1. Introduction
The major factor that negatively affects broiler chickens production in the subtropics and tropics is heat stress (Aluwong et al., Reference Aluwong, Sumanu, Ayo, Ocheja, Zakari and Minka2017; Goel et al., Reference Goel, Ncho and Choi2021; Sugiharto et al., Reference Sugiharto, Yudiarti, Isroli, Widiastuti and Kusumanti2017). The combined effects of high relative humidity (RH) and high ambient temperature (AT) have been reported to induce thermal stress in chickens (Egbuniwe et al., Reference Egbuniwe, Ayo, Mohammed and Aliyu2015; Jiang et al., Reference Jiang, Mohammed, Jacobs, Cramer and Cheng2020; Kim et al., Reference Kim, Lee, Kim and Lee2021; Qaid et al., Reference Qaid, Al-Mufarrej, Azzam, Al-Garadi, Albaadani, Alhidary and Aljumaah2021), therefore, lipid peroxidation resulting from hyperthermia may damage some of their vital organs (Rhoads et al., Reference Rhoads, Baumgard, Suagee and Sanders2013; Al-Zghoul & Saleh, Reference Al-Zghoul and Saleh2020; Aksoy et al., Reference Aksoy, Curek, Narinc and Onenc2021; Gogoi et al., Reference Gogoi, Kolluri, Tyagi, Marappan, Manickam and Narayan2021). Antioxidants are used universally in dietary supplementation, and these supplements are beneficial in ameliorating tissue damages induced by stress (Gouda et al., Reference Gouda, Amer, Gabr and Tolba2020; Sun et al., Reference Sun, Jiang, Xu, Zhang, Xu, Zheng and Qu2015). Fisetin is a flavonoid found in vegetables and fruits, such as grapes, onion, strawberries, and cucumbers (Khan et al., Reference Khan, Syed, Ahmad and Mukhtar2013; Chen et al., Reference Chen, Ho, Wu, Huang, Chen, Tai, Ho and Yen2015; Kikusato et al., Reference Kikusato, Xue, Pastor, Niewold and Toyomizu2021). Fisetin has several pharmacological and physiological activities that are extensive (Qaid et al., Reference Qaid, Al-Mufarrej, Azzam, Al-Garadi, Albaadani, Alhidary and Aljumaah2021). Probiotics are live microorganisms that are beneficial when adequately administered to the host (Aluwong et al., Reference Aluwong, Kawu, Raji, Dzenda, Govwang, Sinkalu and Ayo2013). They function as both antioxidant and antistress agents, and are also gut effective (Sumanu et al., Reference Sumanu, Aluwong, Ayo and Ogbuagu2021).
2. Objectives
Our objective was to determine the cloacal temperature (CT) responses of broiler chickens administered with fisetin and probiotic and exposed to heat stress. We hypothesized that supplementation of fisetin either alone or in combination with probiotic would be beneficial to ameliorate the negative effects of thermal stress in broiler chickens. To test this hypothesis, we assessed broiler chickens CT responses in separate groups of chicks given these supplements alone and in combination.
3. Methods
3.1. Ethics statement
This research was approved by the Ahmadu Bello University, Zaria’s Ethical Committee on Animal Use and Care, with the following reference number, ABUCAUC/2018/021.
3.2. Site of the experiment and experimental animal management
We conducted this experiment at the Department of Veterinary Physiology, Ahmadu Bello University, Zaria (11°10′ N, 07°38′ E). The chickens were exposed naturally to the fluctuating RH and AT conditions (Dzenda et al., Reference Dzenda, Ayo, Lakpini and Adelaiye2013). Sixty newly hatched broiler chicks that were apparently healthy served as the subjects. They were raised in an intensive management system, feed and water were provided to the broiler chicks ad libitum. On Days 1–28, broiler starter were fed to the chicks, whereas broiler finisher was given from Days 29 to 42. The poultry pen was made of concrete floor littered with wood shavings, cement block with aluminum roofing and cardboard ceiling. The dimension of the pen was 8.4 × 5.6 × 1.91 m and the birds were stocked at the density of 15 birds/m2. Protective clothing was made available to ensure optimum biosecurity measures.
