Diagnostic value of integral indexes of intoxication to assess the risk of complications development in gastric and intestinal diseases of calves

. The assessment of the intoxication degree and the state of general organism resistance was carried by identifying "hematological indices" showing the connections between various cell classes of leukocyte formula. Comparison of integral indices of intoxication has recorded “slight degree of intoxication of the organism” in sick calves in 47.6% of cases. In the case of “moderate severity of organism intoxication”, the tension in the hemopoiesis system was recorded in 85.7% of cases, and the imbalance in the reactions of the cellular and humoral link of immunity – in 48.1% cases. For animals with a “severe de gree of organism intoxication” imbalance in the links of immunity was determined in 90.9% of cases. According to the results of the determination of LII, LSI and LI, 31.6% calves were identified as having a risk of gastrointestinal complications development, whereas according to the results of CBC only 9.5% animals were classified as a “risk group”. dangerous parasitic and non- communicable


Introduction
In the nosological structure of diseases of young cattle diseases of digestive organs take the main place . According to veterinary reports, an increase in morbidity by 1.5-2% is registered annually, and this trend is steady [1,2]. Gastrointestinal diseases in calves are characterized by dyspeptic disorders, disorders of water-salt balance and metabolism, changes in immunohematological status and varying severity degrees of endointoxication. Death of young animals in severe toxic forms of dyspepsia can reach 70-85% [3,4,5]. The range of etiological factors causing gastrointestinal diseases in young cattle is extensive: pathogenic and opportunistic microorganisms, viruses, invasive parasites, violations and errors in feeding and keeping animals and so on [2,6]. However, regardless of the nature of the etiological factor, a sick animal develops endogenous intoxication, which is caused by accumulation of toxic substances in the blood stream in concentrations exceeding the functionality of natural disposal systems [3,7,8,9,10]. Specific targets for endotoxins are connective tissue cells, macrophages, neutrophilic leukocytes, platelets and others [10,11,12,13]. In immunohematological studies of blood from sick calves, leukocytosis, shift of leukocyte formula to the left, acceleration of ESR, changes in the composition of immunocompetent cells (T-and B-lymphocytes), increase of the level of circulating immune complexes are registered. Such changes in indicators show the inflammatory reaction of the body as a whole, and are not characteristic of any single pathology. Moreover, changes in the complete blood count characterizing the inflammatory reaction in diseases of the gastrointestinal tract (GIT) in young cattle, with regard to indicators such as leukocytosis and stab shift of leukocyte formula to the left, are observed in 20% (leukocytosis) and 8% (formula shift) cases. Increased ESR occurs only in every third sick calf. Simultaneous combination of leukocytosis, stab shift of formula and elevated ESR is recorded in less than 10% of cases [4,5,6].
In veterinary practice, the issues of complications risk assessment in diseases of the gastrointestinal tract of young animals are particularly acute, as they determine firstly the appropriateness of expenses for therapeutic and prophylactic treatment, and secondly are the basis for the forecast of the population health of breeding young animals and its future productivity. For objective assessment of intoxication degree and the state of general resistance of the organism in medical and veterinary practice "hematological indices" are used, which reflect connections between different classes of cells of leukocyte formula [4,8,11,14,15]. It should be noted that "hematological indices" were mainly determined for adult animals and young cattle aged 6 months and older [17,18]. We did not find the information on "hematological leukocytic indices" self-descriptiveness for assessing intoxication degree and the state of reactivity of the body in gastrointestinal diseases of calves aged from 1 up to 3 months, which was the basis for this study.
Objective: determine the degree of endointoxication and reactivity of calves' bodies to estimate the risk of complications in diseases of the gastrointestinal tract.

