Lip lesions in horses: case report

[Lesões labiais em cavalos: relato de caso]

ABSTRACT

The article presents the results of monitoring studies on the mucous membrane lesions in Trakehner horses. Based on clinical examination, the localization, size, and number of ulcers, as well as the inflammatory reaction to the presence of necrotic tissue, were determined. In addition to clinical examination, planimetric studies were also carried out to determine the size of the ulcers and the dynamics of their reduction during treatment with aerosol application of PVP iodine spray. To determine the nature of inflammation in the lesion focus, smears were taken from the surface of the ulcers. Microbiological and mycological studies of pathological tissues and feed samples were carried out to establish the etiological factors contributing to the ulcers. It was found that the size of the ulcers varied from 1 to 6 cm, and they were located on the upper and lower lips on both the right and left sides. Neutrophilic leukocytosis was observed in the focus of inflammation. Leukocytes actively phagocytized microorganisms present in the pathological focus. Microscopic fungi of the species A. flavus, A. fumigatus, M. species, and P. commune sprouted on Saburo medium from necrotic tissues of the pathological focus. A similar composition of fungal species was obtained by sowing feed samples fed to horses. Microbiological studies of ulcers revealed the growth of two microorganisms, S. aureus and P. vulgaris, on Baird-Parker agar. On the third day of treatment, compared to the first day, the size of the ulcers decreased by 22.8 %. It is recommended to perform primary surgical treatment of the ulcer, apply PVP iodine spray once a day, and replace the affected feed with high-quality feed to achieve a positive therapeutic effect.

Keywords:
horses; mucous membrane ulcers; microscopic studies; treatment

RESUMO

O artigo apresenta os resultados dos estudos de monitoramento das lesões da membrana mucosa em cavalos da raça Trakehner. Com base no exame clínico, foram determinados a localização, o tamanho e o número de úlceras, bem como a reação inflamatória à presença de tecido necrótico. Além do exame clínico, também foram realizados estudos planimétricos para determinar o tamanho das úlceras e a dinâmica de sua redução durante o tratamento com a aplicação de aerossol de iodo PVP. Para determinar a natureza da inflamação no foco da lesão, foram feitos esfregaços da superfície das úlceras. Foram realizados estudos microbiológicos e micológicos de tecidos patológicos e amostras de ração para estabelecer os fatores etiológicos que contribuíram para as úlceras. Verificou-se que o tamanho das úlceras variava de 1 a 6 cm e que elas estavam localizadas nos lábios superior e inferior, tanto do lado direito quanto do esquerdo. A leucocitose neutrofílica foi observada no foco da inflamação. Os leucócitos fagocitaram ativamente os microrganismos presentes no foco patológico. Fungos microscópicos das espécies A. flavus, A. fumigatus, M. species e P. commune brotaram em meio Saburo a partir de tecidos necróticos do foco patológico. Uma composição semelhante de espécies de fungos foi obtida com a semeadura de amostras de ração fornecidas a cavalos. Os estudos microbiológicos das úlceras revelaram o crescimento de dois microrganismos, S. aureus e P. vulgaris, no ágar Baird-Parker. No terceiro dia de tratamento, em comparação com o primeiro dia, o tamanho das úlceras diminuiu em 22,8%. Recomenda-se realizar o tratamento cirúrgico primário da úlcera, aplicar o spray de iodo PVP uma vez ao dia e substituir a ração afetada por ração de alta qualidade para obter um efeito terapêutico positivo.

Palavras-chave:
cavalos; úlceras de membrana mucosa; estudos microscópicos; tratamento

INTRODUCTION

In Ukraine, horses are used in sports, for hippotherapy, transportation of goods and people, as well as for agricultural and forestry work, especially in mountainous regions where access by automobiles is limited.

The most common oral disease in horses is ulceration of the mucous membranes. The main causes of ulcers in horses' mouths are trauma, autoimmune and neoplastic processes, poor coarse feed, and infection with fungi and bacteria (Easley et al., 2010). There are scientific reports that ulcers next to the first and second premolars and near the corner of the lips are caused by tension in the bit and bridle, which is not affected by the normal displacement of teeth (Tell et al., 2008; Björnsdóttir et al., 2014). In Icelandic horses, the use of a curb bit with a port has been associated with a higher risk of lower jaw branch injuries compared to a snaffle bit or a traditional Icelandic curb bit (Björnsdóttir et al., 2014; Tuomola et al., 2021).

