Application of streptococcus thermophilus in preparation of microecological preparation for preventing and / or relieving acute heat stress response of chicken

Abstract

The invention provides an application of streptococcus thermophilus in preparation of a microecological preparation for preventing and / or treating a heat stress reaction of chicken, and particularly provides a microecological preparation for preventing and / or treating the heat stress reaction of the chicken. Experimental animal models prove that the microecological preparation has the effects of relieving core temperature rise and blood biochemical index fluctuation caused by chicken heat stress, reducing oxidative stress and eliminating lipid peroxidation, has the advantages of no drug resistance, low cost, simplicity in operation, high benefit and the like, and can be used as a microecological preparation for relieving chicken heat stress.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and in particular to the use of thermophilic streptococci in the preparation of a microecological preparation for preventing and / or alleviating acute heat stress reactions in chickens. Background Art

[0002] Since poultry is covered with feathers and has no sweat glands, its body temperature regulation function is weaker, and it is more sensitive to high temperature environments, making it prone to heat stress reactions. Studies have shown that heat stress not only causes a significant deterioration in the physiological functions of poultry (such as physiological and biochemical indicators, immune function, and antioxidant levels, etc.) and reduces production performance, but also affects the intestinal health of poultry and the composition, structure, and functional stability of intestinal microorganisms. The intestine is an organ that connects the body of poultry to the external environment, and it is also an important area for absorbing nutrients. The intestinal microorganisms play a vital role in ensuring intestinal health and normal function. Although studies have identified differential intestinal microorganisms and metabolites that change significantly under heat stress, their functions, effects, and effects are not clear and need further research.

[0003] Livestock and poultry suffer from heat stress during the production process. The existing drug formula can only alleviate some of the adverse symptoms caused by heat stress in a targeted manner, but cannot systematically and comprehensively alleviate the series of adverse effects caused by heat stress in chickens. In addition, most drug formulas may produce drug resistance and have residual risks. At present, drugs for treating or relieving heat stress are still under research.

[0004] As antibiotics pose an increasing threat to animal husbandry and humans, "antibiotic ban" has become a hot word in the global animal husbandry industry. my country has officially banned the addition of antibiotics to feed since July 1, 2020. Finding a substitute for feed antibiotics has become a top priority.

[0005] Probiotic preparations refer to products that improve the microecological balance in the host organism by adding or regulating the microbial community. Such preparations usually contain ingredients such as probiotics, prebiotics or symbiotic bacteria. Probiotic preparations can increase the number of beneficial bacteria in the host's intestines and inhibit the growth of harmful bacteria, thereby promoting the balance of intestinal microecology. Probiotic preparations can also improve the host's immune function and enhance resistance. Existing studies have shown that probiotic preparations have shown positive effects in the prevention and adjuvant treatment of some diseases such as enteritis and diarrhea. At the same time, compared with drug treatment, probiotic preparations usually have better safety and tolerability, and fewer side effects. In general, probiotic preparations have important application value by regulating microbial communities and promoting host health.

[0006] Streptococcus thermophilus is one of the most important probiotic lactic acid bacteria (LAB). It is currently widely used in the production of yogurt and cheese because it can produce a large amount of extracellular polysaccharides during the fermentation process, which increases the viscosity of fermented milk and improves the quality of yogurt. Streptococcus thermophilus also has some functional activities, such as producing bacteriocins and lactic acid. The polysaccharides, bacteriocins, lactic acid, etc. produced by it have anti-tumor activity, which resists the occurrence of tumors by activating the body's immune system and inhibiting cell mutation. In addition, Streptococcus thermophilus can produce superoxide dismutase, which can remove excess superoxide anion free radicals produced during metabolism in the body and delay aging. Studies have found that Streptococcus thermophilus has good acid and bile salt resistance, can survive in higher concentrations of bile, and can reach the distal end of the small intestine without being damaged by bile and low pH environment in the body, which lays a good foundation for it as a microecological preparation. Patent CN105961845A discloses that thermophilic streptococci can be added to chicken feed as an antibiotic substitute to improve the immune function of laying hens. Patent CN114601073A discloses that by comparing feed with and without compound bacterial agents, it is concluded that compound bacterial agents can reduce the level of immunoglobulins in the blood of laying hens, that is, compound bacterial agents have the effect of improving immunity. Summary of the invention

[0007] The present invention aims to solve at least one of the technical problems existing in the prior art to at least a certain extent.