Materials and Methods
The study was performed in the Ural Research Veterinary Institute of the Ural Research Institute of the Ural Branch of the Russian Academy of Sciences in the laboratory of immunology and pathobiochemistry of the Department of Ecology and Non-contagious Pathology in the framework of the direction 160 of Fundamental Scientific Studies programs of state academies of sciences for 2013-2020. "Molecular biological and nanobiotechnological creation methods of new generation biopreparations, technologies and ways of their application in order to combat especially dangerous infectious, parasitic and non-communicable diseases of animals".
Object of study: calves at the age of 1-3 months (n=254), blood. According to the results of clinical examination, the following symptoms were observed in calves with gastrointestinal diseases: dyspepsia, dehydration, suppression of motor activity. As a result of hematological studies for the period 2017-2019 the hemograms of healthy calves (n=86) and hemograms of calves with diseases of the gastrointestinal tract (n=168) were compiled and analyzed.
The peripheral blood analysis of calves was performed on an automatic hematological analyzer Abacus Junior Vet (Diatron, Austria) using standard Diatron reagent kits (Austria); the leukocytic formula was calculated with a generally adopted method using the microscope Micros MCX 100 (Austria).
To determine the degree of organism endointoxication and its immunoreactivity in calves, the following "integral leukocytic indices" were calculated: Leukocytic intoxication index in B.A. Reis' modification (LII) [12]. LII is used to determine the degree of intoxication, it indicates the severity of inflammation and the  [12]. LSI is used as a marker of the reactivity of the body in acute inflammation . An increase in LSI indicates an inflammatory process and a breach of immunological reactivity with pronounced endogenous intoxication.
LSI reference value 1.96±0.56 c.u. The lymphocytic index (LI) [12]. LI reflects the connection between the humoral and cellular link of the immune system of the body.