Pain in the horse's mouth is considered a serious welfare problem (Popescu et al., 2014; Tuomola et al., 2019). Conscious perception of pain that occurs in the body as a whole, its role and evaluation through behavior, as well as the sensory function of the horse's oral cavity, are described as a basis for a more detailed assessment of pain in the mouth (Mellor and Beausoleil, 2017; Mellor, 2020).

MATERIALS AND METHODS

Clinical monitoring studies aimed at detecting lesions of the oral cavity were conducted in the Poltava region for nine years (2014-2023). Horses with signs of oral cavity lesions were identified at the Horse farm "Traken" (n=5) in Chutovo village (Kulinich, 2015), and at the Ei-Di equestrian club in Gozhuli village, where a diagnosis of ulceration of the mucous membrane of the lips was established during clinical examination of the animals.

Visual studies were conducted in the following sequence: firstly, a speculum was inserted into the oral cavity of the animal and fixed at the occiput (Figure 1). After that, an examination of the oral cavity was performed. The following were determined: depth of the oral vestibule, color of the mucous membrane, presence of swelling of the mucous membrane (impressions of teeth along the line of their contact), condition of the salivary gland ducts, and the character of secretions from them. The attachment of the upper and lower lip frenula, lateral restraints of the mucous membrane, were also examined. Subsequently, an examination of the oral cavity proper was performed, considering the color, moisture, turgor, integrity, and the presence of pathological formations (desquamation, erosion, ulcers, neoplasms) of the mucous membrane, as well as the state of the tongue (size, mobility, foldability, and the presence of swelling), the floor of the mouth, and the ducts of the sublingual and submandibular salivary glands. The condition of the tongue frenulum, the shape of the palate, the condition of the mucous membrane of the hard and soft palate, the uvula, and the tonsils were also examined.

Laboratory studies were conducted at the Regional State Laboratory of Veterinary Medicine in the Poltava region. Cytological studies were performed by making smears of imprints from ulcers and staining them using the Romanowsky-Giemsa method. Before taking the smears, the mucous membrane of the animals' oral cavity was rinsed with a solution of boric acid and 5-7 drops of a 1% solution of dycaine were applied to the mucous membrane of the lips (Ostrovsky, 1978). We also conducted mycological studies to detect microscopic fungi in the pathological focus, feed, and bacteriological studies. For the former, cultures were made on Sabouraud agar. Identification of fungal cultures was performed by comparing macroscopic and microscopic features of the experimental cultures with previously described features of fungi (Sutton et al., 2001). Microorganisms present in the pathological material were isolated using the Baird-Parker agar method, and their hemolytic activity was determined on Cetler's blood agar. Identification was carried out according to Bergey’s Manual. After performing the initial surgical treatment (removal of purulent-necrotic masses using a swab), PVP iodine spray from the Czech manufacturer Bioveta was sprayed onto the surface of the ulcer from 20-30 cm. In our research, we conducted clinical and planimetric studies to objectively assess changes in the state of the pathological process. To do this, we measured the size of defects on the first, third, and 14th day using a ruler with a division price of 1 mm (DSTU 427-2009).

Figure 1
Preparation of the horse for clinical examination a) pharynx; b) fixation of the pharynx on the animal before examination.

When evaluating clinical changes, the nature of the ulcer surface and the presence of secretions were assessed. The experimental material obtained was processed by the method of variation statistics with determination of arithmetic means (M) and standard deviations (m).

The housing conditions for animals and the methods used in experimental research followed the guidelines set forth in the European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes (Strasbourg, 1985). The research protocol of the current study was approved by the Ethic Committee of Poltava State Agrarian Academi (Aprobal number 2020/04.

