Introduction
Poultry rearing has become a vital economic activity in many countries. In intensive poultry production, newly hatched chicks are more prone to infections due to the slow normal microflora colonization in the intestine. The situation is due to less chance of contact with their mother and acquires microflora from the environment. Due to production pressure, the broiler chickens succumb to stresses, which adversely affects their performance. Under such circumstances, the use of synthetic antimicrobial agents and antibiotics help to mitigate stress and improve feed efficiency and growth. However, with the possibility of antibiotics ceasing to be used as growth promoters in poultry in the coming days, both consumers and manufacturers are looking for alternatives. Europe and South Korea banned the use of antibiotics as growth promoters since 2006 and 2012, respectively, and such a ban affects the rest of the world. For sustainable animal production systems, the development of antibiotic alternatives is the need of the hour. Therefore, the concept of probiotic supplementation emerges, which is highly helpful to fill this gap.
What is Probiotic?
Probiotics are the live microbial feed supplement that beneficially improves the intestinal microbial balance of the host animal. The most used species with two exceptions in probiotic (intestinal strains) preparations are Lactobacillus species (L. acidophilus, L. bulgaricus, L. casei, L. lactis), Streptococcus thermophilus, Enterococcus faecalis, Enterococcus faecium, Bacillus subtilis, and Bifidobacterium spp. The exceptions are Lactobacillus bulgaricus & Streptococcus thermophilus are yogurt starter organisms.
Other types of probiotics such as Bacillus spores (direct-fed microbial – DFM) can be included as feed additives in poultry diets due to their heat resistant capacity and long shelf life. Generally, Bacillus spp. have a distinct advantage over Lactobacillus and Bifidobacterium as DFM, because Bacillus spores are resistant to low pH, bile salts, and other harsh conditions encountered in the gastric environment. Bacillus spores promote gut health by competitive exclusion and produce antimicrobial peptides that are cytotoxic to bacterial pathogens and reduce symptoms associated with enteric infectious diseases, such as necrotic enteritis. Some Bacillus (Bacillus subtilis PB6) strains produce antimicrobial substances with broad-spectrum activity against various strains of Campylobacter and Clostridium species.
Commercially, chickens reared to hatch in clean incubators devoid of organisms commonly found in the gut. Also, shell microbiological contamination may influence gut microflora characteristics. The hydrochloric acid (HCl) gastric secretion that starts at 18 days incubation has an impact on microflora selection. Thus, immediate probiotic supplementation at birth is vital in avian species than in other animals.
Ideal Characteristics of a Probiotic
An ideal probiotic should be characterized as mentioned below.
- Source – animal/ bird origin
- Nonpathogenic and nontoxic
- Resistant and persistent to stress, processing, storage, and gastric acid & bile
- Antimicrobial activity
- To overcome pelleting temperatures and be compatible with most feed additives
- Suitable adherence factors to attach in intestinal epithelium or mucus and compete for binding sites
- Genetically stable and viable at the high population
Probiotic Colonization
Establishing and maintaining healthy gut microflora is essential, which improves the microbial environment of birds’ intestinal tract by displacing harmful bacteria. The gut is sterile in newly hatched chicks and acquires microflora from the environment. Chicks may get infected at this time since pathogenic microbes may multiply faster than beneficial bacteria. However, as the days progress, post-hatch, the microflora stabilizes and attains a balance between the ‘favorable’ and ‘harmful’ bacteria. This balance may be affected by internal factors like stress or surrounding environment like infectious pressure. At this stage, the concept of probiotics supplementation emerges that is helpful for gut microflora. Survival and stability of the microbial strains used, their relationship with the host, host health, stress, nutritional status, dose, usage frequency, age, and genetics are the depending factors for successful colonization of probiotics. The measured colony forming units (CFU) describes the increased colonization at the beak, and that progressed distally to the colon. Small intestines have anaerobes with a population ranging from 104 to 108 CFU/ml (Lactobacilli, Streptococci, and Enterobacteria). The maximum bacterial population, accounting for 1010 to 1013 CFU/ml, has been recorded in the colon and caecum.
