The state of the system of antioxidant protection of the body of dogs during toxocariasis invasion

Among the invasive diseases of dogs, the most common in our country and abroad is gastrointestinal helminthia-sis, among which the leading place is occupied by toxocarosis. Adult toxocara cause an intestinal form of the disease, and the larvae – a visceral one. In the process of migration and vital activity, toxocar larvae cause severe damage to the body of dogs up to death. The work aimed to determine the effect of toxocariasis on the enzymatic and non-enzymatic links of the system of antioxidant protection of the dog's body. Twelve two-to four-month-old dogs were used for experimental research


Introduction
Among the invasive diseases of dogs, regardless of the region, season, age, and breed, helminthiasis of the alimentary canal is the most frequently registered.It is worth noting that toxocariasis in dogs is an essential veterinary medicine issue requiring serious attention from pet owners (Schnieder et al., 2011;Ketzis & Lucio-Forster, 2020;Yuan et al., 2024).This disease is caused by parasitic worms of the genus Toxocara, specifically Toxocara canis, one of the most common helminths infecting dogs.Infestation can have severe consequences for the animal's health and pose a danger to humans, as some of its forms can be transmitted through the feces of infected dogs (Nijsse et al., 2015;Zwickl et al., 2018;Dantas-Torres, 2020).Since parasites can remain in the body for a long time without showing any apparent symptoms, regular preventive examinations and deworming are crucial to maintaining dogs' health and preventing the spread of infestations (Othman, 2011;Zheng et al., 2019;Jenkins, 2020;Chidumayo, 2020).
Toxocariasis is characterized by a variety of disease symptoms, ranging from mild symptoms such as indigestion to severe complications, including growth retardation in puppies and dysfunction of internal organs (Zheng et al., 2021;Mubarak et al., 2023;Ng-Nguyen & Nguyen, 2024).In the case of a significant intensity of invasion, adult parasites cause inflammation of the mucous membrane of the small intestines, stomach, and bile ducts of the liver and pancreas (Jenkins, 2020;Xu et al., 2022;Li et al., 2022;Sieng et al., 2023).Toxocars release waste products, which, being absorbed into the blood, cause general intoxication of the body, which further contributes to the formation of a large number of reactive oxygen species and the strengthening of lipid peroxidation processes (Said et al., 2018).
Oxidative processes that occur in the presence of oxygen are the most important energy source.When the mechanism of biological oxidation is disturbed, these processes acquire an uncontrolled chain character, forming active forms of oxygen, nitrogen, and other free radicals (Martyshuk & Gutyj, 2019;Grymak et al., 2020).
Oxygen, in particular its active free radical forms (superoxidanion radical, hydroxyl radical, hydrogen peroxide), begin to oxidize the structures of the cell itself.Susceptible to such attacks are unsaturated lipids, phospholipids, which are part of cellular and intracellular membranes, proteins and amino acids, DNA molecules that cause mutations, hereditary defects, oncological diseases, diseases of the cardiovascular system, and destruction of cells at the molecular level (Shcherbatyy et al., 2019;Slivinska et al., 2020).
Maintaining the concentration of active forms of oxygen in cells at a relatively stable low level is carried out by a multi-component system of regulation of oxygen activation processes, which is necessary for the organism's normal functioning in aerobic conditions.The body is characterized not so much by a state of complete stability but by a state of dynamic equilibrium; when any deviations arise in the body due to various reasons, the body tries to compensate and return to a particular average value (Vasylyev et al., 2021;Vyslotska et al., 2024).
Excessive production of activated oxygen-and nitrogen-containing metabolites or a malfunction of the antioxidant system leads to the development of oxidative stress, which can result in the death of body cells (Slobodian et al., 2021;Gutyj et al., 2022;Verveha et al., 2023).
In particular, the issue of the influence of toxocars and their by-products on one of the essential protective systems of the body -the antioxidant system, whose task is to maintain a balance between the intensity of radical formation and the needs of the body in the physiological and biochemical aspects of the action of oxygen radicals and their derivatives -remains unsolved, namely synthesis of biologically active substances and regulation of membrane permeability.
Understanding these aspects will help dog owners and veterinary professionals better manage this problem and ensure appropriate pet health and well-being.
The aim of the research: to find out the effect of toxocariasis on the enzymatic and non-enzymatic links of the system of antioxidant protection of the dog's body.