[0008] The present invention proposes the use of Streptococcus thermophilus in the preparation of microecological preparations. According to an embodiment of the present invention, the microecological preparation is used for preventing and / or alleviating the application of acute heat stress damage. Streptococcus thermophilus, a Gram-positive bacterium, is a species of Streptococcus in prokaryotic microorganisms, purchased from Beina Biology. When studying the intestinal flora of laying hens before and after heat stress, the inventor found that the abundance of the bacteria in the intestinal tract of laying hens after acute heat stress was higher than that in the intestinal tract of laying hens before heat stress, and it was inferred that it may have an anti-heat stress effect. Further, by giving Streptococcus thermophilus to laying hens for a period of time and then applying heat stress treatment, the symptoms of heat stress can be effectively alleviated. Thus, it is shown that Streptococcus thermophilus has the effect of preventing and / or alleviating acute heat stress damage, and is simple and safe to use, with good application prospects, and provides new ideas for the prevention and treatment of heat stress in clinical practice.

[0009] Therefore, the present invention proposes a use of thermophilic streptococcus in preparing a preparation for alleviating acute heat stress reaction in chickens.

[0010] The probiotic preparation is used to alleviate the increase in core temperature of chickens caused by acute heat stress. The probiotic preparation is used to alleviate the increase in blood pH caused by acute heat stress in chickens. The probiotic preparation is used to alleviate the disruption of carbon dioxide balance caused by acute heat stress in chickens. The probiotic preparation is used to alleviate the disruption of blood ion balance caused by acute heat stress in chickens. The probiotic preparation is used to alleviate lipid peroxidation caused by acute heat stress in chickens.

[0011] The microecological preparation is administered orally orally, and the dosage is controlled at 2 mL / day.

[0012] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention.

[0013] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) the present invention uses a single liquid preparation of thermophilic Streptococcus, which is administered by intragastric administration, effectively alleviating adverse reactions such as increased core temperature, increased blood pH, disrupted carbon dioxide balance, disrupted blood ion balance, and lipid peroxidation caused by acute heat stress in chickens, thereby enhancing the chickens' ability to resist acute stress; (2) the bacterial agent of the present invention is easily absorbed after entering the body, and can be colonized in the intestines of chickens after continuous intragastric administration for 14 days; (3) the use process of the present invention is simple, suitable for group administration, suitable for production, and saving manpower and material resources. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The above and / or additional aspects and advantages of the present invention will become apparent and easily understood from the description of the embodiments in conjunction with the following drawings, in which:

[0015] Figure 1 The relative abundance analysis of Streptococcus thermophilus in the feces of laying hens at different times of gavage in Example 1 (D0 before gavage, D14, 14 days after gavage) is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. Data are expressed as mean ± standard deviation. Different lowercase letters (a, b) represent significant differences (p<0.05);

[0016] Figure 2 The core temperature change analysis diagram of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. The data are expressed as mean ± standard deviation. Different lowercase letters (a, b, c) represent significant differences (p<0.05);

[0017] Figure 3 The blood pH value analysis chart of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. Data are expressed as mean ± standard deviation. Different lowercase letters (a, b) represent significant differences (p<0.05);

[0018] Figure 4 The TCO2 change analysis chart of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. The data are expressed as mean ± standard deviation. Different lowercase letters (a, b, c) represent significant differences (p<0.05);

[0019] Figure 5 The PCO2 change analysis chart of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. Data are expressed as mean ± standard deviation. Different lowercase letters (a, b) represent significant differences (p<0.05);

[0020] Figure 6 The following is an analysis of the changes in HCO3- concentration in laying hens before and after heat stress treatment in Example 1. One-way analysis of variance (Welch's ANOVA test) was used for significance tests among multiple groups, and the Benjamini-Hochberg method was used for multiple comparisons. Data are expressed as mean ± standard deviation. Different lowercase letters (a, b, c) represent significant differences (p<0.05);

[0021] Figure 7 The graph showing the changes in iCa levels in laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. Data are presented as mean ± standard deviation. Different lowercase letters (a, b, c) represent significant differences (p<0.05);

[0022] Figure 8The TG change analysis chart of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. The data are expressed as mean ± standard deviation. Different lowercase letters (a, b) represent significant differences (p<0.05);

[0023] Fig. 9 The MDA change analysis diagram of laying hens before and after heat stress treatment in Example 1 is shown; the significance test between multiple groups was performed using one-way analysis of variance (Welch's ANOVA test), and multiple comparisons were performed using the Benjamini-Hochberg method. The data are expressed as mean ± standard deviation. Different lowercase letters (a, b, c) represent significant differences (p < 0.05). DETAILED DESCRIPTION

[0024] The scheme of the present invention will be explained below in conjunction with the embodiments. It will be appreciated by those skilled in the art that the following embodiments are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used are not indicated by the manufacturer and are all conventional products that can be obtained commercially.