Mechanism of Action
The probiotics work by competitive exclusion mechanism. Nurmi and Rantala, et. al., 1973, successfully demonstrated that newly hatched chicks develop resistance to Salmonella colonization in the gut by providing a suspension of gut content from healthy adult chickens. Probiotics exclude the pathogen replication sites by competing for the common nutrients in the chicken gut. Competitive exclusion refers to the physical blocking of opportunistic pathogen colonization and altering the environmental niches within the intestinal tract like intestinal villus and crypts that leads to a better immune system. The addition of a nonpathogenic culture by competitive exclusion, either single or multiple strains, reduces the pathogenic bacteria in the GIT. Competitive exclusion by probiotics includes competition for physical attachment sites, host immune system enhancement, and production of antimicrobial compounds like short-chain fatty acids and bacteriocins or colicins from metabolic reactions.
The epithelial barrier enhancement increases adhesion to the intestinal mucosa, modulation of the immune system, and production of antimicrobial substances that are another mechanism of action by probiotics. Probiotics showing antimicrobial effect provides a frontline of defense against the adverse effects of pathogens. For example, Bacillus subtilis PB6 is a natural strain isolated from the healthy chicken gut, shown broad-spectrum activity against various strains of Campylobacter and Clostridium spp. by producing antimicrobial substances in vitro. Some Lactobacillus strains inhibit Shigella and Yersinia virulence factors by directly reducing their invasiveness.
Mechanism of action in probiotics to modulate the immune system mostly depends on the strains of microorganisms or bacteria used, probiotic preparation method, the environment where birds reared, and the route of administration. Through the interaction of host and the probiotic cultures, many studies observed enhancement of both natural and specific antibodies, interferons, or cytokines, as well as activation or suppression of T-cells that lead to the cytokine expression.
The health benefits afforded by probiotics include the formation of low molecular weight compounds (<1,000 Da), such as organic acids, and the production of antibacterial substances termed bacteriocins (> 11,000 Da). Organic acids, acetic acid, and lactic acid have a strong inhibitory effect against gram-negative bacteria, which are the antimicrobial compounds responsible for the inhibitory activity of probiotics against pathogens.
Many Lactobacillus produce antibacterial peptides, including bacteriocins and small antimicrobial peptides (AMPs). Bacteriocins (lactacin B from L. acidophilus) with narrow-spectrum activity act against closely related bacteria. Whereas, some bacteriocins are active against food-borne pathogens. The bacteriocin mediated killing includes the target cell destruction by pore formation and inhibition of cell wall synthesis.
Probiotic Effect on Necrotic Enteritis (NE)
Clostridium perfringens, a gram-positive and spore-forming anaerobe cause an enterotoxemic disease named necrotic enteritis (NE). C. perfringens is commonly found in GIT and develops necrotic lesions on the gut wall resulting in the mortality of poultry. Feed contaminated with C. perfringens implicates outbreaks of necrotic enteritis in chickens. Studies showed that healthy chicken has a relatively low number of C. perfringens in GIT, while the increase in the bacterial concentration correlates with the necrotic enteritis condition.
Usage of antibiotic growth promoters to some extent helped in reducing the incidences of NE. However, isolation of antibiotic-resistant strains of C. perfringens from chickens, a significant shift in consumer attitudes, legislation in the European Union towards raising food animals without drugs and medicines led to search of other non-antibiotic alternatives. Kemin formulated CLOSTAT™, an active microbial strain to inhibit C. perfringens, contains a naturally occurring strain of Bacillus subtilis PB6 isolated from the intestinal tract of a healthy chicken. CLOSTAT™ is a patented proprietary strain that is closely associated with the intestinal epithelium and able to tolerate gastric and bile conditions. The CLOSTAT™ strain can form spores that help in the strains’ survival during pellet formation and for broiler feed production.
