Use of Infrared Thermography for Identifying Physiological and Hematological Conditions of Young Sapera Dairy Goats

Pamungkas FA, Purwanto BP, Manalu W, Yani A, Sianturi RG. 2020. Use of Infrared Thermography for Identifying Physiological and Hematological Conditions of Young Sapera Dairy Goats. JITV 25(3): 120-130. DOI: http://dx.doi.org/10.14334/jitv.v25i3.2522 Infrared thermography (IRT) is an alternative solution that can be applied to replace invasive methods currently used in the monitoring of goats' physiological and hematological parameters. This study was done to compare and correlate the physiological and hematological conditions of young Sapera dairy goats and their correlations with results obtained by IRT. Four young Sapera dairy goats (weight of 26-28 kg) were kept in the individual rearing cage. Skin surface temperature (TS), rectal temperature (TR), body temperature (TB), heartbeat (HR), respiration rate (RR), and IRT at eyes, mouth, nose, legs, left body, right body, vagina, and vulva were monitored from 6 a.m. to 6 p.m. in 2 h intervals. Blood samplings were done at the beginning and the end of the obsevation time. Results showed that IRTs at several body parts were positively correlated with physiological parameters, except for heartbeat. Negatively correlation was observed in hematological parameters. The highest correlation (r = +0.85) was observed in the correlation between the results of the left rear leg IRT on TB. It was concluded that IRT can be applied to examine goats’ physiological conditions especially body temperature.


INTRODUCTION
Body resistance to heat stress is an important factor for dairy goat to maintain optimal productivity according to genetic characteristics. Tyler & Ensminger (2006) report that heat-resistant animals can preserve their body temperatures in a normal range without changing their physiological and productivity status. Goat is a homeothermic animal which can control its body temperature constantly, even in an extreme environmental condition, and can adapt better to the hot environment than other animals (Kawabata et al. 2013;Naandam & Kojo 2014;Hasin et al. 2017;Ribeiro et al. 2018;Façanha et al. 2020).
However, dairy goats can also experience stresses such as physical, nutritional, psychological, and environmental stresses. The most frequently occurred environmental stress is heat stress that indicated by physiologically uncomfortable conditions for the animal (Gupta et al. 2013;Silanikove & Koluman 2015). Dairy goats experience heat stress when there is an unbalanced between heat production in the body or heat obtained from the environment and heat loss to the environment. Homeostatic responses that are generally occurred due to heat stress in goats include the increase in respiration rate, body temperature, and water consumption as well as the reduction in food intake (Gupta et al. 2013;Caulfield et al. 2014), immunity (Tao & Dahl 2013), and even animal death (Sarangi 2018).
On the other hand, several assessments on physiological and metabolic parameters in animal production commonly use invasive methods like the measurements of rectal temperature, respiration rate, and heartbeat as well as blood sample collection (Stewart et al. 2008;da Costa et al. 2015). These methods showed relatively inaccurate results due to an anxiogenic response from the procedure itself that makes difficulties in the interpretation of results (Soerensen & Pedersen 2015). Invasive methods are also subjective, as well as time, and require a lot of labor consumption in the identification of animal production parameters and have less consideration in animal welfare (Blokhuis et al. 2013). Therefore, the use of IRT becomes a good option in the monitoring of goats' physiological and hematological parameters.
IRT is a non-invasive method used in the measurements of heat transfer and blood flow patterns by detecting body temperature changes (Kammersgaard et al. 2013;Alsaaod et al. 2014;Nääs et al. 2014;Roberto et al. 2014;Tattersall 2016). The result obtained from IRT device allows direct monitoring in the temperature distribution at certain objects (Blanik et al. 2014) which helps in understanding the thermoregulation process (Ghahramani et al. 2016). Therefore, the present study aimed to compare and correlate the results obtained from the IRT method at several body objects with the other invasive methods for the assessments of young dairy goats' physiological and hematological conditions.