RESULTS AND DISCUSSION

During the dispensary examination of horses at the Chutivsky stud farm "Traken" (n=5), 5 heads with ulcers on the mucous membrane of the lips were detected. In 100% of cases, the lesions were diagnosed in stallions. The appearance of ulcers on the left lateral surface of the lips was diagnosed in the stallion Hodky (Figure 2a). The size of the ulcer on the upper lip was 1.5x2.2 cm, and on the lower lip, it was 4.3x2.7cm. In the stallion Olimp, the appearance of three ulcers on the right lateral surface of the lips was diagnosed. Two of them were about 0.8x0.6 cm in size, and one was 1.2x0.8cm. In the stallion Piero, during the clinical examination, an ulcer measuring 5.1x6.3cm was found on the left side of the upper lip (Figure 2b). In the stallion Pléro, an ulcer was found on the upper lip in the central part, measuring 7.2x3.6cm. In the stallion Prospekt, ulcers measuring 1.2x1.4cm were found in the corners of the lips. The ulcer had a dirty color, with necrosis of the edges and the presence of purulent exudate under which a wet surface of red color was revealed. Inflammatory swelling and hyperemia around the ulcer were also pronounced. Small ulcers were cleaned of necrotic tissue and purulent exudate, and the appearance of small granulations on the bottom and edges of the ulcer was observed. Along the periphery of the ulcer, scarring and epithelization from the edges were noted. Smears-imprints from the mucous membrane of the lips of sick animals showed an increase in the number of epithelial cells, which were in a state of dystrophy and necrosis.

Uldahl et al. (2022) introduced a protocol (Oral Commissure Assessment) for the classification and registration of pathological lesions on the corners of the lips. The precise anatomical location was considered, pigmentations were classified as natural or potentially pathological, and pathological lesions were characterized. 206 horses were examined, only two had bruises or erosions, five had ulcers, and none had bleeding.

Figure 2
Ulcers: a) on the upper and lower lips on the left side; b) on the upper lip.

We also observed the appearance of neutrophil leukocytosis. The latter were in a state of phagocytosis and actively engulfed microorganisms present in the pathological focus (Figure 3). Neutrophils on the smear were clustered in amounts ranging from 2 to 10 cells.

Figure 3
Native smear of horse lip mucosa ulcers (X400), phagocytosis of microorganisms by neutrophil; a) stallion Hodky; b) stallion Piero

In this case, the cellular composition in 5 fields of view under the microscope was as follows: erythrocytes 12 ± 4.3, neutrophil leukocytes 4.3 ± 2.5, and epithelial cells 3.5 ± 1.5.

Samples of necrotic tissue from the surface of ulcers were placed in bacteriological tubes with isotonic sodium chloride solution and were sown in a dense Sabouraud medium in laboratory conditions. When the culture of the fungus A. flavus (Figure 4) grew on Sabouraud's medium, the growth of mycelium was not intensive and was registered from the 4th day. Later, on the fifth day, an increase in mycelium above the surface of the nutrient medium was noted. By the fifth day, the mycelium had acquired a more intense coloration and darkened.

Figure 4
Mycological studies. a) growth of M. species and A. flavus on Sabouraud medium; b) mycelium of A. flavus (X150)

When growing the mycelium of the A. fumigatus fungus on Sabouraud agar, the first signs of development in the form of semi-transparent formations were observed after 8 hours. After two days, the appearance of a whitish fluffy coating was observed. On the fourth day, the growth of a massive colony of round shapes with a greenish-blue tint in the center and a whitish border on the periphery was recorded. There are descriptions of liver lesions in horses caused by A. Fumigatus, as well as reports of infections of feed with this type of fungus, such as hay, silage, oats, and concentrated feed (Caloni and Cortinovis, 2011). Randleff-Rasmussen et al. (2017) describe the non-classical detection of a skin mycetoma of the lateral wing of the right nostril in a 16-year-old Fjord stallion with suspected dysfunction of the intermediate part of the pituitary gland, in which a painless, hard new growth was found around the lateral wing of the right nostril and lip. Cytological examination of the formation showed marked pyogranulomatous inflammation, and histopathological examination revealed a fungal mycetoma. The fungal culture was identified as Aspergillus terreus. The authors achieved positive results after 5 months of treatment with pergolide, potassium iodide administered orally to the animal (Randleff-Rasmussen et al., 2017).

In addition, growth of fungi of the species Mucor was observed. The growth of the fungus was noted after 36 hours in the form of a white fluffy coating. After three days, the formation of a white voluminous fluffy colony with a strong mycelium was recorded. On the fifth day, the mycelium became tall, gray, and had an airy structure in the form of a canopy. Imogen Johns (2011) reports the fact of the sowing of Mucor fungi from the conjunctiva of horses. Abdullah (2022) identified the growth of Mucor indicus fungi in a mare with inflammation of the nasal cavity.

