INTRODUCTION:

Different kinds of keratoconjunctivitis in animals, and in particular, in cattle, are quite common and often have a massive character, affecting at the same time from 50 to 90% of the livestock. In addition, this ophthalmological pathology causes huge economic damage to farms, which is expressed in a decrease in milk productivity by an average of 50% and an average daily increase in live weight by 30-40%. On average, 25-30% of sick animals become blind, and the same number lose their sight by 50% or more^1-3.

The conjunctiva and cornea are the first to be exposed to harmful environmental factors, and as a result, inflammatory processes develop in the form of keratitis and conjunctivitis, but more often the process captures both membranes together and proceeds in the form of conjunctivokeratitis. These diseases are called massive conjunctivokeratitis due to the disease spread in a large number of animals, which is most often observed in cattle^4-6. The most common forms are infectious and invasive keratoconjunctivitis. Infectious keratoconjunctivitis (IKC) is an acute inflammation of the mucous membrane of the eye and cornea, manifested by lacrimation, redness of the mucous membrane of the eye, photophobia, serous purulent discharge, opacification, and ulceration of the cornea, eyeball deformation, and loss of vision^7,8. Animals of all ages are affected, but calves and young animals from 5 months to 2 years are more susceptible to the disease^9,10.

IKC is most commonly caused by the bacteria Moraxella bovis and Moraxella bovoculi. As a secondary infection, streptococci and staphylococci are often found, complicating the infectious process. The cause may also be various mechanical, physicochemical, infectious, invasive effects, as well as symptomatic effects, developing against the background of various infectious diseases^11-13. Contributing factors are also the movement of sick animals from one farm to another, which in turn contributes to the widespread occurrence of the disease. Infection occurs through the discharge from the eyes and nose both mechanically (with close contact between animals) and with the help of biological vectors (flies). Besides, the source of the spread of infection is the general inventory and elements of outbuildings (feeders, drinkers, walls)¹⁴.

Currently, due to the deterioration of the environmental situation, it is very important to create new phytopreparations that, compared with synthetic ones, have not only symptomatic but also more effective pathogenetic effects^15,16. In the comparative aspect, more than 30% of the medications used in modern medicine are obtained from plant raw materials^17-26. Phytopreparations are used to treat all types of diseases, but in veterinary ophthalmopathology, unfortunately, they are still not used enough. Unlike synthetic medications, phytopreparations have a wider range of pharmacotherapeutic effects, are non-toxic or low-toxic, which makes it possible to use them for a longer time without the risk of any pronounced side effects^27,28.

The study was aimed at development of multicomponent ointment LLP KazSRVI and phyto-ointment KerKon and study of their comparative pharmacotherapeutic effectiveness in IKC in cattle. Objectives of the study:

1. To identify the pharmacotherapeutic effectiveness of the developed eye ointments on sick animals in a comparative aspect;

2. To determine the content of some biochemical and immunological parameters of blood and lacrimal fluid in the treatment of keratoconjunctivitis in cattle.

MATERIALS AND METHODS:

For the preparation of phyto-ointment KerKon, we used the following medicinal plants: drug eyebright grass (Yerhasia officinalis L.), plantain leaves (Plantago stepposa), thyme grass (creeping thyme, Thymus serpyllum L.), and chamomile flowers (Matricaria chamomilla L.). Drying of medicinal plant raw materials was carried out with artificial (air-solar) heat on racks. The average duration of drying was 5-7 days. The quality was determined by special research methods, which include raw material preparation, sampling, and analysis of samples. The authenticity of medicinal plant raw materials was determined by generally accepted pharmacological research methods^29,30.

