Importance of Probiotic for prevention of avian diseases

Abstract:
Disease prevention measure is the second line of defense against diseases challenge after
providing good biosecurity and a hygienic environment. Maintaining chickens free of
diseases is one of the world's significant health challenges to maintain populations' health.
For this purpose, instead of antibiotics, new biological preventative strategies such as
probiotics have been introduced to prevent various avian diseases. Probiotics are live
microorganisms used as feed additives or supplements in animal diets. This review mainly
focuses on some probiotics properties that significantly affect the host to prevent several
avian diseases. Such as Probiotics are non-pathogenic to host and have antagonistic effects
on different microorganisms, including enhancement of the function of the intestinal
epithelial barrier, competition for adhesive receptors, competition for available nutrients,
Production of Antimicrobial Substances, degradation and neutralization of toxins,
immunomodulatory effects, decrease in the growth of pathogenic microorganisms,
decrease the parasitic load in the gut lumen, limiting the amount of ammonia production
and reducing heat-related stress in poultry.

1- Introduction

Probiotics are non-pathogenic living microorganisms, which positively affect the hosts’
health if they enter the body in sufficient amounts. Currently approved Food and Agriculture
Organization/World Health Organization definition of a probiotic is “live microorganisms
which, when administered in adequate amounts, confer a health benefit on the host.” (1.).
More precisely, Probiotics are live microorganisms that are of non-toxic and non-
pathogenic nature (2).
Probiotics can be used to enhance health and well-being in livestock, birds, and human.
Today it is generally accepted that probiotics are unique living microbial cultures that provi
de benefits to a given individual’s body. (6) These living organisms may be bacteria, fungi, or

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yeasts. They are isolated from a healthy adult animal’s gut, typically of the same species to
which the probiotics will be given (3).
probiotics have various species, but the most important ones including, Lactobacillus
bulgaricus, L. acidophilus, L. salivarius, L. lactis, L. casei, L. helveticus, L. plantarum, L.
bulgaricus, Enterococcus faecium, Enterococcus faecalis, Streptococcus thermophilus,
Streptococcus thermophilus Bifidobacterium spp. and E. coli, all of them are intestinal strains
except Lactobacillus bulgaricus and Streptococcus thermophilus, (8,9,11) Some types of
probiotics are microscopic fungi such as yeast belonging to Saccharomyces cerevisiae spp.
(10)

Many types of research indicate that probiotics suppress diseases in chickens and reduce the
shedding of pathogens. (4) A variety of these studies included pretreatment with probiotics
before challenge or at the same time as a challenge. (5) The beneficial effects of probiotics
can differ between strains, so selecting the most effective ones would be necessary to
prevent specific diseases.
Probiotics have essential beneficial properties that are non-pathogenic, adherent to
intestinal cells, and can attach to intestinal cells. (7) Also, probiotics can colonize and
multiply in the host, have the opportunity to be host-specific, resist the passage through
the digestive tract and tolerance to stomach acid and bile, and produce metabolites that
inhibit or destroy harmful bacteria. (4) This review describes some characteristics of ideal
probiotics that have an essential role in probiotics’ disease prevention abilities.

2- Effect of probiotics on gut wall

Probiotics have various effects on enteric pathogens, intestinal morphology, and health.
These probiotics generally inhibit in the lower gut and have a significant impact in preventing
most enteric diseases. (18) Probiotics, including Lactobacillus spp. may have a defensive
effect against enteric pathogen by stabilizing the gut mucus membrane, growing gut mucus
secretion, improving gut motility, (16), and influencing the intestinal microbiota’s structure
and activity. (17)

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2.1. Adherence and colonize to Intestinal Epithelial Cells

Various researches have shown that lactobacilli with a high rate of epithelial adherence
provide the most health benefits for disease prevention. Lactobacillus species that adhered
to chicken crop epithelial cells in vitro, improved growth rates, and tolerance to gram-
negative bacterial infections have resulted. (13) The intestinal epithelium selectively
encourages the ingestion of nutrients but prohibits bacteria from accessing the bloodstream.
L. Plantarum and L. reuteri, when complemented with broiler feed, improved the integrity of
that form of barrier and suppressed such opportunistic or pathogenic bacteria’s entry. (12)
Theories on why adherence to the probiotic effect is essential for disease prevention to
include the need for resident bacteria to overpopulate the intestinal tract when the crop
empties, availability of a ready supply of probiotics to inoculation incoming food,
competition with epithelial binding site pathogens, and cross-talk with epithelial cells to
modulate cell activity and morphology. (14)
One research investigated The inhibitory effects of two different adherent lactobacilli on
epithelial binding to three distinct types of Salmonella. Lactobacillus fermentum reduced
Salmonella Pullorum’s attachment by 77%, while Lactobacillus animalis reduced Pullorum’s
attachment by 90%, S. Enteriditis, 88%, and S. Gallinarum by 78%. (15) Some studies founded
that some strains of lactobacillus inhibited gut pathogens by occupying the intestinal wall.
Fuller found that Lactobacillus was able to decrease E. coli in the crop and small intestine.
(13) other studies indicated that L. salivarius strains decrease the number of Salmonella
enteritidis and Clostridium perfringens. (23)
Probiotics can increase and colonize in the digestive tracts. Blocking receptor locations and
preventing other pathogens’ attachment, including dangerous bacteria such as
enteropathogenic Salmonella or E. E. coli.

