Introduction
The tiger (Panthera tigris) is the largest animal in the cat family, Felidae. Recent population estimation showed that the global wild tiger population was around 5,574 [1], and India has an estimated population of 3,682 tigers [2]. Zoological parks in India stock Royal Bengal tigers (P. tigris tigris), including white tigers. According to a census conducted during 2010–11, the number of captive tigers in India was approximately 381 [3]. Regular hematology and serum chemistry analysis in captive tigers is very important because the data can assist in the diagnosis of many diseases.
Biochemical and hematological studies are crucial tools for health evaluation and help interpret the status of physiological functions of various organs. For example, symmetric dimethylarginine (SDMA), creatinine, and blood urea nitrogen (BUN) are indicators of renal health in domestic and non-domestic cat family animals. A significant correlation was reported between serum SDMA concentration and glomerular filtration rate (GFR) in cats, and SDMA is suggested as a sensitive biomarker for early kidney function detection problems [4]. Viral, bacterial, and parasitic infections may affect the biochemical and hematological parameters [5]. So, the objective of the present study was to conduct regular disease diagnosis which is mandatory for the conservation of captive tigers as they constitute an important gene pool.
A study was conducted from November 2022 to July 2023 to analyze the hematological and serum biochemical parameters of nine Royal Bengal Tigers in Arignar Anna Zoological Park (AAZP) in Tamil Nadu, India.
Materials and Methods
Place of the study and blood collection
The study was conducted on captive Royal Bengal tigers housed at Arignar Anna Zoological Park (AAZP) in Tamil Nadu, India, from November 2022 to July 2023.
The blood was collected from nine tigers from the common tail vein (Dorsal coccygeal vein) by a Veterinary Assistant Surgeon by restraining them properly in a squeeze cage. The collected blood was divided into two parts of 5 ml each. One part of the blood was directly collected in EDTA tubes and gently mixed. The remaining blood was collected in a clot activator tube and allowed to clot for serum separation by centrifugation at 3,000 rpm for 10 minutes. The blood samples were collected from nine tigers with ages ranging from 5 to 20 years. Among the nine tigers, five were males (four normal tigers and one white tiger) and four were females (two white tigers and two normal tigers). All nine tigers were healthy during the study period. Necessary permission was obtained from the Principal Chief Conservator of Forests and Chief Wildlife Warden, Tamil Nadu, for blood sample collection from the tigers of AAZP.
Hematological studies
Hematological analysis was done using “Mindray” (Model: BC-2800 vet.). White blood cells (WBCs), lymphocytes, monocytes, granulocytes, lymphocytes (%), monocytes (%), granulocytes (%), red blood cells (RBC), hemoglobin, hematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), (RBC Distribution Width) RDW, platelets, mean platelet volume (MPV), platelet distribution width, and platelet crit (PCT) were quantified and recorded. Each blood sample was analyzed three times (three replications) to derive a mean value.
Serum parameters
The serum parameters viz., glucose, creatinine, SDMA, BUN, BUN /creatinine, phosphate, calcium, total protein, albumin, globulin, albumin/globulin, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, total bilirubin, cholesterol, amylase, lipase, sodium, potassium, sodium/potassium, chloride, and osmolality were quantified by the instrument IDEXX Catalyst One by using Chem 17 and Lyte 4 clips.
Statistical analysis
Simple linear regression was used to determine the association between age and blood parameters. A two-tailed “t” test determined the significant parameter differences between white and normal tigers.
Results
Hematological values of captive Tigers at AAZP
Table 1 shows the hematological parameters of the Royal Bengal tigers. Except for a few parameters, all were within the reference interval in all tested tigers. Tiger “Anu,” a 20-year-old female, showed the lowest total WBC count (5.9 x 103/μl), while Tiger “Meera” showed the lowest platelet count (71 x 103/μl). The platelet count was significantly high (334 and 318 x 103/μl) in two 9-year-old normal tigers, Deva and Sunitha, respectively. The rest of the tigers showed normal values in other parameters.
Table 1.
Haematological parameters of Bengal tigers in Arignar Anna Zoological Park (AAZP), Vandalur.