3.3. The study area meteorological data and experimental design
The dry-bulb temperature (DBT) and wet-bulb temperature (WBT) (Brannan® Sapphire Instruments, New Delhi, India) were recorded every 2 hr for 3 days, 1 week apart, on Days 21, 28, and 35 of the experiment. RH was calculated using Osmon’s hygrometric table (Narinda Scientific Industries, Haryana, India). The temperature-humidity index (THI) is a measure of the degree of discomfort that the chickens experienced during the stressful season. The index is importantly an efficient temperature based on air temperature and humidity. THI was determined using the formula of Tao and Xin (Reference Tao and Xin2003): THI = 0.85(Tdb) + 0.15Twb, where THI = THI for broiler chickens, Tdb = DBT, and Twb = WBT. The parameters were recorded inside the poultry house on each day of the experiment.
Sixty chicks were divided into four groups of 15 each by simple randomization. Group I control; Group II, fisetin (Sigma Inc., New Orleans, LA) at a dose of 5 mg/kg; Group III, probiotic (Montajat Pharmaceuticals, Bioscience Division, Dammam, Saudi Arabia) at a dose of 4.125 × 106 cfu/100 ml; and Group IV, fisetin and probiotic (same doses as stated above). All administrations were performed orally for the first 7 days of life via gavage.
3.4. CT and AT measurement
CT values, which is an index of the core body temperature of the broiler chickens, were recorded (Sinkalu et al., Reference Sinkalu, Ayo, Adelaiye and Hambolu2015), using a digital clinical thermometer (Krusser Thermometer®, Amazon). CT measurements were taken using procedures of good standard (Minka & Ayo, Reference Minka and Ayo2013) over a period of 24 hr, from 07:00 to 07:00 hr of the next day, on Days 21, 28, and 35 of the study. The AT was recorded simultaneously with CT values using a WBT and DBT (BrannanSapphire Instruments, New Delhi, India). RH and THI were calculated as described by Tao and Xin (Reference Tao and Xin2003).
For data analysis, one-way analysis of variance (ANOVA) was utilized, Tukey’s multiple comparison post-hoc test was used to compare disparateness between the treatments and control groups means. Data obtained were expressed as mean ± standard error of the mean (SEM). GraphPad Prism 5.03 for windows was used (GraphPad Software, San Diego, CA). Significant values were considered to be p < .05.
4. Results
The mean DBT at Day 28 (28.85 ± 0.45°C) of the study was higher compared to those of Days 21 (26.85 ± 0.25°C) and 35 (28.54 ± 0.39°C). Day 21 (77.92 ± 2.56%) had the least RH compared to Days 28 (81.46 ± 3.11%) and 35 (79.00 ± 2.62%). The THI was highest on Day 28 (28.47 ± 0.38) as compared to Days 21 (26.40 ± 0.23) and 35 (28.05 ± 0.33; Table 1). At Day 21 of the study period, overall mean values of the CT in the fisetin group (41.20 ± 0.03°C) showed no difference significantly (p > .05) when compared with that of the control group (41.39 ± 0.03°C). Values recorded in the probiotic (40.32 ± 1.90°C) and fisetin + probiotic groups (40.49 ± 0.03°C) were lower significantly (p < .05) when compared with the control group (41.39 ± 0.03°C; Table 2). The overall mean CT values recorded at Day 28 of the study was evident that fisetin group showed no difference (41.36 ± 0.04°C), whereas the co-administered group had a significant (41.23 ± 0.03°C, p < .05) decrease when compared with the values obtained from the control group. Also, the CT values were different significantly (p < .05) in the chickens supplemented with probiotic (41.10 ± 0.06°C), when compared with the corresponding values of the controls (41.58 ± 0.03°C; Table 3). At Day 35 of the study, the overall mean CT values in the fisetin group (41.68 ± 0.03°C) did not differ when compared with that of the control group (41.78 ± 0.03°C). The overall CT values of fisetin + probiotic-administered group differed significantly (p < .05) from the control group. Nevertheless, probiotic-administered group had the least CT values which differed significantly (40.10 ± 0.11°C; p < .05) as compared to the controls (41.78 ± 0.03°C; Table 4).