[0025] The bacteria used in the following experiments were Streptococcus thermophilus, purchased from BeNa Culture Collection, with strain number BNCC362702. NCBI identified it as Streptococcus thermophilus strain CS18, Accession CP030928.1.

[0026] Example 1

[0027] 1. 115 healthy laying hens aged 45 weeks with a body weight range of 1300±100g were randomly divided into 4 groups: intragastric bacteria group (STH group), intragastric saline control group (NaCl group), blank control group (Control group) and heat stress control group (HS group), with 35, 30, 20 and 30 chickens in each group, respectively.

[0028] 2. Treatment method of the intragastric bacteria group (STH group): intragastric administration was performed at 4 pm every day for 14 consecutive days, and water and feed were stopped half an hour before and after the treatment.

[0029] The temperature of all laying hens was controlled at 22±2℃, and the light intensity was controlled at 16L for 8 days. The laying hens were free to eat and drink water at other times. The ingredients of the feed were corn, soybean meal, bran, vitamin and mineral premix, etc.

[0030] The microecological preparation for oral administration was prepared once a week. The preparation method was as follows: the frozen preserved strains were revived, identified, and then propagated. The bacteria were precipitated by low-speed centrifugation and adjusted to an appropriate concentration using sterile 0.75% saline. After the preparation, the plate spread counting method was used to ensure that the number of viable bacteria was greater than 1×10 8 CFU / mL.

[0031] Chickens in the STH group were gavaged with 2 mL of STH microecological preparation diluted with 0.75% sterile saline per day. After the gavage period, they were subjected to heat stress at 36°C. Chickens in the NaCl group were gavaged with 2 mL of sterile 0.75% saline per day. After the gavage period, they were subjected to heat stress at 36°C. Chickens in the Control group were negative controls and were not subjected to any treatment.

[0032] The feces of the three groups were collected before, 7th and 14th days after the gavage treatment, and the colonization of thermophilic Streptococcus was detected. Chickens in the HS group: no gavage treatment was performed, only 36℃ heat stress treatment was performed.

[0033] 3. Heat stress treatment: The STH, NaCl, and HS groups were subjected to acute heat stress treatment at 36°C for 6 hours on the 15th day. After treatment, the core temperature was measured, blood was collected from the sub-wing vein, and changes in blood gas-related indicators were detected. Serum was separated and changes in biochemical and Elisa indicators were detected. The control group was subjected to the above sample collection and testing under normal temperature.

[0034] The heat stress heating room is constructed and enclosed with polyurethane foam to ensure the heat insulation effect. It is equipped with a heating plate for heating. The temperature of the heating plate can be controlled in the range of 20-40℃. A circulating fan is set above the heating plate to ensure that the hot air circulates in the entire space after the heating plate is heated up, and the temperature difference of the entire vertical plane is kept within 1℃ as much as possible. The heating is controlled by a sensor to turn the power on and off. The heating temperature is 36℃. The temperature sensor temperature is set to start at 35.5℃ and shut down at 36.5℃ to ensure that the temperature in the chicken house is within the range of 36±1℃. The heat treatment humidity is controlled between 65% and 70%, and the treatment method of ground sprinkler and air humidifier is adopted.

[0035] Experimental results:

[0036] like Figure 1The results showed that there was no significant difference in the relative abundance of thermophilic Streptococcus in the feces of the three groups of laying hens before the start of gavage treatment (D0); after 14 consecutive days of gavage treatment (D14), the relative abundance of thermophilic Streptococcus in the feces of laying hens in the STH group was significantly higher than that in the other groups, and thermophilic Streptococcus colonized in the intestinal segments.

[0037] Table 1

[0038]

[0039] like Figure 2 The results showed that after 6 hours of heat stress treatment at 36°C, the core temperature (rectal temperature) of chickens in the HS group was significantly higher than that in the Control group; the core temperature of chickens in the NaCl group was also significantly higher than that in the Control group, and there was no significant difference between the core temperature of chickens in the STH group and those in the HS group; after intervention with thermophilic Streptococcus colonization, the core temperature of chickens in the STH group was significantly lower than those in the HS and NaCl groups, and the significant increase in core temperature of chickens after acute heat stress was alleviated.