In the course of our work, we made officinal mixtures (the size of the plant crushed parts did not exceed 4-6 mm), which were brewed or infused with water in a ratio of 1: 10 (1 part plant, 10 parts water) or prepared as an alcohol extract (dry extract) before use. The phyto-ointment was made in the following ratio: 1 part by weight of dry and finely ground extract from the collection of harvested plants and 9 parts by weight of medical lanolin. The composition of the multicomponent ointment KazSRVI includes vaseline, with additional content of ofloxacin, a 4% aqueous solution of gentamicin sulfate, with the following ratio of the constituent components: ofloxacin 8.0%; gentamicin sulfate (4.0% aqueous solution) 2.0% and pharmaceutical vaseline for the rest of the composition. Preclinical and clinical trials were conducted in the conditions of Baiserke-Agro LLP in the Almaty region. The object of our study was cattle of the Holstein-Frisian breed, as well as mixed cattle of both sexes and different ages. The diagnosis of IKC was carried out taking into account the clinical manifestation of the disease, the epizootic situation, and the laboratory studies.

The study of the distributing effect of manufactured ointments on the mucous membrane of the eye. For this purpose, a group of healthy calves was formed. Experimental animals, in the amount of 5 heads, had the manufactured ointments placed in the lower conjunctival sac. After that, observations were made of changes in the conjunctiva and cornea of the eye. The second eye was left as a control sample. Examination of the mucous membrane was carried out after 5 minutes, then after 30 minutes, and then after 1, 2, 3, 6, 12, and 24 hours.

During the treatment, all animals were monitored. At the same time, attention was paid to changes in the mucous membrane of the eye and cornea, as well as to the time of recovery.

When setting up the research and production experiment on the principle of analogs, 4 groups of animals (15 heads each) with characteristic signs of a pathological process were formed: The 1st experimental group was treated with the use of multicomponent ointment KazSRVI, the 2nd group was treated with the use of phyto-ointment KerKon, the 3rd group was treated with the use of a combined scheme of ointments and the 4th was the control group, which was treated with the use of 1% ocular tetracycline ointment.

The treatment of sick animals was carried out in compliance with the rules of asepsis and antiseptics. First of all, before applying ointments, the affected eyes were thoroughly treated with a solution of furacillin (1:500-5,000.0) or 0.1% potassium permanganate solution (K₂MnO₄). The ointments were used as follows: the lower eyelid was pulled down and a small (0.3 – 0.5 gr.) amount of ointment was placed with a glass spatula in the area of the lower transitional fold (the conjunctiva fornix). The glass stick (spatula) was immersed flat behind the lower eyelid. After the animal closed its eye, the glass stick was removed. The remnants of the ointment on the edges of the eyelids were removed with a sterile swab, and the ointment remaining in the conjunctival bag was distributed by light stroking on the lower eyelid. For therapeutic purposes, ointments were placed under the lower eyelid twice a day, and in the case of the combined scheme, they were alternated.

During the experiment, the animals were monitored. All changes were recorded in the journal. At the same time, blood and lacrimal fluid were collected before the start of treatment and on the 5th, 10th, 15th, and 20th days after the start of treatment.

The blood taken was examined for some biochemical and morphological parameters. Biochemical and immunological studies were carried out in the scientific laboratories of the Department of Clinical Veterinary Medicine of the Kazakh National Agrarian Research University (KazNARU), the Kazakh Scientific Research Veterinary Institute LLP, and the laboratory of the Kazakh-Japanese Innovation Center of KazNARU.

Quantitative values of lysozyme, the bactericidal activity of blood serum (BABS), β-lysine, lysozyme were determined using StatFax 2100 enzyme immunoassay (EIA) reader (USA); EIA automatic analyzer Immulight 1000 (USA), total protein was determined by refractometry; and protein fractions by vertical electrophoresis on agar gel. Lacrimal fluid was taken using an eye pipette from the inner corner of the eye. Lactoferrin concentration in the lacrimal fluid was determined by EIA with sets of reagents Vector-Best (Novosibirsk). The optical density was measured using a Multiskan photometer (Labsystems, Finland) at a wavelength of 492nm.

The obtained digital data were processed by the constant method of variational statistics with the calculation of arithmetic averages and their statistical errors (M ± m), the reliability (P) of the compared indicators was determined by the Student's t-test. The Microsoft Excel statistical analysis package was used for calculations.