Table 1. Variations in thermal environmental parameters on selected days of the study period
Values in parenthesis are minimum–maximum. n = 15.
Abbreviations: DBT, dry-bulb temperature; RH, relative humidity; THI, temperature-humidity index.
Table 2. Circadian variation in cloacal temperature of broiler chickens during Day 21 of study
Values in parenthesis are minimum–maximum.
Abbreviations: F + P, fisetin + probiotic; SEM, standard error of the mean.
a,b,cMeans with different superscript letters across rows are significantly different (p < .05).
Table 3. Circadian variation in cloacal temperature of broiler chickens during Day 28 of study
Values in parenthesis are minimum–maximum.
Abbreviations: F + P, fisetin + probiotic; SEM, standard error of the mean.
a,bMeans with different superscript letters across rows are significantly different (p < .05).
Table 4. Circadian variation in cloacal temperature of broiler chickens during Day 35 of study
Values in parenthesis are minimum–maximum.
Abbreviations: F + P, fisetin + probiotic; SEM, standard error of the mean.
a,bMeans with different superscript letters across rows are significantly different (p < .05).
5. Discussions
From the present study, it was evident that the values of DBT (26.00–36.00°C), RH (49.00–93.00%), and the THI (28.47 ± 0.38) obtained exceeded the thermoneutral zone stipulated for broiler chickens above 3 weeks of age, which are 18–24°C, 65–70%, and 20.8, respectively in the tropics. This further support the fact that the experimental period was thermally stressful to the birds. The values of THI higher than 20.8 which elicited heat stress in broiler chickens agrees with previous findings (Sinkalu et al., Reference Sinkalu, Ayo, Adelaiye and Hambolu2015). Broiler chickens’ energy balance and fitness might be affected negatively during the prevailing thermally stressful season as recorded by previous research (Tao & Xin, Reference Tao and Xin2003; Yin et al., Reference Yin, Tang, Liu, Cao, Xie and Zhang2021). This might subsequently lead to low immunity, decrease performance, suppression, high morbidity, and death; hence, antioxidants, such as fisetin and probiotic could be helpful during the season when administered adequately. Oxidative stress is a resultant of heat stress which consequently, enhances reactive oxygen species (ROS) production (Ogbuagu et al., Reference Ogbuagu, Aluwong, Ayo and Sumanu2018; Shakeri et al., Reference Shakeri, Oskoueian, Le and Shakeri2020; Sumanu et al., Reference Sumanu, Aluwong, Ayo and Ogbuagu2019), therefore, the findings of this study serves as the basis for modulating the detrimental effects of heat stress in animals via fisetin and/or probiotic administration.
Little or no information have been published to the best of our knowledge, on the use of fisetin or its combination with probiotic in modulating the detrimental effects of thermal stress in poultry, using CT as a potent biomarker, during the cause of this study. It is very difficult for the chickens to maintain core body temperature amidst high AT, as they are void of sweat glands and rely more on evaporative cooling (panting) to keep their body cool. Several workers have reported CT increase in older chickens during heat stress (Robinson et al., Reference Robinson, Lida, Jairo, Luciano, Keller and Lina2016; Sahebi-Ala et al., Reference Sahebi-Ala, Hassanabadi and Golian2021). Fluctuations in body temperature depicts the stressful nature of RH and AT, the mechanism of thermoregulation is furthermore required for the maintenance of homeothermy (Makeri et al., Reference Makeri, Ayo, Aluwong and Minka2017; Yin et al., Reference Yin, Tang, Liu, Cao, Xie and Zhang2021). The greater the fluctuation of environmental parameters, the more prominent the thermal stress, and the higher the negative effects upon health and optimal growth (Lee et al., Reference Lee, Park, Heo, Choi and Seo2021).