| Sl. No. | Parameters | Units | Male | Female | |||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Normal/White | --- | NT-1 | NT-2 | NT-3 | WT-1 | NT-4 | WT-2 | NT-5 | WT-3 | WT-4 | |
| Age | Years | 5 | 7 | 9 | 10 | 18 | 8 | 9 | 13 | 20 | |
| 1 | WBC | (×103/μl) | 11.2 | 10.50 | 15.4 | 12.7 | 14.6 | 7.1 | 11.9 | 11.8 | 5.9 |
| 2 | Lymph# | (×103/μl) | 2.0 | 2.16 | 1.8 | 1.8 | 2.5 | 1.7 | 2.5 | 2.0 | 1.6 |
| 3 | Mon# | (×103/μl) | 0.6 | 0.63 | 0.9 | 0.4 | 0.8 | 0.3 | 0.7 | 0.6 | 0.4 |
| 4 | Gran# | (×103/μl) | 8.6 | 7.70 | 12.7 | 10.5 | 11.3 | 5.1 | 8.7 | 9.2 | 3.9 |
| 5 | Lymph% | % | 17.7 | 21.10 | 11.8 | 14.5 | 17.1 | 24.1 | 20.9 | 17.3 | 27.6 |
| 6 | Mon% | % | 5.3 | 6.16 | 5.7 | 3.4 | 5.8 | 4.5 | 5.8 | 5.3 | 6.3 |
| 7 | Gran% | % | 77.0 | 72.73 | 82.5 | 82.1 | 77.1 | 71.4 | 73.3 | 77.4 | 66.1 |
| 8 | RBC | (×106/μl) | 8.07 | 7.73 | 7.13 | 8.94 | 8.47 | 8.79 | 7.63 | 7.09 | 8.48 |
| 9 | HGB | g/dl | 15.2 | 14.16 | 14.5 | 16.3 | 15.1 | 17 | 15.5 | 13.2 | 16.5 |
| 10 | HCT | % | 44.3 | 42.13 | 42.7 | 46.9 | 45.0 | 48.6 | 46.0 | 39.0 | 48.1 |
| 11 | MCV | fL | 55.0 | 55.80 | 60.0 | 52.5 | 53.2 | 55.4 | 60.3 | 55.3 | 56.8 |
| 12 | MCH | Pg | 18.8 | 18.23 | 20.3 | 18.2 | 17.8 | 19.3 | 20.3 | 18.6 | 19.4 |
| 13 | MCHC | g/dl | 34.3 | 33.60 | 33.9 | 34.7 | 33.5 | 34.9 | 33.6 | 33.8 | 34.3 |
| 14 | RDW | % | 16.4 | 16.40 | 15.6 | 16.4 | 19.0 | 16.1 | 15.6 | 16.9 | 15.0 |
| 15 | Platelet | (×103/μl) | 163 | 109.66 | 334 | 201 | 149 | 71 | 318 | 274 | 237 |
| 16 | MPV | fl | 11.3 | 10.20 | 10.5 | 10.6 | 10.4 | 8.3 | 10.6 | 11.5 | 11.1 |
| 17 | PDW | 16.2 | 16.36 | 16.1 | 16 | 15.3 | 17 | 16.2 | 15.3 | 15.7 | |
| 18 | PCT | % | 0.184 | 0.114 | 0.350 | 0.213 | 0.154 | 0.058 | 0.337 | 0.315 | 0.263 |
| 19 | Eos% | 1.8 | 4.60 | 1.7 | 2.0 | 4.8 | 3.2 | 3.6 | 7.2 | 12.9 | |
NT – Normal Tiger; WT – White Tiger.
WBC=White Blood Cells; Lymph=Lymphocyte; Mon=Monocyte; Gran=Granulocyte; RBC=Red Blood Corpuscle; HGB=Haemoglobin; HCT=Haematocrit; MCV=Mean Corpuscular Volume; MCH=Mean Corpuscular Haemoglobin; MCHC=Mean Corpuscular Haemoglobin Concentration; RDW=RBC Distribution Width; MPV=Mean Platelet Volume; PDW=Platelet Distribution Width; PCT=Platelet Crit; Eos=Eosinophil.
The association between age and some hematological parameters, viz., lymphocytes, monocytes, granulocytes, and eosinophils was studied by linear regression model. Eosinophil was strongly associated with the individual’s age in a positive trend (p < 0.05) (Fig. 1A). Lymphocytes and monocytes were also positively associated with age, but there was no significance. A non-significant negative association was observed between age and granulocytes (Fig. 1B).
Figure 1.
Linear model showing the association between age and white blood cell percentage in tigers of Arignar Anna Zoological Park, Tamil Nadu, India.
Serum biochemical values of captive tigers at AAZP
The serum appeared cloudy for an 18-year-old male tiger. This cloudy nature of the serum was due to high cholesterol levels (371 mg/dl), which was significantly higher than other tigers (p < 0.05). BUN was also higher in this 18-year-old tiger (60 mg/dl) (Fig. 2). In general, cholesterol level was higher in older tigers (Fig. 3). ALT was not affected by age factor. A significantly high ALT level of 119 U/l was recorded in an 8-year-old white tiger, and the lowest ALT level (16 U/l) was recorded in a 10-year-old white tiger.
Figure 2.
Symmetric dimethylarginine (SDMA), creatinine, BUN, and total protein in the serum of nine captive tigers at Arignar Anna Zoological Park. NT— Normal Tiger; WT— White Tiger.
Figure 3.
Glucose, BUN, and cholesterol in the different age group of tigers.