2.2. Enhancement of Epithelial Barrier Function

The intestinal epithelial lining plays an essential role against luminal content, such as
pathogens. It serves as a selective barrier for gut contents, Impairment of epithelial barrier
function resulting increase susceptibility of the host to various infectious diseases. (19) many
studies reported the beneficial effects of probiotics on birds’ intestinal morphology and
structures, reported that probiotics affect villus height and crypt depth or number of goblet
cells in the small intestine, and enhancement epithelial barrier integrity (20,40) Increased
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villus height with increased absorption surface area of villi increases absorption of luminal
nutrients and results in improved growth and health. (21) changes in the morphology of GIT
due to Lactobacillus adherence provide a larger surface area for the absorption of nutrients
and increase resistance to the invasion of pathogenic microorganisms. (22)
The alteration produced by probiotics in the epithelial cells causes a decrease of the luminal
PH, antimicrobial activity, and production of antimicrobial peptides that block bacterial
invasion and adhesion to the epithelial cells. On the other side, they maintain the intestinal
membrane, increasing tolerance, and neutralizing dangerous microorganisms. (52)

3- Bacterial Antagonism

The presence of acid-forming probiotics in the gut has significant antagonistic effects against
pathogenic bacteria. Probiotics can secrete and produce such bacteriostatic and bactericidal
substances. Probiotics are capable of producing substances that inhibit pathogens’ growth
for both Gram-positive and Gram-negative bacteria. Also, certain Lactobacillus strains may
produce hydrogen peroxide, which possesses intense bactericidal action on various
pathogens. (24)

Different probiotic strains can generate compounds with bactericidal and bacteriostatic
consequences such as bacteriocins, hydrogen peroxide, and various organic acids. These
substances have a significant effect on harmful bacteria. (25)
Probiotics have been demonstrated to suppress pathogen activity. A variety of antimicrobial
factors, including bacteriocins, have been released. And a variety of acids, such as lactic and
acetic acids, acidify the lumen. Short-chain fatty acids may invade gram-negative pathogenic
bacteria’s outer membranes, contributing to inhibiting the pathogen’s development. (28)
Bacteriocins are active proteins produced by some strains of probiotics in different
environments. (26)
Bacteriocins exhibit powerful antimicrobial activities against various viruses, bacteria, and
fungi, considering significant structural and characteristic variations and have been
recognized as a modern generation of antimicrobials to prevent and control foodborne
pathogens. (27) The typical bacteriocin-mediated killing mechanisms include the
degradation of pathogenic organisms by pore-forming, inhibition of cell wall synthesis, and
destruction of RNA, DNA, and protein synthesis. (38) Reported that Bacillus spp. Produce
antimicrobial peptides (AMP) that are cytotoxic to Eimeria spp., thereby reducing avian
coccidiosis and colonization of Clostridium perfringens. (39)
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 4- Competitive Exclusion of Pathogenic Microorganisms

Competitive Exclusion (CE) is a particular form of probiotic technique that includes adding
a non-pathogenic bacterial culture to the chicken’s intestinal tract to decrease the
concentration of pathogenic bacteria in the gut. (29) This preventive care method controls
pathogenic microorganisms in poultry, especially Day-old chickens, which are young and
hence susceptible to various infectious diseases. (34)
CE in poultry promote the utilization of naturally occurring intestinal microbes in poults and
chicks that have been placed in the brooder area. inoculating a competitive exclusion
microflora to day-old chicks effectively demonstrates intestinal microbiota’s effect on
intestinal function and disease tolerance (30)

