Hemato-biochemical alterations in fish – Cyprinus carpio L. after exposure to ‘Derisom’ a botanical pesticide

Shoeiba Tasneem

Department of Zooogy, School of Biological Sciences, Univesity of Kashmir, Srinagar, Jammu & Kashmir, India, 190006

Corresponding Author Email: shoeiba.tas@uok.edu.in

DOI : https://doi.org/10.51470/ABF.2026.5.1.08

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Introduction: Since few decadesfreshwater aquatic ecosystems are facing the threat of harmful effects of synthetic pesticides which is creating a major problem to non-target organisms especially fish, which constitutes the major protein source for humans [1]. Much research has been done on plants for the production of environment safe pesticides that can be used both in field and in storage of grains. In many tropical countries there is a centuries old traditional practice of applying botanical material to crops as to control pests [2]. Studies have proven that plants possess a number of secondary metabolites some of which possess toxic effects, growth regulating effects and anti-feedant effects [3].

                                      One of the most promising natural compounds known to all since several years is azadirachtin, an active compound extracted from the nee tree (Azadirachtaindica), whose  pesticidal, antiviral, antibacterial and antifungal properties have been established[4], [5], [6]. Like the neem based pesticide a new and novel biopesticide – Derisom has been manufactured by agrilife private limited, Hyderabad. The active component of the biopesticide is an alkaloid – karanjin. The biopesticide Derisom is equally compatible with the modern synthetic pesticides. Derisom is used in all types of crop fields because of its effectiveness against wide range of pests. It is well know that plant based pesticides are eco-friendly and less harmful to the environment. They are degraded easily and do not form residues in the environment.

                              Fish forms an ideal model for toxicology research as it acquires the final trophic level in any given aquatic ecosystem and is very sensitive to any physico-chemical change in the water quality. Contamination of an ecosystem by pesticides causes imbalance and hazard to health of fishes.  Fish forms an important source of protein for human consumption [7]. The common carp Cyprinus carpio is an economically and commercially important fish species cultured worldwide. It is regarded as an important species to be cultured in integrated rice-fish farming system in India and many other countries. The fish Cyprinus carpio is sturdy, easily adapted and maintained in lab conditions, hence it has been used in the present study.

                                 The haematological techniques are considered as important markers and tools in assessing fish health, environmental monitoring and toxicity. Haematological profile (RBC count, WBC count, Hb, PCV, MCV, MCH, MCHC) of a fish acts as an important parameter to detect the physiological response to a contaminated environment [8]. Any kind of significant variations in the morphology and total RBC count,  acts as a major and reliable indicator of stress[9], [10], [11].

                                     Till date, extensive literature is available on the toxixicty and sub-lethal effects of neem based pesticides on various fish species. Studies relevant to the effect of biopesticide- Derisom on haematological profile in fish Cyprinus carpio has not been studies so far. Hence, the present study is conducted to elucidate the sub-lethal exposure of fish Cyprinus carpio to biopesticide-Derisom and analyze variations in haematological parameters (total RBC count, WBC count, Hb, PCV, MCV, MCH and MCHC), comparing them later on with the haematology profile of healthy fishes of the control group.

Materials and methods: The fish common carp – Cyprinus carpio weighing 28.53 ± 1.79 gm and measuring 14 ± 0.83 cm were obtained from a nearby fish farm. The fishes were transported to the Department’s animal house facility in well-aerated tanks. The fishes were then acclimatized for a period of one month in well aerated tanks of 500L capacity. Fish were fed twice daily with commercially available fish feed pellets and de-chlorinated tap water was renewed on a daily basis. The 96 hrs LC 50 value for the biopesticide – Derisom was found to be 2.8 ppm. Fishes were exposed to the sub-lethal concentration of Derisom 0.28ppm for a period of 21 days. Control group was also maintained simultaneously. The experiment was carried out with six replicates. The fishes were fed twice daily during the sub-lethal exposure period and the concentration of Derisom was renewed in a semi-static method.

Estimation of Haematological profile: After successful completion of sub-lethal exposures i.e., 24hrs, 7 days, 14 days and 21 days blood was collected from both control and exposed groups separately in heparinised syringes through caudal vein puncture. Blood was then immediately used for the haematological analysis.  Total RBC count and total WBC count  was measured using Neubaur haemocytometer for which the blood was diluted (200 times for total RBC count and 50 times for total WBC count) with Diace solution containing brilliant cresyl blue 0.1 g, sodium citrate 3.8 g, and formaldehyde 37% 0.2mL in 100mL distilled water [12]. Haematocrit or Packed cell Volume (PCV %) was measured using    micro haematocrit capillary tubes after centrifugation (2500 rpm for 5 min).  Haemoglobin concentration (Hb: g/dl) was measured using cyanomethemoglobin spectrophotometrically. Mean Corpuscular Volume (MCV), Mean Corpuscular Haemoglobin (MCH) and Mean Corpuscular Haemoglobin Concentration (MCHC) were calculated using the standard formulae as follows [12].

MCV (fl/cell) = PCV (%) ×10 cubic microns/RBC (10⁶/µl). 

MCH (pg/cell) = Hb (g/dl) × 10 cubic microns/RBC (10⁶/µl),  

MCHC (g/dl) = Hb (g/dl) ×100ml/PCV (%).

Statistical analysis was performed by one way ANOVA at 5% significant level. Two tests (Tukey & LSD – Least Significance Difference) were performed to compare the significant differences using SPSS V.16 and Graph Pad Prism version 5.