RESULTS:

The results of studies on the study of some immunomorphological parameters of animal blood are presented in Table 1.

Studies have established a reduced hemoglobin content in the blood of patients with keratoconjunctivitis of animals. The results showed that the concentration of hemoglobin in the 1st group after the course of treatment had increased by 6.3% compared to the start of treatment, and in the 2nd and 3rd groups by 7.6 and 13.9%, respectively, while in the control group the increase was only 0.8% (^xR<0.05; ^xxR<0.01; ^x ^xx P<0.001).

The results of the studies showed an intensive increase in the quantitative indicators of blood leukocytes in experimental groups of animals. Thus, the concentration of leukocytes in the 1st experimental group after the course of treatment had increased by 9.1% compared to the start of treatment, and in the 2nd and 3rd experimental groups, by 7.6 and 6.6%, respectively, while in the control group the increase was only 1.4% (^xR<0.05; ^xxR<0.01; ^x ^xx P<0.001).

The findings of our study showed that the concentration of erythrocytes in the 1st experimental group after the course of treatment had increased by 4.9% compared to the start of treatment, and in the 2nd and 3rd groups by 5.2 and 9.6 %, respectively, while in the control group the increase was only 2.3% (^xR<0.05; ^xxR<0.01; ^x ^xx P<0.001). Before the appointment of ointments, the content of total protein and its fractional composition in all groups were approximately equal. The quantitative content of total protein in the 1st experimental group of animals after completing the course of treatment had increased by 12.4% compared to the start of treatment, and in the 2nd and 3rd groups by 16.6% and 21.2 %, respectively, while in the control group the increase was only 4.4% (^xR<0,05; ^XXR<0,01; ^x ^xx Р<0.001). There were no significant changes in the α- and β-globulin fractions of proteins in the experimental groups of animals. With the combined use of ointments in the 3rd experimental group of animals, an increase in the level of the γ-globulin fraction of proteins was observed by 2.5% compared to the beginning of treatment. In animals in the control group, the level of γ-globulins tended to decrease by 2%.

The results of the conducted studies showed that before the appointment of treatment, there were no significant differences in BABS between the experimental and control groups. However, in the course of treatment, the experimental groups showed a positive trend towards an increase in the concentration of BABS. The quantitative content of BABS in the 1st group of animals after completing the course of treatment had increased by 5.1% compared to the start of treatment, in the 2nd and 3rd groups by 5.8 and 12.6%, respectively, while the increase in the control group amounted to only 1.5% (^xR<0.05; ^xxR<0.01; ^xxxP<0.001).

The obtained study results reliably indicate a significant increase in the content of lysozyme in the experimental groups of animals. The quantitative content of lysozyme in the 1st group of animals after the end of the course of treatment had increased by 13.6% compared to the beginning of treatment, in the 2nd and 3rd groups by 12.4 and 16.7%, respectively, and in the control group the increase was only 5.4% (^x P<0.05; ^xx P<0.01; ^xxx Р<0.001).

The studies also revealed positive dynamics concerning β-lysine in experimental groups of animals. The concentration of β-lysine in the blood serum of animals of the 1st group after treatment had increased by 17.1% compared to the start of treatment, in the 2nd and 3rd groups by 15.4 and 25.6%, respectively, while in the control group the increase was only 2.4% (^xR<0.05; ^xxR<0.01; ^xxxP<0.001).

Table 1. Results of immunomorphological blood parameters of experimental animals before and after treatment (n=20; M ± m)

Indicators

Groups

1st experimental (Ointment KazSRVI)

2nd experimental

(Kerkon phyto-ointment)

3rd experimental

(KazSRVI + Kerkon)

IV control

(Tetracycline ointment)

