the combination of wastewater and 2 mg/l microplastics. View evidence ↗
- The chronic bioassay lasted 60 days with 15 fingerlings per replicate (in triplicate), followed by a 60-day recovery period in dechlorinated water. View evidence ↗
- Kidney histomorphology was analyzed using 3–5 µm serial sections stained with haematoxylin and eosin. View evidence ↗
What they found
- After 60 days of exposure, all treated groups (T1-T6) showed significant kidney damage, including necrosis, exudate, and damaged renal tubules. View evidence ↗
- The combined exposure of microplastics and wastewater (COMB1 and COMB2) caused more severe kidney damage than either pollutant alone. View evidence ↗
- Recovery was incomplete for combined treatments (T5, T6), which still showed damaged renal tubules and hematopoietic tissue, while T1 and T2 returned to normal structures. View evidence ↗
- The kidney organ index increased significantly (p ≤ 0.05) after 60 days for all treatment groups compared to the control, except for the 1 mg/l microplastics group. View evidence ↗
What it means
- The authors conclude that combined LDPE microplastics and wastewater cause dose- and duration-dependent kidney damage with limited reversibility, posing long-term risks to aquatic health. View evidence ↗
Visual summary
1 visualsIllustrations explain the paper; they do not show study data.
Synergistic Toxicity Concept
AI-generated conceptual illustration (Cyprinus carpio (common carp) kidney tissue). It explains the idea and does not show study data.
View source evidence ↗Figures & tables
From the original paper
Table 1
Figure 1
Figure 2
Figure 3
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Figure 5
Figure 6Introduction
Water is an essential natural resource that sustains all life forms on Earth; however, the rapid pace of urbanisation and industrialisation has led to the daily discharge of about 2×10 6 tons of different pollutants including domestic sewage, agricultural runoff and industrial waste (Mahmoud et al., 2026) [14] . Wastewater contains a plethora of anthropogenic contaminants such as pharmaceutically active compounds, heavy metals, bisphenol-A, nanomaterials, organic pollutants and endocrine-disrupting chemicals (Klecka et al., 2010; Mehdi et al., 2021; Vaid and Vashishat, 2024) [12,15,23] . Among these, plastic pollution has become a pervasive environmental concern due to its persistence, mismanagement and abundance. Plastic refers to synthetic and organic polymers that are widely used in different industries due to their low production cost, durability, strength and versatility (Kumar et al., 2018) [13] . Plastic particles are categorised from macroplastics (≥ 25 mm) and mesoplastics (5-25 mm) to microplastics (MPs) (20 µm-5 mm) (Gigault et al., 2018) [5] . However, their resistance to biodegradation has led to a significant accumulation in terrestrial and aquatic environments, posing severe ecological and health risks (Rakib et al., 2022) [19] . Among plastic particles of different size groups, MPs are of increasing concern for aquatic organisms due to their toxicity, environmental persistence, abundance and ability to adsorb different contaminants (Vaid and Vashishat, 2025) [24] . Among different aquatic biota, fishes are considered as an exceptional bioindicator to study water pollution due to their sensitivity to environmental fluctuations (Cvetkovic et al., 2010; Vaid and Hundal, 2019) [4,25] . Various studies reported that MPs may induce behavioural alterations, genotoxicity, oxidative stress, histopathology and mortality (Motivarash et al., 2024) [16] . In addition, they are an important protein source and act as a sensitive indicator of overall well-being of the aquatic ecosystem (Vaid et al., 2024; Bakhasha et al., 2025) [26,2] . Among various organs, kidneys are essential excretory organs responsible for eliminating metabolic wastes and xenobiotics; however, prolonged accumulation of various toxic substances may pose significant health issues in fishes (Ortiz et al., 2003) [18] . Moreover, toxicant-induced histological and functional alterations in kidneys can disrupt homeostasis (Iqbal et al., 2004) [9] . Despite escalating concerns over water contamination, research addressing the combined effects of MPs and other contaminants on fish remains limited. Therefore, the present study was designed to study the combined effects of low-density polyethylene (LDPE) MPs and wastewater on histomorphology of kidney of minor carp Cyprinus carpio.
