Clinico-pathological Studies on Canine Mammary Tumors in Dachshund Dog

Mammary neoplasms in dogs are the second most common neoplasms after skin tumors (Rezia et al., 2009; Benjamin et al., 1999) occupying about 52% of all tumors (Egenvall et al., 2005). Nearly 41% to 53% of the mammary tumors that occur in the bitch are malignant (Misdorp 2002). Malignant mammary tumors are most nonresponsive if the treatment is not started earliest (Shoji et al., 2016). Female spayed dogs have low risk of developing mammary tumors before their first oestrus (0.5% incidence compared to intact female dogs) and between first and second oestrus this rate is 8% (Salas et al., 2015). World Health Organization International classification of mammary tumor of the dogs and cat combines histogenic and descriptive morphologic classification, incorporating histologic prognostic features that have been associated with increasing malignancy (Patel et al., 2019). Dog has not only a tremendous mental healing power but also the guarding ability and is also becoming one of the biggest income-generating animals (Ray et al., 2019). But, there are a lot of clinical problem which affects their commercialization. A number of breeds such as Airedale terrier, Brittany spaniel, Boston terrier, Cocker spaniel, English setter, English springer spaniel, Fox terrier, German shorthaired pointer, Great pyrenees, Irish setter, Keeshond, Labrador retriever, Pointer, Dachshund, Poodle and Samoyed have already been reported to be predisposed to mammary tumor (Mitchell et al., 1974; Moe, 2001). Palta (2000) reported that white spitz breed most commonly suffered from canine mammary tumours (34.10%) followed by Doberman (19.65%), German Shepherd (9.83%), non-descript (8.10%), Labrador Retriever (7.57%), Pointer (5.78%), Crossbred (5.20%), Dachshund (2.89%), Boxer (2.31%), Cocker Spaniel (1.73%) and Lhasa (1.16%). Dachshunds and Cocker Spaniels bitches had an increased risk of developing mammary tumours, while Chihuahuas had reduced risk. Beagles had a risk slightly higher than mean risk for all dog breeds (Gupta, 2012).
       
Dachshund breed is prone to developing certain types of tumors such as mammary tumor, oral melanoma, mast cell. However, osteosarcoma, testicular tumor, bladder cancer and lymphoma are also found in Dachshund breed. Mammary gland tumor is the most common type of tumor in the dachshund breed (Moulton et al., 1970). The present study was undertaken to find out the epidemiological data of canine mammary tumor in relation to age, growth rate, lymph node status, type of tumor Besides focusing on their TNM (T means the degree of the primary tumor, N is the condition of the regional lymph nodes, and M means the absence/presence of distant metastases) grading in the Dachshund breed.

This study was performed at the Department of Veterinary Gynecology and Obstetrics, West Bengal University of Animal and Fishery Sciences, Kolkata and different clinics of Kolkata, India. During this period of study, a total of 59 cases of canine mammary neoplasm were detected and out of these 24 cases were in dachshund breed. Also, 6 normal dachshund bitches were included in this study as control animals.
       
Stages of tumors were classified using TNM with slight modification as descried earlier by Owen (1980). The tumor size were classified as T1 (<3 cm), T2 (3-5 cm), or T3 (>5 cm), N indicates the condition of the regional lymph nodes and M for presence of distant metastases. With this system, animal were classified into five clinical stages namely stage I, stage II, stage III, stage IV and stage V. In Stage I, II and III tumor size is ≤3 cm, 3-5 cm and ≥ 5 cm without lymph node metastasis and absence of distant metastases. Where as in stage IV tumor size varies with metastasis in lymph node present and absence of distant metstases. Stage V tumor size varies with or without lymphnode metastasis and presence of distant metastases (Gundim et al., 2016).
 

       
The total erythrocyte count (TEC), total leukocyte count (TLC), platelet count and packed cell volume (PCV %) was carried out as per standard methods described by Schalm et al., (1975). The hemoglobin was estimated by using photoelectric colorimeter by cyanomethemoglobin method as described by Cannan (1958).
       
Pre-operated mammary tumor patients were stabilized by giving balanced electrolyte fluid and vitamins. The mammary tumor patients were operated upon a combination of injectable anesthesia. The bitches were injected with Atropine Sulphate @ 0.04 mg/kg sub-cutaneously. and later Xylazine HCL @ 0.8 mg/kg was injected. Injection of Ketamine HCL @ 7 mg/kg body weight and diazepam @ 0.05 mg/kg body weight was given with Ringer’s lactate drip to maintain the surgical stage of anesthesia.
       
Histological examination was performed by staining with H&E stains and observed under microscope according to the World Health Organizations (WHO) diagnostic criteria (Sharma et al., 2018).
       
Data generated were analyzed using procedure described by Snedecor and Cochran (1968). Data were analyzed by using IBM SPSS version 23.0 statistical software.
 

All animals in this study were intact. Highest number of benign and malignant mammary tumor was observed in the age group of 8 to 12 years but benign tumor was found lowest below 12 years and malignant before 8 years of age. (Table 1). The specific occurrence of mammary tumors in this study showed a trend of increasing the percentage of malignant and benign neoplasia with advancing age till 12 years and reduces thereafter. Our result also indicates that chances of malignant neoplasia is more than Benign after 12 years of age. The growing age results in the accumulation of thrombogenesis factors leading to malignant tumors (Witsch et al., 2010). Findings of the study matches with those of Egenvall et al., (2005), who reported that mammary tumours were extremely rare in dogs younger than five years of age and the incidence increased sharply at approximately six years of age and incidence peaked at about 8-12 years of age. Similar result was also obtained by Taylor et al., (1976).
 