Before treatment

After treatment

Before treatment

After treatment

Before treatment

After treatment

Before treatment

After treatment

Hemoglobin, g/l

96.4±

1.01 ^x

102.5±

0.99 ^xxx

96.3±

1.36 ^xx

103.6±

1.38 ^xxx

92.6±

1.29 ^x

105.5±

1.78 ^xx

92.8±

1.24 ^x

93.5±

0.78 ^xxx

Erythrocytes, ^10-12/l

5.13±

0.07 ^xx

5.38±

0.13 ^x

5.18±

0.17 ^xx

5.45±

0.17 ^xxx

5.64±

0.09 ^xx

6.18±

0.13 ^xxx

5.12±

0.09 ^x

5.24±

0.13 ^xx

Leukocytes, 10 ⁹/l

7.76±

0.15 ^x

8,47±

0.06 ^xx

7.67±

0.19 ^x

8.18±

0.15 ^xx

7.16±

0.08 ^x

8.06±

0.05 ^xxx

7.66±

0.08 ^xx

7.77±

0.06 ^x

Total protein, g/l

59.86±

0.43 ^x

67.3 ±

0.62 ^xxx

59.30±

0.98 ^xxx

69.17±

1.35 ^x

58.38±

1.16 ^x

70.75±

1.15 ^x

59.66±

0.53 ^x

62.3 ±

0.62 ^xx

Albumins, %

43.22±

0.23 ^x

44.37±

0.18 ^x

47.54±

1.35 ^xx

47.38±

0.57 ^xxx

45.03±

0.83 ^x

45.07±

0.58 ^x

43.22±

0.23 ^x

44.07±

0.18 ^x

α - globulins, %

13.98±

0.21 ^xx

12.95±

0.14 ^x

13.75±

0.32 ^x

13.86±

0.39 ^x

13.72±

0.59 ^x

13.42±

0.49 ^x

13.88±

0.20 ^x

13.75±

0.14 ^x

β - globulins, %

18.71±

0.22 ^x

18.64±

0.11 ^xx

17.70±

0.59 ^x

17.23±

0.38 ^xx

15.60±

0.56 ^x

15.23±

0.32 ^xx

18.71±

0.21 ^x

18.44±

0.11 ^x

γ - globulins, %

24.09±

0.19 ^xx

24.04±

0.19 ^x

21.01±

0.91 ^x

21.53±

0.50 ^xx

25.65±

1.04 ^xx

26.28±

0.46 ^xxx

24.19±

0.19 ^x

23.74±

0.19 ^x

BABS, %

73.67±

0.87 ^x

77.86±

0.90 ^xx

73.69±

0.93 ^xx

77.95±

1.47 ^xxx

73.25±

0.51 ^x

82.46±

0.87 ^xxx

73.47±

0.77 ^x

74.56±

0.87 ^x

β - lysine, %

12.02±

0.40 ^x

14.08±

0.69 ^xxx

12.15±

0.71 ^x

14.02±

0.30 ^xx

12.23±

0.15 ^x

16.36±

0.34 ^xx

12.09±

0.36 ^x

12.38±

0.49 ^x

Lysozyme, mcg/ml

1.69±

0.10 ^x

1.92±

0.08 ^xx

1.61±

0.12 ^xx

1.81±

0.14 ^x

1.62±

0.13 ^x

1.89±

0.05 ^x

1.67±

0.10 ^x

1.76±

0.08 ^x

Note: the difference in values in the intergroup comparison ^x P<0.05; ^xx P<0.01; ^x ^xx P<0.001; BABS is the bactericidal activity of blood serum.

Table 2. Dynamics of immunomorphological blood parameters in experimental animals (n=10; M ± m)

Groups

Days of the study

Hemoglobin, g/l

Erythrocytes,

10 ¹²/l

Leukocytes,

10 ⁹/l

Total protein, g/l

Albumins, %

BABS, %

β - lysine, %

Lysozyme, mcg/ml

Before treatment

(background)