Materials and Methods
Materials and Methods
Collection of wastewater and fingerlings
For conducting experiments, wastewater was collected from the sewage water treatment plant, Bhattian, India. The sampling site was situated within the municipal boundaries about 11 km from Punjab Agricultural University, India. C. carpio fingerlings were collected from Aujla fish company fish farm, India and acclimatized for 14 days in 100 l glass tanks. Fingerlings were fed with commercial feed/day at 2% of body weight. Fingerlings were not fed for 24 h prior to the commencement of the experiment. The experimental protocol adhered to the standard guidelines of the Organisation for Economic Co-operation and Development (OECD, 1992) [17] .
Preparation of MPs
Polyethylene plastic pellets (Sigma-Aldrich) were used for the preparation of MPs following the methodology described by Wong et al. (2014) [27] . Briefly, 30 pellets were pre-heated with toluene (6 ml) and Tween-20 (0.2 ml) and subjected to ultrasonication for 30 min, followed by spreading onto a Petri plate and air-drying to completion. The resulting MPs were subsequently utilised for toxicity bioassays.
Toxicity bioassays-Acute and Chronic
The acute bioassay was conducted according to the standard guidelines of the Organisation for Economic Co-operation and Development (OECD, 1992) [17] . For lethal concentration (LC50) calculation, 2 batches (n = 7) of fingerlings each were treated with various concentrations of wastewater (20, 40, 60, 80 and 100%) and LDPE MPs (350-750 mg/l) separately for a period of 96 hours. The results showed no mortality in fingerlings exposed to MPs; thus, sub-lethal concentrations were selected based on environmentally relevant levels, i.e., 1 and 2 mg/l MPs (Rochman et al., 2017) [21] . The LC50 of wastewater, determined using POLO software (Robertson et al., 2007) [20] , was 64.47%. After that, the LC50 value of the combination of wastewater (20, 40, 60, 80 and 100%) and MPs (2 mg/l MPs) was found to be 53.72%. Chronic bioassay concentrations were selected as 1/10 th and 1/20 th of the LC50 for wastewater alone and for the combined treatment [2 mg/l MPs + wastewater (1/10 th and 1/20 th of the LC50)], respectively. The experiment was performed in triplicate (n = 3) with 15 fingerlings in each replicate for 60 days (Table 1). Thereafter, the surviving fingerlings were transferred to tanks containing dechlorinated water for another 60-day recovery period. In parallel, a control (n = 3) with 15 fingerlings in each replicate was also maintained having de-chlorinated water only.
| Groups | Details of treatments |
|---|---|
| T0 (Control) | Fish feed |
| T1 | 1 mg/l MPs + Fish feed |
| T2 | 2 mg/l MPs + Fish feed |
| T3 | 1/10th LC50 of wastewater + Fish feed |
| T4 | 1/20th LC50 of wastewater + Fish feed |
| T5 (COMB1) | 1/10th LC50 of wastewater + 2 mg/l MPs + Fish feed |
| T6 (COMB2) | 1/20th LC50 of wastewater + 2 mg/l MPs + Fish feed |
Table text extracted from the original PDF; superscript letters are shown inline. The original table image is below.
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Table 1: Experimental design for chronic toxicity bioassay
Table 1: Experimental design for chronic toxicity bioassay
Groups
Groups
Details of treatments T0 (Control)
Fish feed T1 1 mg/l MPs + Fish feed T2 2 mg/l MPs + Fish feed T3 1/10 th LC50 of wastewater + Fish feed T4 1/20 th LC50 of wastewater + Fish feed T5 (COMB1) 1/10 th LC50 of wastewater + 2 mg/l MPs + Fish feed T6 (COMB2) 1/20 th LC50 of wastewater + 2 mg/l MPs + Fish feed n = 3 per replicate per treatment
Histomorphology of kidney
Histomorphology of the kidney was done using the procedure of Humason (1975) [7] . Briefly, kidneys were collected and fixed in 10% neutral buffered formalin for 24-48 h, processed with a gradient of ethanol, cleared in xylene and embedded in paraffin. Serial sections (3-5 µm) were obtained using a rotary microtome, mounted on glass slides and stained with haematoxylin and eosin. Microscopic evaluation and image acquisition were performed using a LEICA DM5000 B light microscope equipped with a digital imaging system.