       
Maximum number of mammary tumors were recorded in the inguinal region (Table 2). We observed more involvement of Caudal 4^th and 5^th mammary glands than that of cranial glands. this result is also supported by Misdrop and Hart, (1976); Hellmén et al., (1993). Multiple involvements were recorded in 5 mammary tumor patients. These findings corroborated with the earlier observation made by Moulton et al., (1970). Multiple mammary gland involvement occurs possibly due to the infiltration of tumor cells to the adjacent mammary gland (Else and Hannant, 1979; Rosychuk, 1988).


       
Maximum benign mammary tumor showed a slow growth rate while malignant mammary tumors had a faster growth rate (Table 3). The slow growth rate in benign mammary tumors might be due to less angiogenesis in tumor mass compared to malignant tumors.
 

       
Maximum (79.16%) tumors were found >5 cm in diameter. A high percentage of large size tumors may be due to the result of the owner’s improper attention for slowly growing mammary neoplasm. All the small-sized tumors were benign type and among the large-sized tumors 63.63% were benign type and 92.3% were malignant type (Table 4). Perez-Alenza and Tbarena (2001) reported that 10 to 50% of dogs with mammary tumors have an enlarged lymph node. A similar finding is also reported by Misdrop and Hart (1976).
 

       
Regional lymph node enlargement were observed in 6 out of 24 cases. All cases of regional lymph node enlargement except one were associated with malignant tumor (Table 5). Only 9.10% of benign tumors showed an enlarged lymph node presumably because of local inflammatory reactions. 
 

       
Thoracic radiography revealed lung metastasis only in two dogs (8.33%) suffering malignant mammary tumors (Table 6). Cotchin (1958) reported that 10% of the histologically malignant neoplasms had distant metastasis. However, Krook (1954) found metastasis in 50% of dogs with carcinoma. This lower percentage of distant metastasis in this study might be due to early diagnosis and quick excision of tumor mass in mammary tumor patients.
 

       
Malignant mammary tumors were of clinical stage III (46.15%), 38.46% were of clinical stage IV, 15.38% patient showed stage V and 7.69% patient shows clinical stage II but no malignant of stage I was observed (Table 7). Benign tumors were hardly associated with lymph node or distant metastasis, so clinical stage IV and V were not found. Contrarily in malignant tumor lymph node and distance metastasis was involved, therefore stage IV and stage V found. This finding also supported by those of Gundim et al., (2016). During this study more than 50% tumors were of malignant type. Varallo et al., (2019) reported similar findings of benign and malignant mammary gland tumors with a range from 41-53%. Benjamin et al., (1999) and Sorenmo (2003) reported approximately 40-50% of canine mammary tumors are malignant. Comparatively benign mammary tumors were restricted to stage I, stage II and stage III while malignant mammary tumors were having the stages of III, stage IV and Stage V. Highest incidence of benign and malignant tumor was recorded with Stage III.
 

       
Histopathological  examination (Table 8 and Fig 2 A-D) revealed that out of 24 mammary tumors, 11 (45.83%) were benign tumors and 13 were malignant tumors (54.17%). Among the benign tumors, the maximum percentage having fibroadenoma and lower percentages were with duct papilloma. Moulton et al., (1970) and Bostock (1975) also reported the same. In case of malignant tumors, highest percentage (38.46) of papillary adenocarcinoma and lowest percentage (7.69) of solid adenocarcinoma were observed. This finding also collaborates with Moulton et al., (1970) who also concluded that Benign mixed tumors comprising 45.1% consisting of an adenomatous epithelial component and proliferative fibrous connective tissue and Malignant mixed tumors accounted for only 8.5%. 
 

 

         
Hematological analysis of 24 dachshund bitches and 6 normal bitches are presented below (Table 9). No significant change in any of the hematological parameters were observed in any of the group, Benign and malignant group as compared to normal group. However malignant group showed significant had significantly (P<0.01%) lower platelet count than the normal and benign group. The hematological findings were in contrast to our previous research which showed significant value of Hb, WBC, Eosinophil, BUN and SGOT from day 0 to day 28 during before and after treatment (Kumar et al., 2020) and corroborated with the findings of Sorenmo (2003) who stated that Blood parameters are normal in most dogs with mammary gland tumors but changes may occur in case of some other medical problems and tumoural bleeding. Perez Alenza and Tabanera (2001) reported that thrombocytopenia may be observed in some cases of malignant mammary tumors. Several factors such as polymorphism and mutation in some TFs and cytokines and effects of treatment could be associated with the incidence of thrombocytopenia in solid tumors. In spite of the importance of serious bleeding complication in solid tumors, few studies have yet analyzed the exact mechanisms of the impact of SNPs and mutation in the incidence of thrombocytopenia in solid tumors (Ghanavat et al., 2019).
 

Mammary tumors in Dachshund breeds are heterogeneous in nature. Hematological data is not significant as a prognostic factor for mammary tumor in Dachshund breed.  Biochemical and Molecular study may be carried out to find the exact genetic factor if there responsible to cause the canine mammary tumour in some breeds of dog so as to draw effective breeding strategy to preclude the disease from the population.

The authors are highly thankful to the Vice-Chancellor, WBUAFS for his kind support and help.

Anindita Sarkar and Ajeet Kumar Jha conducted lab work and prepared manuscript, Kalyani Ray designed the work and guided throughout the period. Siddhartha Basu, Anirban Mandal, Durgadas Mandal, Pradip Sarkar and Sanjoy Datta guided throughout the study and analysis of result. Keshav Kumar involved during correction of manuscript. All authors read and approved the final manuscript.