Materials and methods
The work was performed at the Department of Parasitology and Ichthyopathology of Stepan Gzhytskyi National University of Veterinary Medicine and Biotechnologies Lviv.Twelve two-to four-month-old dogs were used for experimental research, and two groups of six animals each were formed: control and experimental.Puppies of the experimental group were experimentally infected with the causative agent of toxocarosis at a dose of 5,000 invasive T. canis eggs per kg of body weight.Puppies of the control group were clinically healthy.
All animal manipulations were carried out in accordance with the "General Ethical Principles of Animal Experiments" (Ukraine, 2001) and the provisions of the "European Convention for the Protection of Vertebrate Animals", which are used for experimental and other scientific purposes (Strasbourg, 1985).

Results and discussion
When researching the catalase activity of blood serum of dogs in the control and experimental groups, it was established that it was 0.210 ± 0.010 mg H2O2 in the control group and 0.207 ± 0.004 mg H 2 O 2 in the experimental group, respectively (Fig. 1).After infesting the animals of the experimental group with the causative agent of toxocarosis, a probable decrease in catalase activity was established on the 7th day of the experiment, respectively, by 23 % compared to the indicators of the control group.On the 14th day of the experiment, the activity of catalase in the blood serum of dogs of the experimental group decreased to 0.152 ± 0.010 mg H2O2.In contrast, this indicator fluctuated within the range of 0.212 ± 0.009 mg H 2 O 2 in the control group.On the 21st and 28th days of the experiment, the enzyme activity continued to decrease reliably in the blood serum of animals of the experimental group.In contrast, compared to the control group, it decreased by 40.5 and 51.9 %, respectively.
On the 35th day of the experiment, a slight increase in catalase activity was found in the blood serum of the dogs of the experimental group compared to the previous day.However, compared to the indicators of the control group, this activity was lower by 46.2 %, respectively.
When studying the activity of superoxide dismutase in the blood serum of experimental animals, it was established that at the beginning of the experiment, the activity of the enzyme was 15.82 ± 0.09 in the control group and 15.87 ± 0.10 c.u./mg of protein in the experimental group (Fig. 2).After infesting the dogs of the research group, a slight increase in enzyme activity was found on the first day of the experiment up to 16.15 ± 0.14 c.u./mg of protein.Subsequently, a probable decrease in the activity of superoxide dismutase was established in the blood of the dogs of the experimental group.Thus, on the 7th and 14th day of the experiment, the activity of the enzyme in the blood serum of the dogs of the experimental group decreased by 6.7 % (Р < 0.001) and 12% (Р < 0.001), respectively.On the 21st day of the experiment, the activity of superoxide dismutase in the blood serum of dogs of the experimental group was 12.38 ± 0.18 c.u./mg protein.In the control group, it was 16.02 ± 0.14 human units/mg protein, respectively.It is worth noting that the lowest activity of the studied enzyme was in the blood of dogs of the experimental group on the 28th and 35th day of the experiment, where compared to the control, this indicator decreased by 33.3 % (Р < 0.001) and 30.9 % (Р < 0.001), respectively.The state of the glutathione link of the system of antioxidant protection of the dog's body was studied by indicators such as glutathione peroxidase, glutathione reductase, and reduced glutathione.Regarding the study of the activity of glutathione peroxidase, it was found that it fluctuated within the limits of physiological values in the blood of dogs of the control and experimental groups.After infesting the dogs of the experimental group with the causative agent of toxocar-osis, a probable decrease in enzyme activity was established throughout the experiment.Only on the first day of the experiment, the enzyme activity in the blood of the experimental group of animals was 18.25 ± 0.13 μmol NADPH2h/mg protein.In contrast, this indicator was 17.90 ± 0.15 μmol NADPH 2 h/mg protein in the control group (Fig. 3).On the 7th day of the experiment, an increase in the activity of glutathione peroxidase in the blood serum of the When studying the activity of glutathione reductase, it was found that in the blood serum of dogs of the control and experimental groups, this enzyme at the beginning of the experiment fluctuated between 6.40 ± 0.02 and 6.38 ± 0.03 μmol NADPH 2 h/mg of protein (Fig. 4).The activity of glutathione reductase in the blood serum of dogs of the control group during the entire experiment ranged from 6.37 to 6.41 μmol of NADPH 2 h/mg of protein.A decrease in the activity of this enzyme was found in a research group of dogs after being infected with the causative agent of toxocarosis.A probable reduction in the activity of glutathione reductase in the blood of the experimental group of animals was established from the 7th day of the experiment, where, compared to the control group, it increased by 6.3 %, respectively.On the 14th day of the experiment, the activity of the enzyme in the blood serum of the infested animals was 5.79 ± 0.04 μmol NADPH2h/mg protein, and on the 21st day -5.41 ± 0.03 μmol NADPH 2 h/mg protein.Compared with the control group's parameters, the enzyme activity decreased by 9.1 and 15.3 %, respectively, during the indicated research periods.
The lowest activity of glutathione reductase was in the serum of dogs of the experimental group on the 28th day of the experiment, where it was 4.99 ± 0.04 μmol NADPH2h/mg protein.In contrast, in the control group, this indicator was 6.41 ± 0.03 μmol NADPH 2 h/mg of protein.
On the 35th day of the experiment, a slight increase in the activity of glutathione reductase was established to 5.05 ± 0.04 μmol NADPH 2 h/mg of protein; however, compared to the control group, the activity of the enzyme was lower by 21.1 % (Р < 0.001), respectively.
The content of reduced glutathione in the blood of dogs infected with the causative agent of toxocarosis is shown in Fig. 5.It was established that the reduced glutathione content in the blood of dogs of the control group during the entire experiment ranged from 0.44 ± 0.02 to 0.46 ± 0.02 mmol/L.In the blood of the experimental group on the first day of the experiment, an increase in the content of reduced glutathione up to 0.49 ± 0.03 mmol/L was established.Starting from the 7th day of the experiment, the content of reduced glutathione in the blood of the experimental group gradually decreased, and on the 14th and 21st days of the experiment, respectively, was 0.36 ± 0. On the 35th day of the experiment, the reduced glutathione content in the blood of dogs in the experimental group was 0.32 ± 0.02 mmol/L.In contrast, this indicator was significantly higher in the control group and was 0.46 ± 0.03 mmol/L.Therefore, based on the research conducted, it was established that during the clinical manifestation of the disease, toxocara secretes metabolic products that contribute to the significant formation of free radicals, which, in turn, suppress the activity of the antioxidant defense system in the blood serum of the dogs of the research group.

