Study on Heat Stress Behavior and Blood H/L Ratio of Ducks Fed Diets Containing Mix Strains of Probiotics in Commercial Farms

: An experiment has been conducted with the purpose of examining the effect of two kinds of probiotic on the behavior and blood H/L ratio of local ducks kept in commercial farms. The study was conducted in collaboration with the ‘Berkah Abadi’ duck farmer group which keeps the bird under an intensive dry system. The intensive system refers to the ordinary way done by the farmers in which ducks were confined around the farmer’s village with a closed fence so the birds had no access to the outside area, and the amount of feed provided could be controlled and measured. The location was in the coastal area of Tegal City as one of the most famous duck centers in Indonesia. The materials used were 13.4+1.3 months old local laying ducks which were reared by the farmer group. The study used a Completely Randomized Design with treatments consisting of 3 doses of P and P+ probiotics respectively. P contained Lactobacillus sp. combined with Saccharomyces cerevisiae, whereas P+ comprised Lactobacillus sp., Saccharomyces cerevisiae, Streptococcus thermophilus, and Lactobacillus bulgaricus. There were 6 treatment units which were replicated 4 times, so in total there were 24 flocks. Each flock had 50 female ducks; therefore this study involved 1,200 ducks. The treatments were applied each morning, and mixed thoroughly in duck ration. Data collected were heat stress-related behavior, body temperature, and H/L ratio. It was concluded that P and P+ had similar effects on the behaviour and H/L ratio of local ducks kept under commercial management.


Introduction
In a tropical country like Indonesia, heat stress is one of the most important environmental stressors challenging poultry production.The detrimental effects of heat stress on poultry have been discussed and studied in depth intensively.Effects of heat stress on broilers and laying hens, for instance, range from reduced growth and egg production to decrease poultry and egg quality and safety (Lara and Rostagno, 2013).Duck, as a homeotermic animal, is susceptible to heat stress which leads to decreasing welfare and productivity.A previous study that compared wet and dry systems of duck husbandry concluded that under dry intensive systems, local ducks suffered from heat stress as indicated by Suggested Citation Suswoyo, I., Tugiyanti, E., & Ismoyowati. (2023).Study on Heat Stress Behavior and Blood H/L Ratio of Ducks Fed Diets Containing Mix Strains of Probiotics in Commercial Farms.European Journal of Theoretical and Applied Sciences, 1(6), 511-517.DOI: 10.59324/ejtas.2023.1(6).51rectal temperature, behavior, and body and plumage condition (Suswoyo et al, 2014).Therefore, releasing heat stress is an essential factor in improving duck welfare.Manipulation on daily management is required to protect the ducks from suffering from heat stress, among others by using mixed strains of probiotics as functional feed.
Several studies have been conducted to investigate the use of probiotic for the alleviation of heat stress in poultry.In broiler chickens, probiotic Lactobacillus strains have proven to be able to restore the microbial balance and maintain the natural stability of indigenous bacterial microbiota following heat stressinduced changes (Lan et al., 2004).Under heat stress conditions, chickens fed with diets containing Lactobacillus cultures have higher antibody production than those on the control diet (Zulkifli et al., 2000).Relating to heat stress, probiotics might be useful for ameliorating the adverse influence of heat on egg production and the gut health of laying hens (Deng et al., 2012).Based on the microbial content, probiotics could be single-strain or multi-strain probiotics.Multistrain probiotics are composed of more than one species or strains of bacteria and sometimes, include fungal species with benefits to human and animal health.Multispecies preparations have advantages compared to single-strain probiotics and, to a lesser extent, multi-strain probiotics (Barton, 2000).
The H/L ratio is commonly used to measure the level of strong physiological stress in birds.Some evidence suggests that the H/L ratio is positively correlated with the strength of the innate immune response (Maccoll et al., 2017).
Information on duck behavior and H/L ratio relating to heat stress is lacking, especially those under commercial duck management.

Method
The study used an experimental method which has been conducted in collaboration with 'Berkah Abadi' duck farmer group keep the bird under a dry system intensively.Under an intensive system, the birds are mostly kept in sheds with rice straw bedding and solid floor ranch in front of the sheds (Setioko dan Rohaeni, 2001).The study site was in Tegal City which is located in the northern coastal area of Java Island.The City and its surrounding areas are one of the most important duck centers in Indonesia with its very famous local laying duck namely Tegal Duck.It is believed that the duck belongs to the Indian Runner family.

Materials
The materials used were local laying ducks which the age of 13.4+1.3months.The intensive system refers to the ordinary way done by the farmers in which ducks were confined around the farmer's village with a closed fence so the birds had no access to the outside area, and the amount of feed provided could be controlled and measured.Drinking water was provided ad libitum three times a day i.e. in the morning, noon, and afternoon, while feed was given twice a day i.e. in the morning and in the afternoon.The feedstuffs were locally available which mainly consisted of rice bran, dried rice, and fresh fish (Leiognathidae) with proportions of 39,65 %, 25,11 %, and 35,24 % respectively.The nutrient content was 26,38 % crude protein, 2.923 kcal/kg metabolic energy, 2,29 % calcium, and 0,78 % phosphorus.
The study used a Completely Randomized Design with treatments consisting of 3 doses of P and P+ probiotics respectively.P contained Lactobacillus sp.combined with Saccharomyces cerevisiae, whereas P+ comprised Lactobacillus sp., Saccharomyces cerevisiae, Streptococcus thermophilus, and Lactobacillus bulgaricus, while doses 1, 2, and 3 were 3, 5, 7 ml/l.There were 6 treatment units which were replicated 4 times, so in total there were 24 flocks.Each flock had 25 female ducks; therefore this study involved 1,200 ducks.The treatments were applied each morning, and mixed thoroughly in duck ration.