Nevertheless, instead of introducing one or a few bacterial species to existing

microorganisms, the competitive exclusion method immediately provides the chick with an
adult intestinal microbiota. Inoculating day-old chicks with competitive exclusion cultures
serves as a successful model for determining these microorganisms’ modes of action and
efficacy. Because of the sensitivity of newly hatched chicks to illness, this method is also of
industrial significance. (31).
Competitive Exclusion is a very successful and effective way of preserving chicks, poults,
pheasants, quails, and pet birds from Salmonellosis and other gut pathogenic. (32)
Competitive Exclusion is the most powerful and safe approach possible for the prevention
GIT disorders in poultry. The preventive method is wholly biological. This method was
initially developed in poultry to successfully prevent and treatment of Salmonella infection.
Later it was extended to include human pathogenic Escherichia coli, Clostridium perfringens,
Listeria, and Campylobacter. (33)
Another role of competitive Exclusion is the subsequent competitive absorption of essential
nutrients essential for pathogen development. The quicker the absorption of nutrients such
as carbon, glucose, and iron provides probiotics with an opportunity to reduce pathogen
growth. (35) Bacillus spp. Produce Siderophores compound, that Quickly uptake iron by
probiotics, plays an essential role in Pathogen exclusion. (36) However, the reduction in
pathogen concentration correlated with the inclusion of probiotics in the chicken gut is an
example of this process in vivo. (37)

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 5- Modulation of the Immune System

The avian Immune system plays a vital role in defense against pathogenic invasion in the
body and subsequent disease prevention and treatment. Probiotics have an essential effect
on the stimulation of poultry’s immunity system against invasive pathogens. Probiotics can
induce innate and adaptive immunity through modulation of Toll-like receptor expression,
activation of dendritic cells, natural killer cells, increased T-helper cells’ responses,
development of cytokines, and immunoglobulins. (41, 42) Probiotics increase the number of
lymphocytes in lymphoid tissues associated with the intestine, such as payer’s patches and
intestinal mucosal cells, therefore supplying local immunity by IgA and plasma cell secretion.
(43)
Two forms of the immune-modulating produced by prebiotics in poultry: (a) from the
microbiota, which passes through the intestinal wall and reproduces to a small degree, (b)
the contaminants produced by the dead and contaminated cells are ingested and thereby
activate the immune system. (49)
Studies reported that peptidoglycans and lipopolysaccharides are released in the gut lumen.
During infection, this release is increased and occurs to build and maintain local immune
function against pathogens. They cause the mucosa immune system’s epithelial cells’
chemotaxis and cause alterations in the intestinal epithelial cells. The chemotaxis activity is
generated by mediators including metalloproteins, prostaglandins, cytokines, and nitrogen
reactive metabolites, elevating the development of IgA, IgM and IgG, activating growth and
division of lymphocytes like NK, CD3, CD4 and CD8, enhancing the movement of lymphocyte
T and generating the interferon. (51, 51)
Newly hatched chicks have a sterile GIT system. Before developing their antibody by their
organisms, pathogens from the environment begin to localize in the digestive tract. The
establishment and activation of the gut microbiota’s growth are primarily influenced by the
humoral and cellular gut-associated immune system greatly influenced by the gut
microbiota. (44)
One study demonstrated NuPro yeast extract’s effect on the intestinal immune system to
increased intestinal IgA production that binds to antigens (such as C. perfringens α-toxin)
prevents them from passing through the mucosal membrane and developing infection and
lesions. Evaluated the effect of NuPro yeast in reduction intestinal C. perfringens levels in
broiler chicks. (45)
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(Elhasany, Hams.) reported that Lactobacillus Plantarum and Lactobacillus Acidophilus
stimulate the immune response and have the ability to reduce the adverse effect of
Salmonella Enteritidis infection in broiler chicks. (46) (Eid, Samah et. All) demonstrated the
immunomodulatory effect of Lactobacillus subtilis for preventing and controlling Necrotic
Enteritis in broiler chicken rather than its prevention by antibiotics. (47) (Knap I, eat all)
approved immunostimulatory results of Bacillus licheniformis for preventing Necrotic enteritis
in chicken. Bacillus licheniformis may be specifically used to avoid antibiotic-
resistant infections in poultry. (48)

6- Effect of probiotics on Gut Microbiota

Intestinal microbiota plays an essential role in growth, immunity, and disease tolerance.
Alterations in the microbial environment adversely effect on growth performance,
production, and disease levels of chickens. The activity and composition of gut microbiota
may be influenced by various factors, including infectious and non-infectious factors. (53)
Probiotics can enhance the equilibrium of intestinal microbiota in the host by Decreasing the
incidence of pathogenic organisms and growing the prevalence of beneficial ones. (54)