MATERIALS AND METHODS
This study was conducted at Goat Research Section, Indonesian Research Institute for Animal Production, Ciawi, Bogor, Indonesia situated at the elevation of 450-500 meters above sea level and rain intensity between 3500 and 4000 mm per year. Using animals in this study has been approved by the Experimental Animal Ethics Committee, Indonesian Agency for Agricultural Research and Development, Ministry of Agriculture, the Republic of Indonesia with Registration No. Balitbangtan/Balitnak/Rm/04/2019. Four young Sapera dairy goats with an average body weight of 26-28 kg were kept in the individual cages of 1.6 m x 1.0 m in size. The cage was located inside an asbestos-roofed building and each cage was made of metal with boarded floor and food container. All goats were fed by C-Prolac concentrate produced by PT. Citra Ina Feedmill Jakarta about 1600 grams per day and king grass silage at the same amount alternately. The feeding schedule was twice a day at 7 a.m. and 3 p.m. Drinking water was given to the animals two hours before feeding schedule using a water bucket provided in each cage.

Physiological responses
Physiological response parameters measured were skin surface temperature (TS), rectal temperature (TR), body temperature (Tb), heartbeat (Hr), and respiration rate (Rr). The physiological responses data were measured from 6 a.m. till 6 p.m. in 2 h intervals. TR was measured by inserting Omron rectal thermometer model Kyoto,Japan) to the rectum about 5 cm deep. TS were measured by Omron digital thermometer model MC-720 specified for measuring skin temperature (Omron Healthcare Co. Ltd., Kyoto, Japan) at four observation points, i.e. back (A), chest (B), upper leg (C), and lower leg (D). The average of TS mean was obtained using formula modified from McLean et al. (1983), while Tb was calculated according to McLean et al. (1983) as follows: TS = 0.25 (A + B) + 0.32 C + 0.18 D Tb = 0.86 TR + 0.14 TS Hr is measured by placing the stethoscope near the left axillary bone, and then the heart rate calculation is performed for one minute. The Rr is measured by placing the stethoscope on the animal's chest, then the calculation of the amount of inspiration and expiration for one minute of respiration.

Hematological parameter profiles
Analysis in hematological parameters was carried out by collecting blood samples at the beginning and the end of heat measurement which was at 6 a.m. and 6 p.m. About 0.5 ml of blood samples were collected from a jugular vein after cleansing the area around the animal's neck with cotton dipped in 70% v/v alcohol so the veins can be easily identified and the blood collection area can be free from dung which may contaminate the samples. The target vein was gently pressed using thumb until puffed. A 20G sterile syringe was employed and injected into the jugular vein at about 1-inch deep. The needle was connected with a vacutainer tube after the blood came out and the angle of the syringe was set to 45º. A little attempt may be required to find the correct needle position for discharging the blood. Each goat was returned to their crib after blood collection and no extra bleeding or inflammation should be confirmed.
Following blood collection, each tube was kept in the iced box and stored in the laboratory for further analysis. Vetscan ® HM5 hematology analyzer (Abaxis, Inc. Union City, CA, United States of America) was employed to check all blood profiles including white blood cell (WBC), red blood cell (RBC), lymphocytes (LYM), monocytes (MON), neutrophils (NEU), hemoglobin (HGB), hematocrit (HCT), mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concentration (MCHC), and red blood cell distribution width (RDWC).

Infrared thermography
Infrared thermography (IRT) was performed using an IR camera FLIR A320 (FLIR Systems Co. Ltd., St. Leonards, NSW, Australia). IRT was employed for thermographic sensing purposes with an emissivity coefficient of 0.98 and carried out in 2 hours interval starting from 6 a.m. till 6 p.m. The IRT was taken at several observation points of each goat including its eyes, mouth, nose, legs, the left side of the body, the right side of body, vagina, and vulva at the distance of about 1 meter from the goats standing point (Fig. 1). All sensing data were saved in the memory card before transferred to a laptop for further analysis using ThermaCAM Researcher Profesional 2.10.

Data analysis
All collected data followed MIXED procedure from SAS (V. 9.1; SAS Institute Inc., Cary, NC, USA). A coefficient correlation between IR thermographic sensing and physiological responses as well as hematological parameters were calculated using the CORR procedure also taken from SAS.