92.6±

1.29 ^x

5.64±

0.09 ^xx

7.16±

0.08 ^x

58.38±

1.16 ^x

45.03±

0.83 ^x

73.25±

0.51 ^x

12.23±

0.15 ^x

1.62±

0.13 ^x

92.8±

1.24 ^x

5.12±

0.09 ^x

7.66±

0.08 ^xx

59.66±

0.53 ^x

43.22±

0.23 ^x

73.47±

0.77 ^x

12.09±

0.36 ^x

1.67±

0.10 ^x

94.2±

1.32 ^x

5.82±

0.06 ^xx

7.49±

0.07 ^x

62.69±

1.15 ^x

45.01±

0.92 ^x

77.45±

0.81 ^x

14.33±

0.19 ^x

1.70±

0.12 ^x

92.9±

1.18 ^x

5.14±

0.05 ^x

7.70±

0.08 ^xx

59.96±

1.47 ^x

43.16±

0.59 ^x

73.62±

0.80 ^x

12.15±

0.28 ^x

1.68±

0.11 ^x

98.5±

1.30 ^x

6.02±

0.08 ^xx

7.95±

0.10 ^x

67.44±

1.19 ^x

45.05±

0.81 ^x

79.27±

0.64 ^x

15.26±

0.23 ^x

1.82±

0.12 ^x

93.0±

1.44 ^x

5.18±

0.09 ^x

7.72±

0.09 ^xx

60.61±

1.52 ^x

43.86±

0.69 ^x

73.89±

0.83 ^x

12.29±

0.44 ^x

1.70±

0.13 ^x

103.6±

1.49 ^x

6.12±

0.04 ^xx

8.02±

0.11 ^x

69.18±

1.23 ^x

45.06±

0.73 ^x

81.38±

0.91 ^x

16.12±

0.35 ^x

1.86±

0.15 ^x

93.3±

1.28 ^x

5.20±

0.09 ^x

7.74±

0.08 ^xx

61.56±

1.50 ^x

43.96±

0.87 ^x

74.29±

0.85 ^x

12.31±

0.36 ^x

1.72±

0.13 ^x

105.5±

1.78 ^xx

6.18±

0.13 ^xxx

8.06±

0.05 ^xxx

70.75±

1.15 ^x

45.07±

0.58 ^x

82.46±

0.87 ^xxx

16.36±

0.34 ^xx

1.89±

0.05 ^x

93.5±

0.78 ^xxx

5.24±

0.13 ^xx

7.77±

0.06 ^x

62.3 ±

0.62 ^xx

44.07±

0.18 ^x

74.56±

0.87 ^x

12.38±

0.49 ^x

1.76±

0.08 ^x

Note: the difference in values in the intergroup comparison ^x P<0.05; ^xx P<0.01; ^x ^xx Р<0.001; BABS is the bactericidal activity of blood serum.

E is the experimental group; C is the control group

The resistance of the outer membrane of the eye to the damaging effect of various infectious agents is largely determined by the state of local (secretory) immunity. The protective properties of the lacrimal fluid are expressed in mechanical cleansing of the eye surface, as well as in the specific and non-specific antimicrobial protection factors. Lactoferrin is one of the factors of nonspecific secretory immunity of the eyes.

As can be seen from Table 3, the level of lactoferrin in the experimental groups of animals tended to increase constantly. Thus, after 5, 10, 15 days of treatment, the level of lactoferrin in the 1st experimental group had increased compared to the baseline, by 3.9 and 6.8 and 21.6%, respectively; in the 2nd experimental group by 4.2; 8.2 and 19.9%, respectively; in the 3rd experimental group the increase was 5.3; 18.9% and 31.2%, respectively, and in the control group no significant changes occurred (^xR<0.05; ^XXR<0.01; ^xxxP<0.001).

It should be noted that the quantitative content of lactoferrin significantly increased in the 3rd experimental group of animals, where the degree of increase compared to the control group ranged from 18.1 to 26.8% (^x P<0.05; ^xx P<0.01; ^xxx Р<0.001).

