Effects of ProbChick® on the immunological response after new castle virus using LaSota stain vaccination in broiler

1-        Effect of ProbChick on the immune response and production properties.

2-        ProbChick enhance the production properties of broiler chickens.

3-        ProbChick enhance the immunological response after vaccination.

Introduction

Probiotics are live bacteria, fungi, or yeasts that replenish the gastrointestinal tract's flora and aid in maintaining a healthy digestive system, which promotes bird growth and is an alternative to antibiotics; probiotics are becoming more widely used in poultry diets (1). Probiotic is derived from two Greek words, pro and biotic, which mean for life (2). In 1965 Lilly and Stillwell were the first to use the word (3); according to a joint FAO/WHO work group, Probiotics are live microorganisms that, when supplied in suitable proportions, impart a health benefit to the host (4). Elie Metchnikoff was the first scientist who discovered that the microbiota present in the intestine plays a vital role in maintaining a healthy body when he found that the Lactobacillus bacteria that produce lactic acid present in fermented milk products were able to increase the longevity of Bulgarian peasants (5). Primarily, live apathogenic bacterial strains in animals and poultry are generally considered probiotics: Lactobacillus acidophilus, L. sporogenes, L. bulgaricus, and Streptococcus thermophilus; Bacillus subtilis, and Saccharomyces cerevisiae (6). Probiotics have various action mechanisms, such as inhibiting all pathogens by producing organic acids and antibacterial substances such as hydrogen peroxide, bacteriocins, and defensins (7). Probiotics compete with pathogenic bacteria on intestinal epithelial binding sites and essential nutrients (8); they enhance the immune response by releasing regulatory T cells, effector T and B cells, and antigen-presenting cells (9). ProbChick^® is a food additive for poultry with multiple beneficial bacteria: Lactobacillus Plantarum, L. sporogenes, L. acidophilus, Streptococcus thermophilus, Bacillus subtilis, Bifidobacterium bifidum, and Saccharomyces cerevisiae. They also contain digestive enzymes, which maintain good gut flora, leading to a high rate of feed utilization, increasing weight gain of the broiler, inhibiting the growth of pathogenic bacteria in the gut, and enhancing the broiler's immune response (10). Probiotic supplementation improved the immune response to the Newcastle disease virus (11). Also, it can boost macrophage and natural killer cell activity, regulate cytokine and immunoglobulin release, and help maintain the integrity of the intestinal epithelial barrier (12). In addition, B lymphocytes will be activated, transforming into antibody-secreting plasma cells, which will then infiltrate the gut-draining lymphoid tissue (13). We aimed in the current study to investigate the effect of ProbChick^® as an immune enhancer during the vaccination process in broilers.

Materials and methods

Experimental designs

A total of 200 one-day-old chicks were included in the current study and divided randomly into five groups (40 chicks in each group), the first group was considered the control group left without any treatment all over the experiment. The second group (positive control) ProbChick^® administration in drinking water from day one at a dose of 1 gram/liter, and the third group vaccinated with live attenuated la sota virus vaccine in drinking water on day 7 of age at dose 2.5*10⁶ (EID). The fourth group was vaccinated with la sota virus at day 7 of age at a dose of 2.5*10⁶ (EID) and of ProbChick^® at day 14 of age at a dose of 1 gram/liter. The fifth group (protective group) was vaccinated with la sota virus on day 7 at a dose of 2.5*10⁶ (EID) and administration of ProbChick^® on the same day at a dose of 1 gram/liter. Ten birds from each group were euthanized at the end of each first, second, third, and fourth week of age. A thin blood smear was stained with Wright-Giemsa to calculate the stress index (14). Weekly body weight, weekly feed consumption, and food conversion ratio (15,16).

Newcastle vaccine

A live, lyophilized vaccine was used for Newcastle virus, La sota strain, containing 10⁶ virus/dose. The vaccine was used according to the manufacturer's instructions. A vaccine package containing 1000 doses was used. It was dissolved in ten liters of clean, cold water, and 2 g/liter of skimmed milk was added to stabilize and increase vaccine efficacy. Chicks were thirsty for 3 hours before vaccination (17).

Stress index

This test was used to identify the effect of E. coli infection and administration of probiotic ProbChick^® on non-specialized cellular immunity in chicks. This index was calculated by staining blood slide smear with Wright- Giemsa stain. The number of lymphocytes and heterophils was recorded for each slide, and the stress index was calculated as follows: heterophils/lymphocytes (18).

Weight gain, FCR, and relative weight of spleen and bursa

The chicks' weight was measured and recorded on the first day of hatching using an electronic scale, and then the results of the weight gain of all chicks were recorded every week and throughout the experiment period to get the weight of the remaining food as compared to the consumed food (19). At the end of the week, the rate of feed consumption was calculated for each group, then the feed conversion ratio (FCR) measures an animal's efficiency in converting feed into increased body mass (19). Spleen and bursa of Fabricius were removed and cleaned of adherent tissues. The weight of each was measured and expressed as relative to final body weight by organs weight/body weight*100 (20).

Newcastle antibody titer

The titer of antibodies against Newcastle disease was measured using the indirect ELISA, where 5 μl of the whole blood was taken and placed in a dilution plate, 200 μl of the dilution solution was added, then 10 μl of the dilution dish was taken. It was placed inside the test plate, then 90 μl of the previous diluted solution was added to it to reach a dilution ratio of 1:500. The test plate was incubated for 30 minutes at a temperature of 22-27ºC. The plate was washed three times, and then 100 μl of the conjugating solution was added and incubated for 30 minutes at a temperature of 22-27ºC. After that, the plate was washed three times, and 100 μl of substrate solution was added and incubated for 15 minutes, and then 100 μl of stopping solution was added. The plate result was read with an e-reader at wavelength 405 (21).