RESULTS AND DISCUSSION
Body temperature, heartbeat, and respiration rate have been extensively examined in goats. Previous investigations using an invasive method in the physiological parameter assessment (Ogebe et al. 1996) showed a coherent association with results produce by a non-invasive method such as IRT (Roberto et al. 2014;Siva et al. 2014;Lecorps et al. 2016). Descriptive analyses of physiological parameters as well as IRT are described in Table 1. Skin temperatures measured by thermometer have values close to the same parameter measured by IRT, particularly at the front and rear legs. Meanwhile, the body temperatures measured physiologically showed almost similar results with body temperature measured by IRT at the vulva. It is important to note that rectal temperature (39.0°C) has a higher value compared to the IR measurement at the vulva (37.7°C). IRT showed high values at vagina and vulva, but low values at body and legs, whereas at eyes and nose it showed medium-ranged temperatures. However, IR measurements at the vagina and vulva showed a less temperature increment (2.1°C) at all observation times compared to the other body parts.
The results obtained from IRT at the leg, either front or rear, closed to the results obtained by invasive method i.e., about ±1°C higher than the skin temperature.  The main reason for these similarities, as reported by Popoola et al. (2014), is that skin temperature is an adapted condition of the blood flow in the skin that ends up in the heat regulating process of the body and the skin, in which cattle usually release heat load from their bodies through skins at the neck, ears, and legs in a particular contribution to all body surface. Piccione et al. (2013) report that legs play an important role in the thermoregulation compared to the other body parts due to vasoconstriction in the legs can cause a better mechanism in the heat conversion.
The IRT result at the vulva (37.67°C) closed to the result obtained from body temperature measurement (37.97°C). Hoffmann et al. (2012) obtained a similar result in the IR thermographic sensing at cow's vulva (37.2°C). Different results have been reported by Stelletta et al. (2017) in which IRT at the vulva of Angora goats before lust synchronization is 36.78°C and decreases to 35.40°C at the end of the estrous period. Hoffmann et al. (2012) and Talukder et al. (2015) reported that vulva temperature was the indicator used to identify changes in body temperature as general features of animals' physiology and health. The difference in body temperature measured was due to the difference in the variety of goats used. Simoes et al. (2014) and Stelletta et al. (2017) reported an increase in vulva temperature during the estrous period until approaching ovulation, and then the vulva temperature decreased at the end of the estrous period. The absolute point of vulva temperature can be affected by an environmental condition such as relative humidity, wind speed, and solar radiation. According to Talukder et al. (2014), reduction in vulva temperature at the end of the estrous period is associated with a reduction in estrogen concentration.
The result found a higher rectal temperature compared to the IRT at the vulva. This difference can be acceptable due to rectal temperature measurement using a digital thermometer was considered as the best method to identify goats' condition. However, this method needs time and direct contact with the measured animals. Besides, the method of rectal temperature measurement using a digital thermometer depends on the depth of the penetration. Also, the type of the thermometer as well as the stress level of the animals may affect the value of temperature obtained (Burfeind et al. 2010;Naylor et al. 2012;Hoffmann et al. 2012).
The IRT at vulva showed the highest temperature (37.67°C) but the lowest temperature increased (2.1°C) across the observation time compared to those measured at the other body parts. This finding agrees with the finding reported by Hooper et al. (2018) in which they found the highest value (38.29°C) at vulva using IRT compared to the other body parts and the lowest temperature increment (0.66°C) was found at observation time from 6 a.m. to 6 p.m. In this present study, the lower temperatures at body and legs than at the vagina and vulva were possibly affected by the high thermal isolation due to the difference in skin thickness and fur compactness (Arkin et al. 1991). Moreover, Bianchini et al. (2006) and Radon et al. (2014) suggested that skin and fur may influence heat transfer depending on their color, compactness, diameter, and depth, as well as on heat transmissivity and absorption.