The growth of the P. expansum fungus was characterized by the appearance of green mycelium on the surface of the medium from the third day. The colonies of the P. commune fungus grew in the form of radial, fluffy, blue-gray colonies, with orange-brown on the reverse side of the cup; their edge thickened more towards the center. According to the analysis of research results, it was found that in cases of ulcerative lesions of the lips, microscopic fungi were more often detected in the form of associations.

In order to identify the source of mycotic pathogens, a study of feeds that are included in the diet of horses was conducted. Samples of feed were sown on Sabouraud's medium and incubated at 25°C for five days. Analysis of the results showed that the species composition was practically identical to that isolated from the ulcerations on the lips.

During bacteriological studies, two microorganisms were isolated by the Baird-Parker agar method: S. aureus and P. vulgaris. Both species grew on blood agar but did not produce hemolysis (Figure 5).

After removal of purulent-necrotic masses using a tampon, PVP-Iodine spray (PVP YOD spray) from the Czech manufacturer AO "Biowet" was aerosolized onto the surface of the ulcer from 20-30 cm (Fig. 5a). In our study, for an objective assessment of changes in the state of the pathological process during treatment, clinical and planimetric studies were conducted. For this, the defects were measured using a ruler on the first, third, and 14th day. Clinical changes were evaluated by the character of the surface of the ulcers and the presence of secretions (Figure 5b).

Figure 5
Microbiological studies a) Growth of P. vulgaris on Baird-Parker agar b) Growth of S. aureus on Zeissler blood agar

The area of the ulcer surface in animals during treatment significantly decreased (Table 1). Thus, on the third day relative to the initial data, the average reduction in the defect was 22.8%. Accordingly, on the 14th day, the reduction of the defect relative to the initial data was 4.2 times, and 3.3 times relative to the indicators on the third day. Clinically, starting from the third day, a decrease in the defect and the disappearance of signs of purulent exudation were observed. In the future, epithelialization was noted creeping onto the surface of the ulcer.

Table 1
Dynamics of wound surface area in the treatment of horses with lip ulcers, cm², M±m

Indicators		Defect size
Day of Treatment	1	14.5±0.72
	3	11.2±0.36

While studying a 6-month-old colt named "Gai" at the AD equestrian club in Gozhuli village, bilateral lesions of the lips on the right side were identified. Accordingly, the ulcers had a pink color with a gray border on the periphery. In some places, the accumulation of gray necrotic tissue was noted on the surface. On the upper lip on the right side, one large lesion was found, measuring 8.2 x 4.1cm, and on the lower lip, there were two separate lesions of regular shape measuring 6.2 x 2.2cm and less than 5.4 x 2.1cm. On the left side, a lesion measuring 5.4 x 2.6cm was found on the upper lip, and two oval-shaped lesions on the lower lip merged into one and measured 5.6 x 1.8cm.

Cytological examination of a native smear of the mucous membrane of the affected lips revealed neutrophil infiltration. It was found that neutrophils were at different stages of maturation: promyelocyte, segmented neutrophil, phagocytosis of microorganisms by neutrophils.

Nicolas J. Vos recommends using iodine solutions for flushing after surgical treatment of lip ulcers in ponies caused by Actinomyces fungi. Dixon & Dacre (2005) report successful treatment of ulcers caused by stomatitis with diluted iodine solution. These reports are consistent with the use of iodine solutions in our studies.

CONCLUSION

The prevalence of lip ulcers in horses has been established through cytological and mycological research, as well as treatment of affected animals. The effectiveness of the procedures performed was determined based on clinical and planimetric data analysis.

It has been proven that clinically, ulcers had a dirty color, with necrosis at the edges, the presence of purulent exudate under which revealed a wet surface of red color. Inflammatory swelling and hyperemia around the ulcer were also pronounced.

Mycological studies of samples of necrotized tissue identified the presence of microscopic fungi species such as A. flavus, A. fumigatus, Muсor species, P. expansum, P. commune, and bacterial cultures S. aureus and P. vulgaris.

Removing the affected feed from the diet, replacing it with quality feed, and subsequent treatment of sick horses with aerosol application of the PVP YOD spray from the Czech manufacturer AO "Biovet" was found to be quite effective, as the area of the wound surface in animals decreased 4.2 times by the end of the treatment.

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Publication Dates

• Publication in this collection
  12 Feb 2024
• Date of issue
  Mar-Apr 2024

History

• Received
  24 June 2023
• Accepted
  06 Oct 2023