Results: The various haematological parameters estimated in fish Cyprinus carpio on sub-lethal exposure to Derisom are presented in Table: 1.

The total RBC count showed significant changes in the blood of fishes during the sub lethal exposure to Derisom when compared to the control group fishes. The total RBC count increased after 24 hrs exposure as compared to the control group. The total RBC count remained almost equal to the control group after 7^th day of exposure, the value decreased significantly after 14^th day and the least value was recorded on the 21^st day. The total WBC count significantly increased as the days of exposure was increased. The total HB concentration remained almost equal to the control group after 24hrs exposure. The Hb concentration decreased significantly as the exposure period increased. The PCV value showed an increase after 24 hrs of exposure as compared to the control group, the PCV value decreased significantly after each consecutive exposure period.

                      The blood indices parameter MCV value remained similar to the control value after completion of 24 hrs and 7 days. MCV value significantly increased after 14^th days and was recorded as the highest value after 21^st day. The blood indices parameter MCH decreased significantly after 24 hrs exposure, remained equal equal to control group after 7 days exposure period. The MCH value significantly increased after 14^th day and 21^st day. The blood indices parameter MCHC decreased significantly after 24hsrs exposure, the value remained almost equal to the control group value after 7days, 14 days and 21 days.

Discussion: The present study clearly shows significant alterations in haematological profile of Cyprinus carpio on sub-lethal exposure to Derisom. Altered haematological status of a fish species indicates the toxic impact of xenobiotics on its health[13]. Blood parameters generally change in the presence of stressors and pollutants[14]. Our findings presented in this article provide a proof that long-term exposure of fish species to botanical/plant based pesticides cause significant deviations in haematological profile [15].  Similar changes in total RBC count and Hb concentration were also reported by other researchers [16]. Another study also showed similar decreasing trend in total RBC count in Cirrhinus mrigala exposed to sub-lethal concentrations of dyes[17]. A decrease in PCV is directly proportional with the decrease in total RBC count, as reported in the present study. Similar kind of variations in haematological profile were reported by researchers in Coho salmon and in Anabas testudineus [18], [19]. Literature shows that total RBC count and packed cell volume values decreased significantly in Cyprinus carpio exposed to combined heavy metals[20]. Few other researchers[21] proved that malachite green is also able to produce similar kind of haematological variations in Nile tilapia on sub-lethal exposures. Few reports support the present findings wherein the haemoglobin concentration decreased in Indian carps, Labeo rohita and Catla catla exposed to sub-acute concentrations of Cr and Cd [22], [23]. A significant reduction in total RBC count and Hb content was also reported in Labeo rohita exposed to aquatic contaminants[24], [25].

                         Total RBC count and Hb concentration may act as important markers which help in estimating the overall health condition of fish. Any significant deviations in these values may lead to conditions such as partial anaemia, hypoxia, erythrocytopenia, decreased haemopoiesis or damage in the haematopoietic tissues. Other reasons for decrease in Hb content reflects the anaemic state of the fish which may be a result of iron deficiency and its consequent decreased utilization for Hb synthesis [22]. The above mentioned deviations in the haematological profile may lead to the structural damage in RBC membranes resulting in haemolysis and impairment in haemoglobin synthesis, stress related release of RBCs from the spleen and hypoxia condition in fish. Anaemic condition might also result due to haemo-dilution caused by imbalanced osmoregulation across gill epithelium. Condition like naemia may affect the general well-being of the fishes.  The overall reduction in total RBC count and Hb content may be the result of damage and destruction of red blood cells[26], [27].

 Present study shows a significant increase in total WBC count throughout the sub-lethal exposure period. This significant increase in total WBC count may be considered as an adaptive immunological defence mechanism, leucocytosis or lymphocytosis against sub-lethal exposure to Derisom. Similar deviations in haematological profile was observed by other researchers[28] in Poecilia reticulate exposed to methyl red. Some other researchers recorded Leucocytosis in Cirrhinus mrigala exposed to various pollutants [29].

 Red Blood cell indices – mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC) are very sensitive indicators of overall fish health and significant deviations from their normal range may cause reversible changes in the homeostatic environment of fish. Variations in these blood indices are directly proportional to the total RBC count, haemoglobin concentration and packed cell volume. Results of the present study showed a significant variation in these indices in Cyprinus carpio during sub-lethal exposure to Derisom. Similar responses were also recorded in common carp and other freshwater fish on exposure to various pesticides [30], [31], [32], [33].

Conclusion: Water borne pollutants like pesticides may alter the haematological parameters in fish. Survival of fish in an aquatic ecosystem depends not only on their biological state but on the quantity and type of pollutants. The results obtained in the present study clearly show that Derisom is moderately toxic to Cyprinus carpio. Exposure of common carp to sub-lethal concentrations of Derisom resulted in significant haematological alterations. Hence the above mentioned changes in haematological profile suggest that the fish during sub-lethal exposure to Derisom are facing some metabolic problems which are not normal. The decreasing values of haematological parameters such as total RBC count, haemoglobin concentration and PCV in the exposed fish may act as indicators of stress. The increased WBC value indicates the immunological response of fish against the biopesticide Derisom. Therefore it may be concluded that the usage of pesticides both synthetic and botanical, either in the agricultural fields should or aquaculture ponds should be monitored well as they may be a threat to both aquatic fauna and flora including humans.

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