Feeding Feeding
In general, results by IRT showed a similar curve pattern with the RR compared to the HR pattern along the observation time. The mean HR was 95 beats/minute and the RR was 52 times/minute. The IRT results at eyes, mouth, and nose, and the left/right body is in the range between the TS and TB (Fig. 2A). TR and TB have relatively stable changes and have an almost similar pattern with IR image results at the vulva. Similarly, the TS curve performs the same pattern with IRT at the left rear leg (Fig. 2B). All parameters showed significant increments in the feeding time at 7 a.m., but no increase in the feeding time at 3 p.m, except for HR which still showed a significant increase. HR showed two peaks during the whole day which occurred 1 hour after feeding time. A high increase was found in the respiration rate from 8 a.m. to 12 a.m., whereas a big decrease started from 4 p.m. to 6 p.m. (Fig. 2).
Two major peaks observed for the HR (Fig. 2) represents an increase in heat energy release due to feeding that usually occurs about 1 to 2 hours after feeding time. This condition is usually followed by an increase in the TS at several body parts (Montanholi et al. 2008) as is found also in this present study. The increase in heat energy release occurred for 3 to 4 hours after feeding (Puchala et al. 2007). This increase was associated with the protein mobilization process to trigger glucose synthesis (Lawler dan White, 2003). Puchala et al. (2007) suggested that the increase in an HR was caused by an intention to get food and hunger condition as a response of lateral hypothalamus and projected to the lateral medulla (as host for cells actuating autonomous system including parasympathetic vagal nucleus) and a group of sympathetic system cells in the bone marrow. Moreover, Olsson and Carlsson (1999) suggested that the provision of water and food at the same time may stimulate sensory input in the faring area caused by temporary activation of the sympathetic neural system which may increase HR.
In this study, the average HR about 95 (min 88.5 and max 99.5) beats/minute and RR of 52 (min 39 and max 66) times/minute were obtained. Afshar et al. (2005) reported an average RR and HR of Iranian female goats were as many as 20.2 and 114.2 times/minute, respectively. Hooper et al. (2018) obtained an average RR of Saanen goats about 49.5 times/minute, whereas de Lima et al. (2019) obtained the same from similar breed about 72.6 times/minute after giving birth. In another experiment, Phulia et al. (2010) found an increase in RR from 43.7 to 77.3 times/minute when Sirohi goats stood in a hot environment for six hours. HR and RR increased due to the effects of environmental conditions and body temperature. Increased respiratory action is an attempt to stabilize heat loss by evaporative cooling (Gupta et al. 2013), whereas increased HR is due to the increased muscle activity in combination with the increase in respiration speed and reduction in the peripheral resistance of blood vessel. The increased HR can cause increased blood flow to the body surface that could increase heat loss via conduction, convection, radiation, as well as diffusion from the skin (Marai et al. 2007;Spiers 2012;Okoruwa 2014). Al-Haidary et al. (2012) suggested that the RR could be used to estimate the negative effect of ambient temperature as an indicator of stress due to heat stress. Okoruwa et al. (2013) also suggested that the RR of more than 12-20 times/minute in goats and sheep could be used as an indicator of heat stress. Swollen breath is a mechanism of evaporation due to heat loss and respiration rate tends to follow heat loss because of that evaporation (Marai et al. 2007).
Evaluation in hematological parameters plays an important role to identify animals' prosperity because blood is an important indicator in response to stress due to heat (Okoruwa 2015). Hematological profiles of young Sapera dairy goats did not show significant changes across the observation time (from 6 a.m. to 6 p.m.). However, some changes were found in a few blood parameters, but the increments were very small ( Table 2). The blood counts obtained in this present study were still in the reference hematological range for goats (Piccione et al. 2014;Arfuso et al. 2016) and higher values generally occurred in the morning. A study related to heat tolerance in goats has reported a decrease in few blood components in the evening such as HGB concentration (Correa et al. 2012).