Table 3. Dynamics of lactoferrin level in lacrimal fluid in keratoconjunctiva in cattle, mcg/ml, (n=10, M ± m)

Groups	Before treatment	Terms of treatment,
		5	10	15	20
		days after treatment
1st experimental	27.85±0.58 ^x	28.94± 0.55 ^x	29.74± 0.51 ^xx	33.86± 0.92 ^xxx	27.18± 0.61 ^x
2nd experimental	27.59±0.62 ^x	28.75± 0.53 ^xx	29.85± 0.53 ^x	33.09± 0.94 ^xx	27.87± 0.85 ^x
3rd experimental	27.49±0.82 ^x	28.96± 0.63 ^x	32.69± 0.62 ^xx	36.05± 0.96 ^xxx	27.67± 0.79 ^xx
Control	27.47±0.91 ^x	27.49± 0.42 ^x	27.69± 0.65 ^xxx	28.42± 0.97 ^x	27.33± 0.70 ^xx

Note: ^x P < 0.05; ^xx P < 0.01; ^xxx P<0.001 compared to the start of treatment and to the control group.

The next stage of our study was the evaluation of the comparative pharmacotherapeutic effectiveness of the use of ointments in the treatment of bovine IKC by setting up a research and production experiment in farm conditions. Before the start of treatment, all sick animals had a generally depressed state and decreased appetite. Severe bleeding and blepharospasm of the diseased eye were noted. The conjunctiva of the eye was reddened and edematous. Catarrhal purulent exudate outflows were observed from the inner corner of the eye. There were crusts of dried exudate on the edges of the eyelids. The cornea was diffusely clouded, from grayish-white to milky in color (Figs. 1, 2).

In animals of the 1st, 2nd, 3rd experimental groups, a slight improvement in the general condition and nutritional activity, a decrease in swelling, and eyelid soreness were noted before the 5th-6th day of treatment. The conjunctiva of the diseased eye was hyperemic. We noted a decrease in the intensity of catarrhal purulent expirations, as well as the absence of photophobia.

Fig. 1. Catarrhal purulent keratoconjunctivitis of the right eye

Fig. 2. Catarrhal purulent keratoconjunctivitis of the left eye before treatment

Fig. 3. Catarrhal purulent keratoconjunctivitis of the right eye on the 10th-12th day of treatment

Fig. 4. Catarrhal purulent keratoconjunctivitis of the right eye on the 15th day of treatment

On clinical examination on the 8th-10th day, the animals showed some improvement in their general condition, slight pain on palpation, moderate hyperemia of the mucous membrane. The sphericity of the cornea remained unchanged, active corneal clearing had occurred.

By the 12th-13th day of treatment, the animals had a satisfactory general condition and ate willingly. Soreness and blepharospasm had disappeared, weak hyperemia of the conjunctiva of the eye was observed, as well as scanty outflows from the inner corner of the eye. The area of opacity was reduced to the minimum size (Fig. 3).

Complete absence of signs of an inflammatory reaction and recovery occurred on average on the 15th- 16th day after the start of treatment. At the same time, normal nutritional activity was observed in the animals, and their general condition was satisfactory. The cornea of the eyes was smooth, shiny, without signs of damage. The conjunctiva of the eye was pale pink and smooth (Fig. 4).

The comparative pharmacotherapeutic effectiveness of the ointments used in the course of treatment in the context of groups is shown in Table 4.

The pharmacotherapeutic effectiveness can be judged by the following indicators: out of 15 sick animals of the 1st experimental group, 4 heads (26.7%) had recovered by the 10th day from the moment of application of the multicomponent ointment KazSRVI, 10 heads (66.7%) recovered in 11-16 days, 1 head (6.6%) in 17-20 days, 4 heads (26.7%) recovered in the 2nd experimental group by the 10th day from the moment of application of phyto-ointment KerKon, 9 heads (60%) in 11-16 days, and 2 heads (13.3%) in 17-20 days.

In the course of treatment, we observed the greatest therapeutic effect in the 3rd experimental group of animals that used a combined scheme, that is, with the use of multicomponent ointment KazSRVI and phyto-ointment KerKon alternatively. After applying such a combined scheme, by the 10th day from the beginning of treatment, recovery had occurred in 5 heads (33.3%), and full recovery of the majority of sick animals (10 heads, 66.7%) occurred within 11-16 days.