At Days 21, 28, and 35 of the study, the CT values recorded in the control group were higher than the values recorded within the treatment groups. This may be attributed to the fact that the birds were naturally exposed to heat stress and their endogenous antioxidants were not potent enough to mitigate its adverse effects. These findings are in agreement with previous studies in the tropic (Egbuniwe et al., Reference Egbuniwe, Ayo, Mohammed and Aliyu2015). The CT values recorded in the fisetin-administered group were higher than the values recorded in the probiotic and the co-administered groups, respectively. It may be deduced that fisetin is devoid of antistress effect in mitigating heat stress in broiler chickens as evident in this study. This further rejects the hypothesis which was earlier stated that fisetin administration would be advantageous in ameliorating the detrimental effect of heat stress in broiler chickens. Values of CT recorded in the probiotic group during the period of study were lower as compared to all other groups. It could be speculated that probiotic was able to function not only as an antioxidant, but also as an antistress agent in mitigating heat stress, hence it could be potent in enhancing thermoregulation which reduces the metabolic heat production of the chickens, vis a vis increasing their productivity. Previous studies on broiler chickens demonstrated similar findings (Aluwong et al., Reference Aluwong, Sumanu, Ayo, Ocheja, Zakari and Minka2017; Sugiharto et al., Reference Sugiharto, Yudiarti, Isroli, Widiastuti and Kusumanti2017). The co-administered group had CT values that were lower than that of the control and fisetin groups, respectively during the study period. This may be attributed to the thermoregulatory effect of the probiotic and not the fisetin. Nevertheless, this is the first time that fisetin, a phytonutrient, would be tried on food animals to the best of our knowledge. Therefore, fisetin alone was not potent in alleviating heat stress in broiler chickens.
The molecular mechanism by which probiotic was able to elicit thermoregulation should be further investigated.
6. Conclusion
CT was significantly increased by the thermal environmental parameters, indicative that the study period was thermally stressful. Therefore, the administration of fisetin singly was not potent in eliciting antistress effect in broiler chickens exposed to thermal stress. Although, probiotic was potent in alleviating heat stress, hence its use singly is advocated.
Acknowledgments
Profs J. O. Ayo and T. Aluwong are highly appreciated for designing this research and for their mentorship all through the study. Their constructive criticism and meticulous corrections in bringing out the best in me and this research can never be underestimated.
Authorship Contributions
J.O.A. and T.A. conceived and designed the study. V.O.S., T.A., J.O.A., and N.E.O. conducted data gathering, V.O.S. performed statistical analyses, V.O.S., J.O.A., and T.A. wrote the article.
Data Availability Statement
The authors confirm that the data supporting the findings of this study are available within the article.
Conflict of Interest
The authors declare no conflict of interest.
Funding Statement
This research received no specific grant from any funding agency, commercial or not-for-profit sectors.
Open access
Comments
Comments to the Author: Dear Editor,
Thank you for considering me as a reviewer for this publication in your esteemed journal Experimental Results. I have provided my comments as follows.
The authors have achieved their objective and the manuscript is, generally speaking, well-written.
1- There are some grammar issues in L11 (There is a death), L19 (The probiotic-supplemented), L90 (CT values were), L127 (These findings are), L135 (also as an anti-stress).
2- In L79 and 80 (add the full name of the DBT and RH). In addition, please delete all the means values throughout the result in the text since you have mentioned them in the tables, this will be a repetition of the result.
3- In L114 (change the sentence to “fisetin or its combination with probiotics”).
4- Could the authors explain what are these numbers between brackets in all tables, which is very confusing? since the authors have provided the mean values ± SEM.
Thank you
Mohammed Ibrahim