Probiotics have a substantial effect on the composition and functioning of the gut
microbiota, including competition with other microorganisms for binding sites, nutrients,
and receptors on the intestinal mucosa and inhibit the overgrowth of harmful organisms.
(58)

Several studies demonstrated the beneficial effects of probiotics on the microbiome, enzyme
activities, and fermentation activity in broiler chickens’ digestive tract. (55, 56, 57) a further
study demonstrated that supplementation of B.toyonensis and B. bifidum decreases fungal
growth and E.coli populations. (59)

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 7- Production of Antimicrobial Substances

Probiotics can inhibit pathogenic growth by releasing numerous antibacterial substances

such as bacteriocins, defensins, and lactic and acetic acids, which drop the luminal ph. Short-
chain fatty acids destroy gram-negative bacterial outer membranes causing bacterial
effectiveness. (60) Antimicrobial compounds like bacteriocins produced by lactic acid
bacteria, including plantaricin, lactacin B, and nisin, have narrow activity against some
bacterial strains. But certain bacteriocins are also effective against foodborne pathogens.
Mechanisms of bacteriocin-mediated killing include damaging of cells by pore-forming and
inactivation of cell wall synthesis. (61, 62)

8- Decrease parasitic load

Lee et al. (63) Demonstrate that in-feed supplementation of Pediococcus-Based Probiotic

enhances bird’s resistance against coccidiosis, especially effective against Eimeria tenella and
E. acervulina. They identified that orally feeding 0.1 percent probiotics decreased the
shedding of oocysts in broilers, Eimeria-specific antibody levels were significantly increased
in the same experiment. Compared with the un-supplemented control birds. This experiment
founded that probiotics are an effective preventive method for removing coccidiosis in the
birds. Kyung eat all. (64) Found that dietary Bacillus used reduced the incidence and clinical
signs of Eimeria maxima infection in broiler chickens and improved their natural host’s innate
and acquired immunity. This experiment founded that probiotics are an effective preventive
method for removing coccidiosis in the birds. Further study found the effect of probiotic
supplementation on broiler chickens’ performance experimentally infected with a mixed
infection of E. maxima, E. acervulina and E. tenella. Probiotics improve both performance in
growth and disease prevention. (65)

9- Elimination of toxic compounds in the gut

Probiotics can improve health and vigor by reducing the toxicity produced in the gut from
many Compounds such as ammonia, aflatoxins and ochratoxins.

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Ammonia production in poultry farms can adversely affect chicken’s growth, feed
conversion, carcass condemnation rate, birds’ immune system, and susceptibility to various
diseases. (69) Probiotics are used as one preventive method for removing ammonia in
poultry farms. Bacillus Subtilis produce subtilin, which can decrease urease production
generated by microbiota in the gut lumen. Thus ammonia release decrease. (66)
Consumption of ammonia by Bacillus spp. is another mechanism for reducing ammonia in
the gut, eliminating ammonia toxicity resulting from hydrolyzed uric acid. (67) Pezzuolo et
al., 2019 found that litter treatment with probiotic bacteria would considerably minimize
ammonia in a broiler farm. (68)
Aflatoxins are mycotoxins produced by Aspergillus species, cause various diseases in chicken
and other species, and are common in poultry feed. Probiotics are used as a biological
strategy to prevent aflatoxins in poultry farms.(70) Fan et al. (2015) found that the
supplementation of B. subtilis ANSB060 significantly decreased the amount of aflatoxin levels
in broiler chickens’ duodenum and prevented aflatoxicosis. (71) Another research selected
supplements with probiotics for removing aflatoxin and proved its in-vivo efficacy in
Japanese quails. (72)

Ochratoxin is a nephrotoxic mycotoxin produced chiefly by Aspergillus ochraceus and

Penicillium viridicatum in grains and feed. Ochratoxicosis mainly causes kidney disease but
also affects the liver, immune system, and bone marrow. (73) Biological detoxification of
ochratoxins by probiotics is a promising method for preventing ochratoxins in the poultry
farm.

Śliżewska K. et al. (2014) found that probiotics containing Lactobacillus strains and yeasts
Saccharomyces cerevisiae effectively detoxify ochratoxin A added to a feed. (74)
Piotrowska M . et al. (2005) examined 29 strains of lactic acid bacteria of the Lactobacillus
and Lactococcus genera, demonstrating that all strains can decrease Ochratoxin A levels in
the liquid medium. (75) in another study, Piotrowska M. (2014) found that three species of
lactic acid bacteria: Lactobacillus plantarum, L. brevis, and L. sanfranciscensis can remove
and absorb ochratoxin A. (76)

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 10- Effect of probiotics on heat-related stress

Heat-related stress in poultry farms has become a severe issue. Causes adverse effects on
birds’ physiology, immunology, and microbiology, resulting in abnormal physiological
functions and impaired performances. (77) Several studies found the effect of dietary
supplementation of probiotics on reducing heat stress in poultry.

