COMPARISON OF PATHOLOGICAL CHANGES IN THE STUDY OF DOGS AFFECTED BY PARVOVIRAL ENTERITIS AND INTESTINAL YERSINIOSIS

Aim of the study: to examine the pathological changes in dogs that died due to confirmed intestinal yersiniosis and parvoviral enteritis and establish trends for characteristic organ lesions for both diseases in comparison, then evaluate our findings with the existing published material of sufficient evidence quality regarding differential postmortem diagnosis of spontaneous intestinal yersiniosis and parvoviral enteritis in dogs. Materials and methods: the study examined the cases of 25 dogs from 2 month to 1.3 y. o. of various breed, gender that died due to either confirmed Canine parvoviral enteritis (CPV) infection or intestinal yersiniosis and subsequently divided into two groups based on their diagnosis. The definitive diagnosis has been confirmed by performing rapid ELISA diagnostics (SensPERT®, VetAll Laboratories, Kyunggi-Do, Korea) for CPV and serological tests for Y. enterocolitica sera AT were performed using yersiniosis antigens from the «Component set for serological diagnosis of animal yersiniosis» (Kharkiv, NSC IEKVM, TS 46.15.091-95) in accordance with the «Interim guidelines for the use of a set of components for serological diagnostics». A dilution of 1:200 has been considered as the diagnostic titer. After the definitive diagnosis had been confirmed, the animals were divided into two groups, depending on diagnosis: CPV (n = 14) or IY (n = 11). The autopsies of twenty randomly (10 from each group) selected dog corpses have been conducted utilizing standard methodology. Results: according to the results of autopsy of dogs afflicted by CPV, the main pathological changes were found in the small intestine – catarrhal-desquamative inflammation (in 100 % of cases), serous-hemorrhagic mesenteric lymphadenitis (90 %) large intestine (70 %) in the stomach (60 %), in the liver (50 %), in other organs – less than 40 percent of cases and most notably caused lung damage (edema and local atelectasis) in 90 % of the animals in the study, which was not the case for intestinal yersiniosis with only 20 % incidence of lung damage. Conclusions: Spontaneous intestinal yersiniosis in dogs was pathologically manifested by pronounced catarrhal-desquamative processes mainly in the stomach and small intestine (70, 100 and 80 % of cases, respectively), inflammation of the mesenteric lymph nodes (90 %) and large intestine (80 %), dystrophy and congestive processes in the liver (80 %). Low incidence and type of lung damage (congestive hyperemia in 20 % of reviewed cases compared to 90 % of local atelectasis add edema in CPV group) was noteworthy. Cases of 25 animals that died due to either confirmed CPV or Y. enterocolitica infection were analyzed, and 20 animal corpses were autopsied during study. Dogs that died from intestinal yersiniosis had significantly higher frequency of pathological findings in kidney (200 %) and liver (60 %) in particular. Changes in stomach and large intestine were also more frequent. At the same time, we observed a lower frequency of pathological changes in spleen (33 % lower), heart (25 % lower) and the lowest Original Research Article: full paper (2021), «EUREKA: Health Sciences» Number 2

Veterinary Science and Veterinary Medicine the heart; granular dystrophy and necrosis of cardiomyocytes with destruction of myocardial muscle fibers. Also non-specific permanent morphological signs arising from circulatory disorders and heart failure were encountered like venous hyperemia of the liver, kidneys and pulmonary edema [9,10]. At the same time, mutations in viruses that cause CPV in dogs in recent times, somewhat change both the mechanisms of pathogenesis and pathological manifestations of the disease [4,11].
In canine intestinal yersiniosis, along with the asymptomatic course of the disease and fecal shedding of the pathogen, clinically pronounced forms of the disease are determined, accompanied by diarrhea, joint damage and cases of a generalized condition, leading to death [12][13][14]. Most reports indicate the dominance, prevalence and growth of pathogenicity of Y. enterocolitica, among other biovars of yersinia. Therefore, when describing the pathological picture of yersiniosis, the greatest attention was paid to intestinal yersiniosis. In addition, this is due to the important epidemiological significance of the disease. In published materials for intestinal yersiniosis described at autopsy catarrhal-hemorrhagic gastroenterocolitis, serous peritonitis, synovium, female genital lesions [15,16], rarely other pathologies [17], especially with complications of the underlying disease.
Express-diagnosis of these diseases is not always effective, due to the presence of a certain amount of antibodies and other reasons [18]. Thus, the information provided indicates the need to have criteria for pathological assessment of two similar clinical manifestations of diseases of different etiologies.
The aim of the study was to identify the comparative incidence of pathological signs caused by CPV and intestinal yersiniosis that can enable practitioners to conduct a differential postmortem diagnosis.