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Body temperature: measuring rectal temperature; 10 ducks from each flock were measured their rectal temperature.

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Daily ambient air temperature and humidity as indicators of environmental condition were measured at 6 am, noon, and 3 pm.

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Blood H/L ratio Hematological examination was carried out by taking blood samples taken from two ducks in each flock.Blood was taken from the wing vein (brachial vein), 3 ml with a syringe, then put into a sterile tube containing anticoagulant (EDTA/Ethylene Diamine Tetraacetic Acid), labeled according to the duck number, the tube was put into a thermos and taken to the laboratory.

Data Analysis
Data collected were analyzed using Variance Analyses at 5% confident, and descriptive analyses.

Heat Stress Relating Behavior
Heat stress-related behavior is presented in the table below.Table 1 shows that drinking was the highest response of the ducks during this study.Lara and Rostagno (2013) reported that birds subjected to heat stress conditions spend less time feeding, more time drinking and panting, as well as more time with their wings elevated, less time moving or walking, and more time resting.This finding indicated that most of the ducks suffered from heat stress.By comparing the treatments, it looks like ducks with P were less stressed than those with P+ supplementation.Heat stress significantly affects daily behavior, including feeding, drinking, lying, standing, and walking (Li et al., 2015).The behavior of chickens can significantly influence their growth rate, consequently influencing production costs (Neves et al .2014).

Body temperature
The body temperature of ducks as indicated by rectal temperature during the study is presented in Table 2.  (Smith, 1976).
However, these finding was close to the upper level for normal body temperature.The body temperature of domestic poultry is maintained within a relatively narrow range that is usually reflected by the upper and lower limits of a circadian rhythm in deep body temperature.
After birds are exposed to a high ambient temperature, their body temperature rises above their normal body temperature.High heat caused tissue damage and an increase in the workload of the physiological system (Aengwanich et al., 2003).Statistical analyses indicated that the data were not significantly (P>0,05) different.It implies that P and P+ had similar effects on duck body temperature.

Daily Ambient Air Temperature and Humidity
The environmental condition at the study site is presented in Table 3. Table 3 indicates that the environmental temperature of the study site was 28,22+0,71 o C on average which was higher than the maximum temperature needed by poultry.The study site which was a coastal region of Tegal City is considerably suffering from climate change.At the same time relative humidity was also high with an average of 83,91+3,70%.It seems that the environmental condition in the area was higher than the thermo-neutral zone required by ducks.Thermo-neutral zone for poultry is between 18 to 25ºC, and the most efficient temperature for ducks ranges between 23 to 25 o C (El-Badry et al., 2009).
Controlling the ducks' environment, particularly temperature, humidity, litter moisture, and ammonia is crucial to duck welfare (Jones and Dawkins, 2010).If the ambient temperature is higher than the thermoneutral zone, panting will increase ten times which decreases productivity (Ahmad and Sarwar, 2005).Qualitative observation during the study showed that ducks with treatments had more comfort conditions indicated by less drinking and panting activities.This situation becomes worse with a natural phenomenon i.e. climate change.Climate change is one of big the issues that concern the world.It is understood that climate change has a severe impact not only on human beings but also on animals.Indonesian National Aviation and Space Agency reported that the climate in Indonesia has become warmer during the 20th century with an average annual temperature increased by about 0.3°C since 1900 (LAPAN, 2012).

H/L ratio
The blood H/L ratio of the ducks during the study is presented in the table below.P + 1 0,81+0,07.5.
P + 1 0,76+0,01 The H/L ratio in this study ranged from 0.65+0.04 to a high of 0.81+0.07.The results of this study are lower than the results of previous research which obtained an average figure of 1.45 + 0.2 in dry systems (Suswoyo et al., 2016).This result is also still lower than the results of previous research which reported that in the confined method, the average H/L ratio was 2.32 (Suswoyo and Ismoyowati, 2010) (Minias, 2019).Supplementation of probiotics in the diet significantly increases white blood cell count and decreases the H/L ratio which is important in the reduction of stress effect in poultry (Rahimi and Khaksefidi, 2006).

Conclusion
It was concluded that P and P+ had similar effects on behavior, body temperature, and H/L ratio of local ducks kept under commercial management.

Table 2 . Rectal Temperature under Treatments
These temperatures inTable 2 were in the normal range which is 38-42 o C

Table 4 . The Blood H/L Ratio of the Ducks during the Study
Kramer (1998)l., 2014)havior of ducks show that during the day most of the ducks suffer from heat stress as indicated by the number of ducks panting.The results of this research are in accordance with the results of previous research that during the day 36-70% of ducks experience panting(Suswoyo et al., 2014).Kramer (1998)stated that, in terms of immune function, birds with initially low H/L ratios have stronger antibody responses to Brucella abortus than those with high H/L ratios.In his research, Wang (2022) concluded that chickens with low H/L ratios are more resistant to Salmonella Enteritidis.Chickens with low H/L are superior to chickens with high H/L in terms of survival, immune response, and resistance to Salmonella infection