Heat can reduce blood count due to hemodilution effect by which lots of water component will be transported by the blood circulation system to help evaporative cooling (Seixas et al. 2017). Almost similar HGB value was found in the experiment on West African Dwarf goats by Opara et al. (2010), whereas higher HGB, MCHC, and WBC values were found in the experiment on Etawah breed goats (Yupardhi et al. 2013) and on the crossbreeding of Saanen and Ettawah (Sarmin et al. 2019). Okonkwo et al. (2011) suggested that results on goat's hematology depended on the type and age of the goats. High HGB value tends to transport more oxygen to blood (Okonkwo et al. 2011). Similarly, higher erythrocyte (MCV, MCH, and MCHC) and WBC indices show a higher oxygen transport capacity on blood (Tsai et al. 2010) and a higher immune system to fight against infection agents (Piccione et al. 2014), especially on young goats than on does.
The associations between IRT results with physiological and hematological parameters are presented in Table 3 and Table 4. The results obtained from IRT sensing were positively correlated (p < 0.0001) with goats' physiological parameters (Table 3) such as TB (r = 0.69 to 0.85), TS (r = 0.63 to 0.76), and  (Table 3). IRT was negatively correlated with hematological parameters (r = -0.01 to -0.71) as described in Table 4, except for neutrophil count (r = 0.32). The data showed that IRT at different body parts have different correlations with skin temperature, rectal temperature, body temperature, heartbeat, and respiration rate as well ( Fig. 2 and Table 3). The difference in the measured body parts is relevant to the heat storage and release (Kenny & Jay 2013). Fluctuations in rectal temperature and respiration rate across the observation time were also investigated by Hooper et al. (2018) with the lowest value occurred at 6 a.m. morning. Several studies (Paim et al. 2014;McManus et al. 2015;Seixas et al. 2017) reported that the lowest temperatures occurred in the morning and reached its peak at noon.
Temperature measurements at several body parts are important to identify the capacity of physiological adaptation of cattle to heat stress on the environment (Silanikove 2000). Experimental results showed a higher positive correlation between IRT testing with TB compared to that with TS or TR and no significant correlation was found between IRT with RR and HR. In this context, IRT is a possible non-invasive method to study temperature and metabolic response due to thermal stress on animals (Paim et al. 2013;Martello et al. 2016;Hooper et al. 2018). Evidence revealed that right and left legs, either front or back, showed a higher correlation with TB, TS, and TR. Piccione & Refinetti (2003) found a strong daily rhythm between TR and leg temperature due to the input rhythm from the suprachiasmatic nucleus at the hypothalamus that acts as a thermoregulating agent. D' Alterio et al. (2011) have described body extremity as the main factor in regulating heat storage or loss. Further, D' Alterio et al. (2011) report that goat's legs play important role in the mechanism related to heat loss by which these body parts have vein tissue rich in branches that make heat transfer be possibly mediated by the increase in blood flow.
A nonsignificant correlation between IRT result and HR was probably due to the absence of thermal pressure during the observation period. Another study suggests that heat stress in animals' cardiovascular systems may have an effect on the increase in heartbeat, and this effect is associated with the decrease in blood pressure that stimulates HR (Du Prezz 2000). In general, the results of IRT were negatively correlated with hematological parameters. Similarly, some studies found negative associations between ambient air temperature, as well as the temperature-humidity index (THI), and hematological parameters including RBC and HGB concentrations (Correa et al. 2012;Seixas et al. 2017). This finding was associated with hemodilution effect and low RBC count as well as HGB concentration in the animals' performing high adaptation capacity to environmental condition and showing a negative effect with heat released by their bodies (Seixas et al. 2017).  *p < 0.05, **p < 0.01, ***p < 0.001, ns = non significant. RBC = red blood cell, HGB = hemoglobin, HCT = hematocrit, MCV = mean cell volume, MCH = mean cell hemoglobin, MCHC = mean cell hemoglobin concentration, RDW = red blood cell distribution width, WBC = white blood cell, LYM = lymphocyte, MON = monocyte, NEU = neutrophil

CONCLUSION
Infrared themography (IRT) shows a curve pattern that was almost the same as the physiological parameters in young Sapera dairy goats from the beginning until the end of the observation. Significant correlations were found between physiological parameters (except HR and hematological) of goats on thermography measurements results at several body parts. The highest positive correlation (r = 0.85) was found between TB and IRT of the left rear leg. Therefore, it was concluded that IRT can be applied to measure goats physiological conditions especially body temperature.