Materials and methods of research
The study analyzed cases of 25 dogs of various breeds, gender and weight, aged from 2 m. to 1.3 y. o., which were admitted to veterinary clinics «Hels» and «Vetservis» in Sumy, Ukraine throughout 2019-2020. The dogs presented with melena, severe dehydration, vomiting, various levels of ataxia and perished during the course of the disease. The work was performed in accordance with the research programs of Sumy National Agrarian University: «To develop an epizootic welfare control system for infectious animal diseases based on monitoring, diagnosis, forecasting and safety assessment of livestock and poultry products in North-Eastern Ukraine» (No. DR 0114U000261). The work was carried out for the period from 2019 to 2020 at the Department of Virology, Pathological Anatomy and Poultry Diseases of Sumy National Agrarian University branch, located at a small animal clinic «Vetservis» Sumy, Ukraine.
The definitive diagnosis has been confirmed by performing rapid ELISA diagnostics (Sens-PERT ® , VetAll Laboratories, Kyunggi-Do, Korea) for CPV and serological tests for Y. enterocolitica sera AT were performed using yersiniosis antigens from the «Component set for serological diagnosis of animal yersiniosis» (Kharkiv, NSC IEKVM, TS 46.15.091-95) in accordance with the «Interim guidelines for the use of a set of components for serological diagnostics» approved by HOVM with the State Inspectorate on November 16, 1995 according to No. 15-14/52). A dilution of 1:200 has been considered as the diagnostic titer.
After the definitive diagnosis had been confirmed, the animals were divided into two groups, depending on diagnosis: CPV (n = 14) or IY (n = 11). The autopsies of twenty randomly (10 from each group) selected dog corpses have been conducted utilizing standard methodology [19] at «Vetservice» veterinary clinic in Sumy. During the autopsies, the following criteria were taken into account: the location and type of lesions (Tables 1, 2), frequency of occurrence of different lesions in each group (Tables 1, 2).
Pathoanatomical research was conducted on diseased animals. The animal corpses were stored in laboratory freezers at -5 °C. After the autopsy all the specimens were destroyed in ac-

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We used percentage calculation to compare the incidence of different pathoanatomical phenomena in both research groups, results were statistically analyzed using Mann-Whitney U criteria. The results were calculated using Microsoft Excel 2016.

Results
During the autopsy of dogs afflicted with CPV mild stage of rigor mortis was observed. Serous fluid or crusts of mucus or dried serous exudate were found around the nose, and around the mouth -varying from gray to pale pink. The anal area and hind limbs were contaminated with liquid bloody feces. Dissection of the corpses revealed effusion into the abdominal cavity. In the stomach, the mucous membrane was grayish-red with hemorrhages. Inflammation of the gastric mucosa was localized mainly on the folds (Fig. 1). The intestine, where the main pathological changes were localized, was almost void of content. The mucous membrane was in a state of acute catarrhal-hemorrhagic desquamative inflammation ( Fig. 2, a, b), mainly in the small intestine. Also, some areas with erosive hemorrhagic lesions were spotted. In some cases, peeling of the mucous membrane was observed in areas of several centimeters.
The liver was enlarged, elastic and firm consistency, brown, with grey focal lesions on the surface of a parenchymal cut (Fig. 3).
Visible anatomical changes in the kidneys were almost not determined. There were hemorrhages in the spleen. Pleural hemorrhages, foci of atelectasis and pulmonary edema were detected. The lungs were mostly elastic and loose when pressed. The incision surfaces of both lungs were spotted with hemorrhages (Fig. 4).

Fig. 4. Lesions in canine lungs with CPV
The results of the generalization of the pathological and anatomical picture in the study of dogs that died from CPV are presented in Table 1.