Bacillus subtilis PB6 produces a specific bacteriocin in the GIT that has inhibitory activity against pathogenic strains of Clostridium perfringens and Campylobacter spp; produces specific surfactins that have efficient anti-inflammatory properties. Bacillus subtilis PB6 significantly improves the intestinal morphology, growth performance, carcass traits, and control necrotic enteritis in broilers.
Effect of Probiotics on Immune system
Using probiotics regularly stimulates the humoral and cell-mediated immunity through enhanced production of natural interferons/ cytokines, increased macrophage, lymphocyte, and natural killer (NK) cell activity. It upregulates oxidative burst in heterophils and increases immunoglobulin (IgG, IgM, and IgA). Probiotics produce a gut-stabilizing effect and immune regulation through balanced control of pro-inflammatory and anti-inflammatory cytokines.
Probiotics inhibit the growth of infectious organisms by increasing the number of lymphocytes and lymphoid cells in lamina propria and intraepithelial lymphocytes (IEL) in the small intestine. Cell-mediated immunity stimulation reduces the flock mortality caused by immunosuppressive diseases [infectious bursal disease (IBD), chicken infectious anemia, reoviral infection, Mareks’ disease, and mycotoxins] and fights against viral infection. Their metabolites act as immunomodulatory agents by activating specific and non-specific immune responses of chicks, which in turn enable them to prevent various infectious diseases. Feeding probiotics could improve antibody titers against Infectious bursal disease and Newcastle disease (ND). By increasing the immune status and checking/ preventing enteric infections (bacterial, coccidian), probiotics could help in alleviating the losses due to secondary infections in birds, observed during viral diseases or immunosuppressive conditions.
Probiotics enhance the resistance of birds and partially protect from the ill effects on growth associated with pathogenic microbes. Probiotics help to reduce the pathogenic microbial load in the intestine, thereby reducing the risk of spread in the house through fecal contamination. A multi-strain probiotic use in time (before the infection sets in) and regularly in feed helps to prevent various infectious agents like bacterial, fungal, protozoan, and viral agents.
Chickens fed with dietary Bacillus subtilis PB6 for 28 days tend to display better growth performance and pronounced intestinal morphology, including consistent cell mitosis, extended cell area, and prominent villus height, compared to those fed on a control diet. In a challenge study with C. perfringens, B. subtilis PB6 supplemented diet, broilers had significant feed conversion ratio (FCR) and increased intestinal villi length between 10.88% and 30.46% over infected control. The study demonstrated that Bacillus subtilis PB6 supplementation can improve the host gut physiology and intestinal health in the presence of pathogens.
Application of Probiotics
Probiotics maintain the proper balance of beneficial microbial population in the intestine of birds, which is vital for productivity, growth, efficient feed conversion, and stimulation of birds’ immune mechanisms. The mechanism of the action of probiotics in the poultry production system include: competitive exclusion of harmful bacteria/ pathogens, production of antibacterial peptides like bacteriocins and small AMPs, establishing and maintaining a healthy gut microflora, improved digestion, and utilization of nutrients, decreased pH, the release of various antibacterial substances, toxins neutralization, competition for nutrients with pathogens, reduced ammonia production, and immune system stimulation. Effective probiotic accelerates the development of normal microflora in chicks and poults, improves egg production, weight, and size, and helps in better poultry performance.
Conclusion
Probiotics usage helps to improve gut health by reducing enteric pathogens in poultry. Probiotics act as growth promoters, immunostimulants, and optimize the microflora balance in the avian gut. Bacillus subtilis spores can be included as feed additives in poultry diets due to their remarkable heat stability and long shelf life. Bacillus subtilis spores promote gut health, not only by competitive exclusion but also by producing antimicrobial peptides that are cytotoxic to bacterial pathogens and reduce symptoms associated with enteric infectious diseases, such as necrotic enteritis. Bacillus subtilis PB6 controls C. perfringens induced necrotic enteritis in broiler birds and improves gut health and gut integrity by increasing the villus height and villus height to crypt depth ratio. Studies proved that probiotics are a potential alternative to antibiotic growth promoters used in the poultry industry.
by Dr. Srinivas J, Kemin Industries