Thus, after the treatment of animals with IKC with the proposed scheme, we achieved a positive effect, and the treatment lasted on average 14-15 days. Therefore, we believe that the daily application of ointments twice a day leads to positive dynamics in the treatment of this disease in animals.

Hence, it should be noted that the most effective was the use of a combined treatment regimen, where the absolute effectiveness in all experimental groups was 100%.

Table 4. Comparative pharmacotherapeutic effectiveness of ointments in the treatment of bovine keratoconjunctivitis (M ± m; n =60)

Indicators	Animal groups
Indicators	1st experimental	2nd experimental	3rd experimental	Control
Number of animals, heads	15	15	15	15
Number of recovered animals, heads	15	15	15	12
in %	100	100	100	80
Including:
Within 6-10 days, heads	4 ± 0.62 ^xx	4 ± 0.54 ^x	5 ± 0.73 ^xxx	-
in %	26.7	26.7	33.3
Within 11-16 days, heads	10 ± 0.93 ^x	9 ± 0.82 ^xx	10 ± 0.91 ^x	3 ± 0.32 ^x
in %	66.7	60	66.7	20
Within 17-20 days, heads	1 ± 0.09 ^x	2 ± 0.08 ^xxx	-	5 ± 0.49 ^x
in %	6.6	13.3		33.3
Within 20 or more days, heads	-	-	-	4 ± 0.56 ^xx
in %			-	26.7
Not recovered, heads	-	-	-	3 ± 0.14 ^x
in %				20
Absolute effectiveness, %	100	100	100	80
Note: ^x P < 0.05; ^xx P < 0.01; ^xxx P< 0.001 are the reliability of indicators.

A different picture is observed in the control group of animals that used the traditional 1% ocular tetracycline ointment, where the therapeutic effectiveness was significantly low compared to the indicators of the experimental groups of animals, and recovery was significantly delayed. Thus, within 11-16 days from the beginning of treatment, recovery occurred only in 3 animals (20%), within 17-20 days, recovery occurred in 5 heads (33.3%), and within 20 or more days in 4 heads (26.7%), while 3 heads (20%) did not recover. The absolute effectiveness in the control group was only 80%.

DISCUSSION:

In the course of the study, we observed low concentrations of blood hemoglobin in all experimental groups of animals. Low hemoglobin content in the blood of animals with keratoconjunctivitis indicates the development of endogenous intoxication of the body, which entails a decrease in the antioxidant capabilities of the body³¹.

The obtained study results showed that the hemoglobin content in the experimental groups of animals after the end of the course of treatment had significantly increased compared to the beginning of treatment in the range from 6.3 to 13.9%, and compared to the control group by up to 12.8%.

The results of the studies showed that in the experimental groups there was an intensive increase in the quantitative indicators of blood leukocytes. The concentration of leukocytes in the experimental groups of animals had significantly increased compared to the start of treatment, on average from 7.6 to 9.1%, respectively, and in the control group, the increase was only 1.4%. An increase in the number of leukocytes in the blood should probably be considered as a compensatory reaction, which quantitatively compensates for the insufficiency of the functional activity of the cellular link of the immune system with a significant antigenic load that occurs with pronounced inflammatory processes on the mucous membranes of the eyes³².

The obtained study results showed that the concentration of erythrocytes in the experimental groups of animals compared to the control group had significantly increased on average from 4.9 to 9.6%, and in the control group the increase was only 2.3%. An increase in the number of erythrocytes in peripheral blood in experimental groups of animals may be associated with a compensatory reaction since erythrocytes take an active part in the transport of microbial antigens as part of immune complexes due to the absorption of immune complexes on the erythrocyte membrane due to specific receptors³³.