Dietary Supplementation of Bacillus subtilis could help improve intestinal epithelial

characteristics and microflora balance of heat-stressed broiler chickens. But there was no
improvement in Birds’ thermotolerance. However, heat stress mortality was decreased. Also,
intestinal microflora environment and epithelial cells structure are well regulated, it can
reduce the harmful effects of heat stress on broilers health and efficiency. (78) another study
found dietary supplementation with either man-nan-oligosaccharide (MOS) or Lactobacillus-
based probiotic (LBP) alone or combined can minimize cyclic heat stress’s adverse effects
broilers. (79)

Conclusions

Over the past five decades, antibiotics are used mostly to reduce harmful bacterial diseases
and then as preventive measures in poultry farms to control diseases. Therefore, antibiotic
resistance and the rise in diseases are becoming a significant problem in the poultry
population, with the poultry industry now approaching natural products such as probiotics.
There is a significant impact of probiotics on the prevention of various diseases. The
antagonistic effect of probiotics on pathogens is illustrated by several mechanisms, including
competition for binding receptors, competition for nutrient uptake and inhibitory substance
production, improvement of intestinal barrier function, toxin degradation, and neutralization
of immune-stimulatory effects. Probiotics are also used as antibiotic alternatives to limit the
chances of infections caused by various pathogenic bacteria such as: such as Salmonella,
Campylobacter, Clostridium perfringens. E. coli, and elimination effects of many harmful
toxins like Aflatoxin and Ochratoxin. This review show that probiotics successfully used in
avian feed for the prevention of many diseases by modulation of intestinal microflora,
pathogen inhibition, and immunomodulation.
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References:

1. FAO/WHO. Health and nutritional properties of probiotics in food including powder milk with live lactic
acid bacteria. Report of a Joint FAO/WHO Expert Consultation on Evaluation of Health and Nutritional
Properties of Probiotics in Food Including Powder Milk with Live Lactic Acid Bacteria; FAO/WHO:
Amerian Córdoba Park Hotel, Córdoba, Argentina, 2001; pp. 1-34.

2. Guillot, J.F. Les probiotiques en alimentation animale. Cah. Agric. 1998, 7, 49-54.

3. R.U. Khan & S. Naz (2013) The applications of probiotics in poultry production, World's Poultry Science
Journal, 69:3, 621-632, DOI: 10.1017/S0043933913000627

4. Jeanne Marie Smith "A Review of Avian Probiotics," Journal of Avian Medicine and Surgery 28(2), 87-94,
(1 June 2014). https://doi.org/10.1647/2012-031

5. Smith JM, Selim SA. Performance of neonate cockatiels given cockatiel derived Lactobacillus. Proc Annu
Conf Avian Vet. 2003:157–159.

6. Fesseha H. Probiotics and its potential role in poultry production: A review. Vet Med Open J. 2019; 4(2):
69-76. doi: 10.17140/VMOJ-4-138

7. KHAN, R., & NAZ, S. (2013). The applications of probiotics in poultry production. World's Poultry Science
Journal, 69(3), 621-632. doi:10.1017/S0043933913000627

8. Fuller, R. Probiotics in man and animals. J. Appl. Bacteriol. 1989, 66, 365-378.

9. O'Dea EE, Fasenko GM, Allison GE, Korver DR, Tannock GW, Guan LL. Investigating the effects of commercial
probiotics on broiler chick quality and production efficiency. Poult Sci. 2006 Oct;85(10):1855-63. doi:
10.1093/ps/85.10.1855. PMID: 17012181.

10. Thomke, S.; Elwinger, K. Growth promotants in feeding pigs and poultry. III. Alternatives to antibiotic
growth promotants. Ann. Zootech. 1998, 47, 245-271.

11. Kechagia M, Basoulis D, Konstantopoulou S, et al. Health benefits of probiotics: A review. ISRN Nutrition.
2013; 2013. 1-7. doi: 10.5402/2013/481651

11
12. Meyer, M.M.; Fries-Craft, K.A.; Bobeck, E.A. Composition and inclusion of probiotics in broiler diets alter
intestinal permeability and spleen immune cell profiles without negatively affecting performance1. J. Anim.
Sci.2020, 98, skz383. [CrossRef]

13. Fuller R. The importance of lactobacilli in maintaining normal microbial balance in the crop. Br Poult Sci.
1977;18(1):85–94.