[0040] Table 2

[0041]

[0042]

[0043] like Figure 3 The results showed that after 6 hours of heat stress at 36°C, the blood pH of chickens in the HS and NaCl groups increased significantly compared with the Control group, which was related to respiratory alkalosis. The blood pH of chickens in the STH group was normal, with no significant difference from the Control group, and the increase in blood pH of chickens after acute heat stress was significantly inhibited.

[0044] Table 3

[0045] Group Category Blood pH Control Group 7.491±0.058 HS group 7.585±0.062 NaCl group 7.554±0.055 STH group 7.488±0.068

[0046] like Figure 4 and Figure 5 The results showed that after 6 hours of heat stress at 36°C, the total carbon dioxide (TCO2) and carbon dioxide partial pressure (PCO2) levels of chickens in the HS and NaCl groups were significantly reduced, and the blood carbon dioxide balance was disrupted. The TCO2 and PCO2 of chickens in the STH group were significantly higher than those in the above two groups, but there was still a significant difference between them and the Control group, indicating that thermophilic Streptococcus alleviated the decrease of TCO2 and PCO2 after acute heat stress.

[0047] Table 4

[0048]

[0049] like Figure 6 and Figure 7 The results showed that after 6 hours of heat stress at 36°C, the blood ion balance of chickens in the HS and NaCl groups was disrupted, and HCO 3- The HCO and iCa concentrations in the blood of the chickens in the STH group were significantly decreased. 3- The reduction degree of iCa concentration in the H1N1 group was significantly lower than that in the above two groups, but there was still a significant difference between it and the Control group. Streptococcus thermophilus alleviated the destruction of blood ion balance after acute heat stress to a certain extent.

[0050] Table 5

[0051] Group Category <![CDATA[HCO 3- Concentration mmol / L]]> iCa concentrationmmol / L Control Group 23.51±2.24 1.63±0.16 HS group 20.50±2.61 1.36±0.06 NaCl group 20.35±2.96 1.35±0.10 STH group 21.85±2.02 1.47±0.17

[0052] like Figure 8 and Fig. 9 The results showed that after 6 hours of heat stress at 36°C, lipid peroxidation occurred in the chickens in the HS and NaCl groups, and the levels of triglyceride (TG) and malondialdehyde (MDA) in their serum were significantly reduced and increased, respectively. However, after heat stress, the TG level in the serum of the chickens in the STH group was not significantly different from that of the non-heat stressed Control group, and the MDA level was significantly lower than that of the Control group. Thermophilic Streptococcus can eliminate lipid peroxidation after heat stress.

[0053] Table 6

[0054] Group Category Triglyceride (TG) mmol / L Malondialdehyde (MDA) mmol / L Control Group 1.88±0.04 15.01±1.31 HS group 1.80±0.07 16.32±1.61 NaCl group 1.77±0.08 16.29±1.97 STH group 1.87±0.04 13.90±1.35

[0055] In summary, the chickens colonized with Streptococcus thermophilus effectively alleviated the increase in core temperature, decrease in blood pH, decrease in TCO2 and PCO2 levels, disruption of ion balance, and lipid peroxidation levels caused by acute heat stress. This provides a theoretical basis for Streptococcus thermophilus to be used as a probiotic microecological preparation for the prevention and treatment of heat stress.

Claims

1. Use of thermophilic streptococci in the preparation of a microecological preparation for preventing and / or alleviating acute heat stress reaction in chickens.

2. A microecological preparation for alleviating acute heat stress in chickens, characterized in that: The preparation contains the thermophilic streptococcus according to claim 1, and the viable count of the thermophilic streptococcus is greater than 1×10 8 CFU / mL.

3. The microecological preparation for alleviating acute heat stress in chickens according to claim 2, characterized in that: The microecological additive also includes an auxiliary material, and the auxiliary material is maltodextrin.

4. The microecological preparation for alleviating acute heat stress in chickens according to claim 2 or 3, characterized in that: The additive is in the form of liquid, powder or granule.

5. The microecological preparation for alleviating acute heat stress in chickens according to claim 4, characterized in that: The microecological preparation is prepared by resuscitating, identifying and amplifying the thermophilic streptococcus, and then adjusting the concentration to a suitable concentration using 0.75% sterile physiological saline to obtain a liquid.

6. The microecological preparation for alleviating acute heat stress in chickens according to claim 5, characterized in that: The administration method of the microecological preparation is oral gavage, and the dosage of the microecological preparation is 2 mL / day.

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