BUN was found to increase with the age of the tigers. However, the association between BUN and age was not statistically significant (p > 0.05) (Fig. 4). SDMA level was also not affected by the age of the individual. The range for SDMA calculated from the samples lies between 6 and 11 µg/dl and was within the reference internal provided by IDEXX (0–14 µg/dl). Though creatinine showed a negative association with age, it was not statistically significant (p > 0.05). A graphical summary is presented in Figure 2.
Figure 4.
Simple linear model comparing age and some serum parameters, glucose, creatinine, BUN, SDMA, cholesterol, and ALT levels in tigers of AAZP.
The average serum parameters of all age groups of white tigers were compared with normal tigers by t-test, and no significant difference was found between them (Table 2).
Table 2.
Comparison of White and Normal Bengal tigers of all age groups for some serum parameters using t-test.
| Sl. No. | Serum parameter | White tiger (n=4) |
Normal tiger (n=5) | t-value | df | p-value |
|---|---|---|---|---|---|---|
| 1 | Glucose (mg/dl) | 94.50 ± 14.97 | 87.0 ± 7.07 | 1.00 | 7 | 0.35 |
| 2 | SDMA (µg/dl) | 9.75 ± 1.50 | 7.40 ± 1.67 | 2.19 | 7 | 0.06 |
| 3 | Creatinine (mg/dl) | 2.37 ± 0.53 | 2.52 ± 0.35 | 0.49 | 7 | 0.63 |
| 4 | BUN (mg/dl) | 28.25 ± 8.65 | 31.0 ± 16.29 | 0.30 | 7 | 0.77 |
| 5 | Cholesterol (mg/dl) | 211.50 ± 43.91 | 229.20 ± 79.73 | 0.39 | 7 | 0.70 |
| 6 | Bilirubin (mg/dl) | 0.52 ± 0.36 | 0.26 ± 0.09 | 1.61 | 7 | 0.15 |
| 7 | ALT (U/l) | 65.25 ± 42.32 | 69.0 ± 16.23 | 0.18 | 7 | 0.85 |
Discussion
Captive animals have longer life expectancy compared to their wild counterparts. Due to this reason, age-related diseases are common in captive animals [6,7]. Hematological and serum biochemical analysis in captive animals helps to identify health problems at the early stages. Serum parameters such as creatinine, BUN, and SDMA are markers of chronic kidney diseases [8,9]. Hall et al. found that serum SDMA concentrations significantly correlated to GFR and increased earlier than serum creatinine [10]. Also, SDMA had higher sensitivity compared to creatinine. Lamglait and Vandenbunder-Beltrame have reported an SDMA range from 14 to 67 µg/dl in cheetahs that died due to chronic kidney disease [11]. In the present study, the SDMA level in nine tigers ranged between 6 and 11 µg/dl. Brans et al. [12] reported that a serum SDMA level of less than 14 µg/dl could be accepted as a normal value in cats. Langhorn et al. [13] in studying cats with hypertrophic cardiomyopathy and diabetes mellitus, estimated that healthy cats with no medical issues showed an SDMA level of 15 ug/dl.
In the present investigation, the BUN level ranged between 17 and 60 mg/dl. A significantly high BUN (60 mg/dl) was recorded in an 18-year-old animal, which also showed a significantly high cholesterol level. These complications in this older animal indicated cardiovascular/renal issues due to age factors. In free-ranging tigers, the BUN range was recorded as 6.5 to 48.2 mg/dl, with a mean of 27.9 mg/dl [5]. Elevation of BUN is usually associated with renal disease, congestive heart failure, shock, hypertension, and so on [14]. Shrivastav and Singh reported a mean creatinine range from 1.6 to 4.6 mg/dl and a BUN range from 6.5 to 48.2 mg/dl in Bengal Tigers [5].
Diagnosis of diseases using blood and serum parameter levels has several limitations due to sensitivity and inter-individual variation. The limitation of the present study is that the sampling size is low. A reasonable sample size is essential to understand the age-related deviations in blood parameters.
Conclusion
The findings of the present study contribute to understanding the health status of Royal Bengal Tigers in AAZP. Significant variations were observed in WBC, platelet, BUN, and ALT levels in some individuals. Eosinophil count increased significantly with age. There was no significant difference in these parameters between white and normal tigers. The study revealed that all captive tigers in AAZP, with the exception of one or two individuals, were in normal health.
Acknowledgement
The authors greatly acknowledge the financial support provided by the Tamil Nadu Forest Department, Government of Tamil Nadu. The Principal Chief Conservator of Forests (PCCF) and Head of the Forest Force, and PCCF and Chief Wildlife Warden are acknowledged for their help, support, and encouragement. The authors acknowledge Dr K. Sridhar, Forest Veterinary Assistant Surgeon, AAZP, for his help in blood sample collection from tigers.