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Analysis of pathological and anatomical findings shows that despite the age difference (from 2.5 months to 1.5 years) of animals the majority of subjects demonstrated significant damage of the abdominal organs. The severity of pathological changes depended primarily on the duration of the clinical period and the severity of the course.
In most cases, the heart muscle damage could not be detected during an autopsy, but this does not exclude changes in the organ at the tissue level. Hemorrhages were found in the thoracic and abdominal cavities in 20-30 % of cases. The variety of changes in the lungs (from hyperemia and edema to focal atelectasis), which was observed in 90 % of cases, is noteworthy.
Pathological examination of dogs with spontaneously occurring yersiniosis revealed mainly catarrhal gastroenteritis, catarrhal-hemorrhagic gastroenteritis or enterocolitis, characterized by uneven hyperemia of the mucous membrane, loss of luster, swelling, loosening and excessive folding (Fig. 5, a, b; 6, a, b) with excessive accumulation of turbid mucus in the form of strands or clots. The contents were mostly liquid, cloudy, laced with mucus and often contain blood impurities. The mesentery vessels were overflowing with blood; the lymph nodes were enlarged, with their dissection revealling signs of serous lymphadenitis and hyperemia (Fig. 7, a, b).
Examination of the liver revealed deep dystrophic processes with blunting of the edges of the organ, discoloration and increased tension of the capsule. The color of the parenchyma is heterogeneous varying from dark cherry to clay. Dissection reveals, smoothed parenchymal pattern and a scrape giving off moderate amount of content. The gall bladder was overflowing with bile in all cases.
The kidneys showed an insignificant increase in volume, mainly due to congestive hyperemia. Other changes were found less frequently ( Table 2).
Upon receiving data from the autopsies and incidence percent calculation, a percent bias was determined ( Table 3).

Discussion
The literature analysis revealed insufficient data regarding small animal yersiniosis, especially in the aspect of pathoanatomical diagnostics, which shows current lack of knowledge in this area possibly because of main research efforts being directed towards food safety and prevention of disease spread.
Comparative pathological studies have shown that it is extremely difficult to detect significant differences in the incidence of lesions of the gastrointestinal tract. Interestingly, for CPV, the phenomena of lung hyperemia, local atelectasis and edema were found in 90 % of cases, against 20 % of cases of congestive hyperemia in intestinal yersiniosis, which corresponds with available data from other sources [6,20]. At the same time examination of pleura and peritoneum in CPV revealed hemorrhages, which was not observed in intestinal yersiniosis.
Pathologically, both diseases show signs of granular dystrophy in the heart and kidneys further corroborating previous studies [4,9]. Moreover, CPV manifests pronounced desquamative processes, often with detachment of the mucous membrane on the background of the inflammatory process [3], which was not observed in intestinal yersiniosis [10,20]. In the case of bloody diarrhea, when the stools contain only small pieces of epithelium, the accurate clinical diagnosis is problematic [5].
Hemorrhages can be found in the body during CPV, but venous hyperemia was only detected in two cases of intestinal yersiniosis. Interestingly, for CPV, the phenomena of hyperemia, local atelectasis and edema were found in 90 % of cases, against 20 % of cases of congestive hyperemia in intestinal yersiniosis. Pathologically, both diseases show signs of granular dystrophy in the heart and kidneys.
Depending on the timing of the disease, the use of specific diagnostic tests may also be unsuccessful [2] and definitive tests for Y. enterocolitica require bacterial cultures or sera titer diagnostics, which can be time-consuming and ineffective as a rapid diagnostic test in-vivo [12].
Research limitations. The diagnostic potential of the described method is limited due to: 1. Small quantity of specimens, which is a result of seldomly conducted autopsies of deceased patients at small animal clinics due to absence of mandatory autopsy provisions in veterinary regulatory documents, inability to properly perform the autopsy or to adhere to proper pathological material disposal protocol and ethical concerns of the owners, unwilling to give their consent for an autopsy. Further studies will enable proper statistical analysis of incidence of different pathoanatomic findings.
2. Potential concurrent course of both diseases and/or presence of secondary or underlying pathological process/status severely limits the possibility of accurate pathoanatomical diagnosis.
Further studies. Further research will be directed towards the increase of the quantity of autopsies being performed on target population animals in order to perform more accurately evaluations of the incidence of certain pathological changes in various organs and systems which will result in more accurate postmortem diagnostics.
Potentially, concurrent course of CPV infection and intestinal yersiniosis can be studied to determine the impact of secondary bacterial infections on the outcome of CPV.

Conclusions
1. Cases of 25 animals that died due to either confirmed CPV or Y. enterocolitica infection were analyzed, and 20 animal corpses were autopsied during study.
2. Dogs that died from intestinal yersiniosis had significantly higher frequency of pathological findings in kidney (200 %) and liver (60 %) in particular. Changes in stomach and large intestine were also more frequent.
3. At the same time, we observed a lower frequency of pathological changes in spleen (33 % lower), heart (25 % lower) and the lowest frequency was in lungs (77 % lower) presented by edema and local atelectasis in animals afflicted by yersiniosis compared to CPV.
4. Incidence and manifestation of pathological findings was mesenteric lymph nodes and abdominal cavity were similar, and could not be considered during posthumous diagnostics.