Results and discussion
To assess the severity of endogenous intoxication in calves with gastrointestinal diseases, we studied the results of a complete blood count (CBC). It was found that in all sick animals hemoglobin concentration in blood was significantly reduced and was determined in the range from 49.72 to 90.62 g/l. The total number of white blood cells in 54.2% of sick calves was 1.57-2.9 times higher and ranged from 12.4х109/l to 21.3х109/l. In healthy calves of similar age the total number of white blood cells was determined in the range from 4.5x109/l to 7.0х109/l. Stab shift of leukocyte formula to the left was registered in 11.3% of cases; increased erythrocyte sedimentation rate (ESR) -in 25.0% of cases. Combined diagnosis of leukocytosis, stab shift of leukocyte formula and increased ESR was observed in 9.5% of examined patients. Thus, if we evaluate the main markers of the inflammatory reaction of the body to endogenous intoxication only based on the results of the CBC, then the risk group for the complications development of the gastrointestinal disease should count as 16 (9.5 %) individuals (Fig. 1).  In determining the integral indices of intoxication LII, LSI and LI in the examined calves a slightly different picture of the nature of the inflammatory reaction was established. Thus, the degree of endogenous intoxication according to LII index in the examined calves in 6.6% of cases was interpreted as "severe" (5.8 c.u. ≤ LII ≤ 8.5 c.u.); in 45.8% of cases -as "medium"; in 34.5% cases -as "mild". In 13.1% of cases calves condition could be characterized as "borderline" [8,10,12,19]. The LII index in these animals was 2.47±0.18 c.u., which is on average 1.4 times higher than the normative indicator (N LII from 0.62±0.09 c.u. to 1.60±0.5 c.u.), but lower than the indicator of "mild degree of intoxication" -from 2.7 to 3.7±0.67 c.u. The share distribution of severity degrees of endogenous intoxication in calves with different spectrum of inflammatory reaction markers of the organism is presented in Figure 2. Group 1 -body inflammatory reaction marker "Leukocytosis"; group 2 -"Leukocytosis + elevated ESR"; group 3 -"Leukocytosis + "left shift in leukocyte formula"; group 4 -"Leukocytosis + increased ESR + "left shift in leukocyte formula".
The increase in the blood leukocyte shift index (ESR) by 2.2-3.5 times was recorded in 3.57% of cases (Fig. 3). Two of the calves of the 3rd group had LSI values of 2.12 c.u. and 2.74 c.u. Four calves of the 4th group had LSI values in the range of 3.07 to 3.58 c.u. LSI increase indicated an active inflammatory process in the body with pronounced endogenous intoxication in these examined animals. In 51.19% of cases there was an increase in LSI by 1.5-1. segmentonuclear neutrophils in the blood of sick calves, which testified to the predominant role of granulocytes in the immune response. In 45.24% of cases the value of LSI was determined in the range of physiological norm (from 0.93 c.u. to 1.07 c.u.).
Leukocytic index (LI) in our studies was used as an integral indicator of response reaction balance of cellular and humoral link of immunity to active inflammatory process in diseases of the gastrointestinal tract in calves (Fig. 4). In healthy animals, LI values were recorded between 0.96 c.u. and 1.68 c.u. In 66.7% of cases, the immune response had a balanced character as evidenced by the LI values, which were determined in the range of 1.01-1.70 c.u. The reaction imbalance of the cellular and humoral link of immunity was revealed in 33.3% of cases. In 48 sick calves (28.6%) there was a decrease in LI to 0.47-0.84 c.u., which can be interpreted as the predominance of cellular link reactions of immunity over the humoral link. In 4.7% of cases, humoral reactions prevailed in sick animals (LI 1.98-2.30 c.u.). Group 1 -body inflammatory reaction marker "Leukocytosis"; group 2 -"Leukocytosis + elevated ESR"; group 3 -"Leukocytosis + "left shift in leukocyte formula"; group 4 -"Leukocytosis + increased ESR + "left shift in leukocyte formula".
In comparison of the integral indices of intoxication LII, LSI and LI it was found that with diseases of the gastrointestinal tract in the examined calves in 47.6% of cases identified degree was "mild degree of body intoxication". Compensatory and adaptive reactions from the immune system were adequate (Fig. 5). In case of "moderate severity of body intoxication" tension in the hemopoiesis system was recorded in 85.7% of cases; imbalance in reactions of cellular and humoral link of immunity -in 48.1% cases. In animals with a "severe degree of body intoxication" tension in the system of hemopoiesis had a pathological character, imbalance in the links of immunity was determined in 90.9% of cases. Thus, according to the results of intoxication indices determination LII, LSI and LI in 53 calves (31.6%) there was a risk of complications development of gastrointestinal diseases. According to the results of CBC, only 16 sick animals (9.5 %) were classified as "risk group" (Fig. 1).
It is known that the cause of leukocytosis in diseases is leukopoietic function stimulation of hematopoietic organs as a result of the influence of pathogenic originators and inflammation endogenous factors. Leukocytic cells that have not managed to completely differentiate in the hematopoietic organs, in the form of immature form elements enter the blood, thereby determining its pathological picture [11,12,14]. When assessing the leukocyte formula in a particular sick animal, the total percentage of granulocytes is generally taken into account. Increasing it in 2 or more times above normal indicators shows the risk of pathological inflammatory process development in the body [1,9,17]. Quite often, the assessment of these data is subjective, because it does not give a complete picture regarding the adequacy and timeliness of the body's response to inflammation in general. In addition, numerous studies have proved that individual features of the immune reactivity of the organism are caused by the age of the individual, its physiological state, the influence of a complex of negative exogenous factors [5,13,17,18].

Conclusions
The studies have shown that in order to assess and predict the course of gastrointestinal diseases in calves aged 1-3 months, together with the CBC studies, to obtain expanded information on homeostasis condition it is efficient to apply "hematological leukocytic indices": leukocytic index of intoxication -LII, blood leukocyte shift index -LSI, lymphocytic index -LI. A comprehensive assessment based on the comparison of the LII, LSI and LI indices is more informative than a simple hemogram analysis. Firstly, it allows to characterize the severity and course of the inflammatory process and the level of endogenous intoxication. Secondly, to judge changes and disorders in some segments of homeostasis, such as resistance, immunoreactivity, immunocondition of the body. Thirdly, to determine the "key facts" indicating an increased probability of complications development risk of the disease and formation of pathological conditions in an individual.