Organ index of kidney
Organ index of kidney for different groups was determined following the classification system described by Bernet et al. (1999) [3] . For each experimental group, kidneys from 3 fingerlings were analysed and 3 randomly selected histological sections per fingerling were evaluated. Histological lesions were categorised into reaction patterns. Each lesion was assigned a severity score ranging from 0 (Absent) to 6 (Severe) based on the degree of tissue damage, and an importance factor (1-3) was applied according to its pathological relevance. The total histopathological index and the organ-specific index were subsequently calculated using the formula 1, where rp, alt, a, org and w indicate reaction pattern, alteration, score value, organ and importance factor respectively.
Organ index = ∑ ∑ (a org rp alt alt rp × 𝑤 org rp alt )
Statistical analysis
Data were expressed as mean ± standard error. Significant differences among treatment groups were evaluated using two-way analysis of variance (ANOVA) followed by Tukey's honestly significant difference (HSD) post hoc test in Statistix 10.00 software, with significance set at p ≤ 0.05.
Results
Results
Histomorphology of kidney
Normal morphological structure of kidneys of C. carpio consists of renal tubules, glomerulus and hematopoietic tissue. After 15 days exposure, fingerlings belonging to control group showed renal tubules and hematopoietic tissue (Figure 1A). In case of T1 group, the morphological examination of kidneys depicted detachment of renal tubule from basal lamina, paracellular spaces and damaged hematopoietic tissue (Figure 1B). Similarly, fingerlings exposed to T2 group showed detachment of renal tubule from basal lamina, paracellular spaces, vacuolization and damaged hematopoietic tissue (Figure 1C). Likewise, fingerlings reared in T3-T6 groups for 15 days revealed detachment of renal tubule from basal lamina, paracellular spaces, enlarged spaces in lumen, vacuolization and damaged hematopoietic tissue (Figures 1D-G). The results showed increase in severity of damage with increase in the concentration of test chemicals. Similarly, fingerlings belonging to control group after 30 days showed normal renal tubules (Figure 2A). However, fingerlings reared in 1 and 2 mg/l MPs for 30 days revealed vacuolization, detachment of renal tubule from basal lamina, damaged renal tubule and damaged hematopoietic tissue in kidneys of C. carpio fingerlings (Figures 2B-C). Similar alterations were observed in kidney of fingerlings exposed to T3 and T4 groups (Figures 2D-E). Kidney of fingerlings exposed to T5 and T6 groups also indicated alterations such as detachment of renal tubule from basal lamina, paracellular spaces, vacuolization and damaged hematopoietic tissue (Figures 2F-G). Apart from these, severe renal alterations such as tubular necrosis and irregular arrangement of tubular cells, enlargement of Bowman space tissue, collapsed and degenerated glomeruli and enlarged space in lumen were also observed in T5 and T6 groups. After 45 days, normal renal tubules were found in kidney of C. carpio belonging to control group (Figures 3A).
Morphological examination of kidney of fingerlings exposed to different treatment groups i.e. T1-T6 for 45 days exhibited aberrations such as detachment of renal tubule from basal lamina, paracellular spaces, vacuolization, damaged hematopoietic tissue, necrosis, tubular necrosis, irregular arrangement of tubular cells, collapsed and degenerated glomeruli; and enlarged space in lumen (Figures 3B-G). Similarly, normal renal tubules were found in kidney of C. carpio belonging to control group (Figures 4A). Kidneys of fingerlings exposed to different groups i.e. T1-T6 for 60 days showed similar aberrations such as detachment of renal tubule from basal lamina, paracellular spaces, vacuolization, damaged hematopoietic tissue, necrosis, exudate and damaged renal tubules (Figures 4B-G). The results clearly indicated dose and durationdependent effect of LDPE MPs and wastewater on fingerlings. Additionally, the results indicated that the effect of exposure of COMB1 and COMB2 was more severe in kidneys than MPs and wastewater alone. After recovery period, less severe alterations were observed in the kidney of treatment groups indicating incomplete recovery. After recovery period, fingerlings exposed to control, T1 and T2 showed normal structures of kidneys (Figures 5A-C). Similarly, fingerlings exposed to T3 and T4 groups revealed normal histological structure with slight aberrations such as exudate formation, detachment of renal tubule from basal lamina and vacuolization in kidney (Figures 5D-E). Alterations such as damaged hematopoietic tissue, paracellular spaces, exudate formation, vacuolization and damaged renal tubules were observed in T5 and T6 groups after recovery period (Figures 5F-G).