Conclusions
Under the conditions of experimental toxocarosis invasion, a decrease in the activity of the enzyme link of the system of antioxidant protection of the dog's body was established, as indicated by a reduction in the activity of catalase by 51.9 %, superoxide dismutase by 33.4 %.The development of toxocariasis in dogs is also accompanied by depletion of the glutathione-dependent link of the antioxidant defense system.In infected dogs, a decrease in the content of reduced glutathione in their blood was established by 31.1 % (Р < 0.01), glutathione peroxidase activity -by 26.6 % (Р < 0.001), glutathione reductase activity -by 22.2 % (Р < 0.001).** ** ** *

Fig. 2 .
Catalase activity in the blood serum of dogs infected with the causative agent of toxocarosis, mg Н 2 О 2 (М ± m, n = 6); a -control; b -experimental a b The activity of superoxide dismutase in the blood serum of dogs infected with the causative agent of toxocarosis, c.u./mg protein (М ± m, n = 6); a -control; b -experimental

Fig. 4 .
experimental group was established by 7.2 % compared to the control.It is worth noting that the lowest activity of the enzyme was on the 28th day of the experi-ment in the blood serum of the experimental group, where it was 12.97 ± 0.14 μmol NADPH 2 h/mg of protein, which was 26.6 % (Р < 0.001) lower than the indicators control group.a b Fig. 3.The activity of glutathione peroxidase in the blood serum of dogs infected with the causative agent of toxocariasis, μmol NADPH 2 h/mg protein (M ± m, n = 6); a -control; b -experimental a b Glutathione reductase activity in blood serum of dogs infected with the causative agent of toxocarosis, μmol NADPH 2 h/mg protein (M ± m, n = 6); a -control; b -experimental 02 and 0.34 ± 0.02 mmol/L.The lowest content of the studied indicator was on the 28th day of the experiment, where, compared to the control group, it decreased by 31.1% (Р < 0.01), respectively.The content of reduced glutathione in the blood of dogs infected with the causative agent of toxocarosis, mmol/L (M ± m, n = 6); a -control; b -experimental