14. Fuller R. Probiotics for farm animals. In: Tannock GW, ed. Probiotics: A Critical Review. Norfolk, England:
Horizon Scientific Press; 1999:15–22.

15. Gusils C, Gonzalez SN, Oliver G. Some probiotic properties of chicken lactobacilli. Can J
Microbiol.1999;45(12):981–987

16. Liu, G.; Griffiths, M. W.; Wu, P.; Wang, H.; Zhang, X. & Li, P. (2011). Enterococcus faecium LM-2, a
multi-bacteriocinogenic strain naturally occurring in “Byaslag”, a traditional cheese of Inner Mongolia in
China. Food Control, Vol.22, No.2, (February 2011), pp. 283–289, ISSN 0956-7135

17. Castilho, N.A.; De Leblanc, A.M.; Galdeano, C.M. & Perdigón, G. (2012). Probiotics: an alternative strategy
for combating salmonellosis immune mechanisms involved. Food Research International, Vol.45, No.2,
(March 2012), pp. 831-841, ISSN 0963-9969

18. Sri-Harimurti, Huda M, Kistiani AD (2013b) The dynamics of indigenous lactic acid bacteria probiotics on
carcass yield, abdominal fat and intestinal morphology of broilers. In: Proceeding of the 3rd AINI
international seminar, Padang, West Sumatera, Indonesia, 24–26 Sept 2013

19. Madsen, Karen. (2012). Enhancement of Epithelial Barrier Function by Probiotics. Journal of Epithelial
Biology and Pharmacology. 5. 10.2174/1875044301205010055.

20. Abd El-Moneim, E. A., El-Wardany, I., Abu-Taleb, A. M., Wakwak, M. M., Ebeid, T. A., & Saleh, A. A.
(2020). Assessment of in ovo administration of Bifidobacterium bifidum and Bifidobacterium longum on
performance, ileal histomorphometry, blood hematological, and
biochemical parameters of broilers. Probiotics and Antimicrobial Proteins, 12(2), 439–450.
https://doi.org/10.1007/s1260 2-019- 09549 -2

21. Singh, K., Kallali, B., Kumar, A., & Thaker, V. (2011). Probiotics: A review. Asian Pacific Journal of
Tropical Biomedicine, 1(2), S287–S290. https:// doi.org/10.1016/S2221 -1691(11)60174 -3

22. Sieo CC, Abdullah N, Tan WS, Hot YW (2005) Influence of glucanase-producing lactobacilli strains on
intestinal characteristics and feed passage rate of broiler chickens. J Poult Sci
84:734–741

12
23. Kizerwetter-Swida, M.; Binek, M. Protective effect of potentially probiotic Lactobacillus strain on infection
with pathogenic bacteria in chickens. Pol. J. Vet. Sci. 2009, 12, 15-20.

24. Edens F (2003) An alternative for antibiotic use in poultry’ probiotics. Revista Brasileira de
Ciência Avícola 5:75–97

25. Ewing WN, Cole DJA (1994) The living gut: an introduction to micro-organisms in nutrition.
Context Publications, Dungannon, UK

26. Willey JM, Van Der Donk WA (2007) Lantibiotics: peptides of diverse structure and function.Annu Rev
Microbiol 61:477–501

27. Zasloff M (2002) Antimicrobial peptides in health and disease. N Engl J Med 347:1199–1200

28. Mulaw G, Sisay Tessema T, Muleta D, Tesfaye A. In vitro evaluation of probiotic properties of lactic acid
bacteria isolated from some traditionally fermented Ethiopian food products. Int J Microbiol. 2019; 2019:
7179514. doi: 10.1155/2019/7179514

29. SCHNEITZ , C. 2005 . Competitive exclusion in poultry – 30 years of research . Food Cont. , 16 , 657 –
667 .

30. Stern, N.J.; Cox, N.A.; Bailey, J.S.; Berrang, M.E.; Musgrove, M.T. Comparison of mucosal competitive
exclusion and competitive exclusion treatment to reduce Salmonella and Campylobacter spp. colonization
in broiler chickens. Poult. Sci. 2001, 80, 156-160.

31. Fritts, C.A.; Kersey, J.H.; Motl, M.A.; Kroger, E.C.; Yan, F.; Si, J.; Jiang, Q.; Campos, M.M.;Waldroup,
A.L.; Waldroup, P.W. Bacillus subtilis C-3102 (Calsporin) improves live performance and microbiological
status of broiler chickens. J. Appl. Poult. Res. 2000, 9,149- 155.