ProbCkick^®

ProbChick^® contains 10 billion CFU/gram strains of beneficial bacteria like Lactobacillus Plantarum, Lactobacillus sporogenes, Lactobacillus acidophilus, Streptococcus thermophilus, Bacillus subtilis, Bifidobacterium bifidum, and Saccharomyces cerevisiae.

Statistical analysis

The means included in the current study were analyzed using one-way ANOVA, with Duncan's post-Hock test as P<0.05.

Results

Average body weight

The effects of the LaSota vaccine and probiotic ProbChick^® are shown in table 1. On day one, no significant variation of P

Table 1: Effects of LaSota vaccine and probiotic on weekly average body weight

-Different letters within the same column mean statistically significant differences at P<0.05.

Body weight gain and feed conversion rate

The average weekly weight gain and feed conversion rate FCR are summarized in table 2. The effects of LaSota vaccine and ProbChick^® on weekly weight gain at week 1 no significant variation P

Table 2: Effects of LaSota vaccine and probiotic on weekly body weight gain and FCR

-Different letters within the same column mean statistically significant differences at P<0.05.

Spleen relative weight

The effects LaSota vaccine and ProbChick^® on spleen relative weight are shown in table 3. Week 1 showed no significant difference between groups at P

Table 3: Effects of LaSota vaccine and probiotic on spleen relative weight

-Different letters within the same column mean statistically significant differences at P<0.05.

Bursa of Fabricius (FB) relative weight

The effects of the LaSota vaccine and ProbChick^® on relative weight are shown in table 4. On weeks 1, 2, and 3, the G3, G4, and G5 are significantly higher P

Table 4: Effects of LaSota vaccine and probiotic on Bursa of Fabricius relative weight

-Different letters within the same column mean statistically significant differences at P<0.05.

Stress index

The effects LaSota vaccine and ProbChick^® on stress index are shown in table 5. On day one, old chick, no significant variation P

Table 5: Effects of LaSota vaccine and probiotic on relative kidney weight

-Different letters within the same column mean statistically significant differences at P<0.05.

New castle antibody titer

The effects of ProbChick^® on the Newcastle antibody titer range are shown in table 6. Day one and week one showed no significant variation P

Table 6: Effects of probiotic on antibody titer for Newcastle virus using indirect ELISA test

-Different letters within the same column mean statistically significant differences at P<0.05.

Discussion

The Newcastle vaccine in broilers did not affect the average weekly weight, weight gain, and feed conversion ratio compared to the control group. The mechanism through which the probiotic works in improving the weight gain and, as a result, the feed conversion ratio is that giving the beneficial bacteria in poultry will increase in number in the small and large intestines, which helps significantly in improving the quality of the natural flora in the intestines of these birds and improving its ph in the middle of the intestine which is Close to neutral. These changes improve the food digestion and absorption from the intestine and its introduction into the blood circulation of poultry (22).

As for the relative weight of the spleen and the bursae of Fabricia, the results of the current study indicated that giving the probiotic has no effect on the relative weight of these two organs in the group given the probiotic only compared to the control group. The groups that received the vaccine dose against Newcastle disease Using the Lasota strain alone or with the probiotic showed an increase in the relative weight of each of the spleen and bursae of Fabricia compared with the control group and the group given the probiotic alone. This result was in agreement with many other studies that indicated that the poultry vaccination process increases the organs' weight. Primary immunoglobulins such as the spleen, the bursae of Fabricia, and the thymus gland will be responsible for the immune response and the production of antibodies that help immunize the bird through vaccination (23,24).

The results of the current study indicated that giving the probiotic reduced the stress factor in the groups that were given the probiotic alone or on the same day of vaccination or seven days after it compared to the control group. The immune system will, in turn, cause the production of additional amounts of antibodies in the bird's body (25).

The results of the current study indicated that the level of antibodies in the group that was given the Newcastle vaccine and the probiotic on the seventh day caused an increase in the level of antibodies. They were observed in the second week, and then there was a gradual decrease in the following weeks, but the level of decrease in the concentration of antibodies in the blood was less than What was observed in the other groups. The natural flora in the intestines of birds plays an essential role in the formation and function of the immune system in poultry (26), as the commensal bacterial present in the intestine and adjacent to the intestinal cells have an essential role in stimulating the intestinal immune tissue (27). Moreover, it is also due to the oral administration of probiotics in poultry that activates the spleen's natural killer cells and stimulates these cells' phagocytic properties (28). In addition to the birds that were given probiotics containing yeast and lactobacilli bacteria whose serum contained a higher concentration of antibodies against sheep red blood cells (as an immune stimulus) than the birds that did not receive these probiotics (29), and the administration of probiotics had a significant effect on the response. Immunogenicity increases the antibody concentration when immunizing broiler chicks against Newcastle disease, infectious bronchitis, and bird flu (29).

Conclusions

We conclude that adding ProbChick^® to the broiler at 1 g/litter of drinking water will enhance the immune response during and after vaccination. This will improve the efficiency and titer of antibody production.

Acknowledgments

The authors wish to thank the College of Veterinary Medicine, University of Mosul, Mosul, Iraq, for supporting the current study.

Conflict of interest

No conflict.