Discussion
Kidney is an essential excretory organ and it plays a pivotal role in maintaining homeostasis (Iqbal et al., 2004) [9] . Kidneys of fish are sensitive to chemical alterations in the aquatic bodies because they are constantly exposed to chemicals dissolved in water (Hussain et al., 2019) [19] . Their roles in excretion and detoxification of toxicants from blood expose them to contaminants (Kim and Kang, 2015) [11] . Kidney plays role in excretion of metabolic waste and selective reabsorption which helps in maintaining volume and pH of various body fluids. Therefore, any alterations in histomorphology of kidney could lead to corresponding alterations in behavioural and physiological parameters (Ortiz et al., 2003) [18] . Histopathological alterations such as destructive renal tubules, necrosis and shrinkage of renal cortex were observed in the kidney of fish. Histological effects of 1, 3 and 5% of LDPE and polyethylene terephthalate MPs on kidneys of Oreochromis niloticus were studied by Ali et al. (2023) [1] . In a study conducted by Hamed et al. (2021) [6] , effects of different concentrations of MPs (1, 10 and 100 mg/l) were examined on O. niloticus juveniles for 15 days and 15 days post-exposure. Concentration dependent histopathological alterations such as vacuolated tubules, glomerular atrophy, increase in inter-tubular space, inflammatory cells, congestion of blood capillaries, loss of basophilic cytoplasm, shrinkage of convoluted tubules, deformation, vacuolization and fatty tubules were observed in kidneys of experimental groups. The results also showed alterations in kidney even after post-exposure. The results suggested that MPs cause range of alterations from biochemical alterations in single cells to lesions in entire tissue affecting vitality of fish. The degree of histological alterations in kidneys of fish O. niloticus obtained from a polluted site (downstream of Chakbandi drain) was observed by Shahid et al. (2022) [22] . Histopathological alterations such as degenerated renal tubules, necrosis, delineated lymphocytic infiltration and obliteration in Bowman's space were observed in kidneys of MPs-exposed groups. Similarly, the effects of water pollution on kidneys of fishes (Wallago attu and Cirrhinus mrigala) collected from Chenab River were studied by Hussain et al. (2019) [8] . Alterations such as dilation of glomerular capillaries, hyperaemia, degeneration of renal tubule cells, glomerulonephritis, absence of Bowman's space, vacuolization, necrosis, degeneration, hypertrophy and shrinkage of glomeruli were observed in sections of kidneys. However, fish belonging to control group showed normal histoarchitecture of kidneys. In support of the findings of the current study, histopathological alterations such as necrosis, vacuolization, hypertrophy, degeneration, dilation of glomerular capillaries and contraction of glomeruli were observed in kidneys of Channa punctatus exposed to different concentrations of wastewater (60-100%) from Tung Dhab Drain for 15 and 30 days by Kaur and Dua (2014) [10] . Zhu et al. (2019) [28] conducted the experiment to study the effects of different concentrations of polystyrene MPs (500, 1000 and 2000 µg/g) on kidneys of Oreochromis melastigma for 10 weeks. The alterations such as glomerulopathy and nephrogenesis were observed in kidneys of O. melastigma exposed to PS MPs. However, the alterations might have occurred due to leaching of additives from MPs.
Conclusion
The present study showed significant alterations such as detachment of renal tubule from basal lamina, paracellular spaces, vacuolization, damaged hematopoietic tissue, necrosis and exudate in kidneys of fingerlings belonging to treated groups. Likewise, organ index of kidney of different groups also showed a significant (p ≤ 0.05) increase in comparison to the control group except fingerlings treated with 1 mg/l MPs. Moreover, the results showed incomplete recovery in different groups compared to control group except 1 mg/l LDPE MPs. The results indicated that the combination of LDPE MPs and wastewater may impact kidneys of C. carpio fingerlings in a duration-and dosedependent manner with limited reversibility following exposure, highlighting potential long-term risks to aquatic health.
Figures and tables from the paper
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