32. Schneitz, C. Competitive exclusion in poultry––30 years of research. Food Control 2005, 16, 657-667.

33. Carita Schneitz, Competitive exclusion in poultry––30 years of research, Food Control, Volume 16, Issue
8, 2005, Pages 657-667, ISSN 0956-7135,

34. Pan, D., Yu, Z., 2014. Intestinal microbiome of poultry and its interaction with host and diet. Gut Microbes
5, 108–119.

35. Slepecky, R.A., Hemphill, H.E., 2006. The genus Bacillus—nonmedical. In: Dworkin, M., Falkow, S.,
Rosenberg, E., Schleifer, K.H., Stackebrandt, E. (Eds.), The Prokaryotes: A Handbook on the Biology of
Bacteria 4. Springer, New York, USA, pp. 530–562.

36. Lalloo, R., Maharajh, D., Görgens, J., Gardiner, N., 2010a. A downstream process for production of a viable
and stable Bacillus cereus aquaculture biological agent. Appl. Microbiol. Biotechnol. 86, 499–508.

13
37. La Ragione, R.M, Woodward, M.J, 2003. Competitive exclusion by Bacillus subtilis spores of Salmonella
enterica serotype enteritidis and Clostridium perfringens in young chickens. Vet. Microbiol. 94, 245–256

38. Bermudez-Brito, M., Plaza-Díaz, J., Muñoz-Quezada, S., Gómez-Llorente, C., Gil, A., 2012. Probiotic
mechanisms of action. Ann. Nutr. Metab. 61, 160–174.

39. Lee, K.-W., Lillehoj, H.S., Jang, S.I., Li, G., Lee, S.-H., Lillehoj, E.P., Siragusa, G.R., 2010a. Effect of
Bacillus-based direct-fed microbials on Eimeria maxima infection in broiler chickens. Comp. Immunol.
Microbiol. Infect. Dis. 33, e105–e110.

40. Chichlowski, M., Croom, J., Mcbride, B., Havenstein, G., Koci, M., 2007. Metabolic and physiological
impact of probiotics or direct-fed-microbials on poultry: a brief revie of current knowledge. Int. J. Poult.
Sci. 6, 694–704.

41. Alkhalf A., Alhaj M. and Al-Homidan I. (2010). Influence of probiotic supplementation on immune
response of broiler chicks. Egyptian Poult. Sci. 30, 271-280.

42. Tsai Y.T., Cheng P.C. and Pan T.M. (2012). The immunomodulatory effects of lactic acid bacteria for
improving immune functions and benefits. Appl. Microbiol. Biotechnol. 96, 853-862.

43. Haghighi H.R., Gong J., Gyles C.L., Hayes M.A., Zhou H., Sanei B., Chambers J.R. and Sharif S. (2006).
Probiotics stimulate production of natural antibodies in chickens. Clin. Vaccine Immunol. 13, 975-980.

44. Ouwehand AC, Kirjavainen PV, Shortt C, Salminen S (1999) Probiotics: mechanisms and established
effects. Int Dairy J 9:43–52

45. Thanissery R, McReynolds JL, Conner DE, Macklin KS, Curtis PA, Fasina YO. Evaluation of the efficacy
of yeast extract in reducing intestinal Clostridium perfringens levels in broiler chickens. Poult Sci. 2010
Nov;89(11):2380-8. doi: 10.3382/ps.2010-00879. PMID: 20952700.

46. Elhasany, Hams. (2016). Effect of Probiotic on Salmonella Enteritidis infection on broiler chickens.
Egypt.J.Chem.Environ.Health,2(2):298-314. 2. 298-314.

47. Eid, Samah & Atfehy, Nayera & Amer, Fatma & Tolba, Hala & Hamed, Rehab. (2020). Prevention of
Necrotic Enteritis in Broiler Chickens by Prebiotics and Probiotics VS Control by Antibiotics, in Vivo
Study. Alexandria Journal of Veterinary Sciences. 64. 143. 10.5455/ajvs.76994.

48. Knap I, Lund B, Kehlet AB, Hofacre C, Mathis G. Bacillus licheniformis prevents necrotic enteritis in
broiler chickens. Avian Dis. 2010 Jun;54(2):931-5. doi: 10.1637/9106-101509-ResNote.1. PMID:
20608542.