Thus, an increase in the quantitative content of hematological indicators characterizes the activation of the immunobiological reactivity of the body, since they provide a better supply of oxygen to tissues, increase the intensity of metabolic reactions and thereby contribute to a faster recovery of animals. Deficiency of total protein in the blood serum of animals predisposes to inflammation of the mucous membranes of the eyes^34,35. As a result of our experiments, we obtained data indicating an increasing content of the protein composition of the blood serum of sick animals during treatment. These changes affect the content of both the total protein and its fractions. The highest total protein content was detected after the use of the combined treatment, where the increase compared to the start of treatment was 21.2%, and in the control group, it was only 4.4%. There were no significant changes in the α- and β-globulin fractions of proteins in the experimental groups. There was only an increase in the level of the γ-globulin fraction of proteins by 2.5% compared to the start of treatment.

The dynamic increase in the amount of total protein in the blood may be associated with an increase in the level of specific blood proteins, immunoglobulins, in response to bacterial infection, which is a consequence of the formation of the primary immune response to the antigens of microorganisms^{32, 33}. The use of ointments developed by us has a beneficial effect on the healing process of keratoconjunctivitis, and the confirming factor is an increase in the humoral links of immunity protection.

The results of the conducted studies showed that under the influence of the ointments used, there was a positive trend towards an increase in the concentration of BABS. The quantitative content BABS in the 1st group of animals after completing the course of treatment had increased by 5.1% compared to the start of treatment, in the 2nd and 3rd groups by 5.8 and 12.6%, respectively, while the degree of increase in the control group amounted to only 1.5%. A significant increase in the concentration of BABS in the experimental groups directly reflects the patterns of the normal reaction of the cattle organism to the introduction of microflora and indicates the tension of the body's defenses³⁶. A decrease in lysozyme activity correlates with a decrease in antioxidant systems (ceruloplasmin). This is explained by the fact that lysozyme, being a natural antioxidant present in the body of animals, is partially spent on the elimination of free radicals, and is also used to participate in the infectious process³⁷.

The obtained study results reliably indicate a significant increase in the content of lysozyme in the experimental groups of animals. The quantitative content of lysozyme in the experimental groups of animals after the end of the course of treatment had increased compared to the beginning of treatment by an average of 13.6 and 16.7%, and in the control group, the increase was only 5.4%.

The studies also revealed positive dynamics concerning β-lysine in experimental groups of animals. The concentration of β - lysine in the blood serum of animals after the end of the course of treatment in the experimental groups had significantly increased compared to the beginning of treatment and was equal from 17.1 to 25.6%, and in the control group, the increase was only 2.4%. An increase in lactoferrin concentration of lacrimal fluid with a maximum on the 15th day of treatment can be regarded as a positive effect, since lactoferrin has not only a direct bactericidal effect, but also an anti-inflammatory effect, inhibiting the inflammatory process due to its binding to bacterial endotoxins and, as a consequence, limiting the production of pro-inflammatory cytokines³⁸.

The level of lactoferrin in the experimental groups of animals tended to increase dynamically significantly in comparison with the control group. Thus, by the 5th, 10th, 15th day of treatment, the level of lactoferrin compared to the baseline in the 3rd experimental group had increased by 5.3, 18.9, and 31.2%, respectively, and there were no significant changes in the control group. In addition, an increase in the concentration of lactoferrin can sufficiently ensure its physiological regulation of granulocytopoiesis and provide a potentiating effect on the bactericidal effect of lysozyme on gram-negative bacteria, since, without lactoferrin, lysozyme has a detrimental effect only on gram-positive bacteria^{39, 40}. Dynamic changes in the concentration of antimicrobial protection factors of the eye mucosa can be highly informative in assessing the effectiveness of the treatment⁴⁰.

CONCLUSION:

Thus, after treating animals with infectious keratoconjunctiva with the proposed scheme, we achieved a positive effect, since the complete recovery of animals occurs in 15-16 days. Therefore, we believe that the daily application of ointments twice a day leads to positive dynamics in the treatment of this disease in animals. The most effective method is the use of a combined treatment regimen with developed ointments.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

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Received on 08.02.2022 Modified on 13.05.2022

Research J. Pharm. and Tech 2023; 16(1):46-54.

DOI: 10.52711/0974-360X.2023.00009