14
49. Cox CM, Dalloul RA. Immunomodulatory role of probiotics in poultry and potential in ovo application.
Benef Microbes. 2015 Mar;6(1):45-52. doi: 10.3920/BM2014.0062. PMID: 25213028

50. Ng, S.C.; Hart, A.L.; Kamm, M.A.; Stagg, A.J. & Knight, S.C. (2009). Mechanisms of Action of Probiotics:
Recent Advances. Inflammatory Bowel Diseases, Vol.15, No.2, (February 2009), pp.300-310, ISSN 1536-
4844

51. Zhang, Z.; Hinrichs, D.J.; Lu, H.; Chen, H.; Zhong, W. & Kolls, J.K. After interleukin-12p40, are
interleukin-23 and interleukin-17 the next therapeutic targets for
inflamematorybowel disease? Int Immunopharmacol, 200 Vol.7, No.4, (April, 2007), pp.
409–416, ISSN 1567-5769

52. Gupta, V. & Garg, R. (2009). Probiotics. Indian Journal of Medical Microbiology, Vol.27, No.3, (July-
September 2009), pp. 202-209, ISSN 1998-3646

53. Shang Y, Kumar S, Oakley B and Kim WK (2018) Chicken Gut Microbiota: Importance and Detection
Technology. Front. Vet. Sci. 5:254. doi: 10.3389/fvets.2018.00254

54. Yadav, S.; Jha, R. Strategies to modulate the intestinal microbiota and their e_ects on nutrient utilization,
performance, and health of poultry. J. Anim. Sci. Biotechnol. 2019, 10, 2.

55. Nakphaichit, M.; Thanomwongwattana, S.; Phraephaisarn, C.; Sakamoto, N.; Keawsompong, S.;
Nakayama, J.; Nitisinprasert, S. The e_ect of including Lactobacillus reuteri KUB-AC5 during post-hatch
feeding on the growth and ileum microbiota of broiler chickens. Poult. Sci. 2011, 90, 2753–2765.

56. Martínez, E.A.; Babot, J.D.; Lorenzo-Pisarello, M.J.; Apella, M.C.; Chaia, A.P. Feed supplementation with
avian Propionibacterium acidipropionici contributes to mucosa development in early stages of rearing
broiler chickens. Benef. Microbes 2016, 7, 687–698.

57. Asghar, S.; Arif, M.; Nawaz, M.; Muhammad, K.; Ali, M.A.; Ahmad, M.D.; Iqbal, S.; Anjum, A.A.; Khan,
M.; Nazir, J. Selection, characterisation and evaluation of potential probiotic Lactobacillus spp. isolated
from poultry droppings. Benef. Microbes 2016, 7, 35–44.

58. Abd El-Moneim, E. A., El-Wardany, I., Abu-Taleb, A. M., Wakwak, M. M., Ebeid, T. A., & Saleh, A. A.
(2020). Assessment of in ovo administration of Bifidobacterium bifidum and Bifidobacterium longum on
performance, ileal histomorphometry, blood hematological, and
biochemical parameters of broilers. Probiotics and Antimicrobial Proteins, 12(2), 439–450.
https://doi.org/10.1007/s1260 2-019- 09549 -2

59. Abou-Kassem, D., Elsadek, M., Abdel-Moneim, A., Mahgoub, S., Elaraby, G., Taha, A., Elshafie, M. M.,
Alkhawtani, D. M., Abd El-Hack, M. E., & Ashour, E. (2020). Growth, carcass characteristics, meat quality

15
 and microbial aspects of growing quail fed diets enriched with two different types of probiotics (Bacillus
toyonensis and Bifidobacterium bifidum). Poultry Science https://doi.org/10.1016/j.psj.2020.04.019in press.

60. Mulaw G, Sisay Tessema T, Muleta D, Tesfaye A. In vitro evaluation of probiotic properties of lactic acid
bacteria isolated from some traditionally fermented Ethiopian food products. Int J Microbiol. 2019; 2019:
7179514. doi: 10.1155/2019/7179514

61. Dittoe DK, Ricke SC, Kiess AS. Organic acids and potential for modifying the avian gastrointestinal tract
and reducing pathogens and disease. Front Vet Sci. 2018; 5: 216. doi: 10.3389/ fvets.2018.00216

62. Bermudez-Brito M, Plaza-Díaz J, Muñoz-Quezada S, Gomez- Llorente C, Gil A. Probiotics mechanisms of

action of. Ann Nutr Metab. 2012; 61: 160-174. doi: 10.1159/000342079

63. Lee SH, Lillehoj HS, Dalloul RA, Park DW, Hong YH, Lin JJ. Influence of Pediococcus-based probiotic
on coccidiosis in broiler chickens. Poult Sci. 2007 Jan;86(1):63-6. doi: 10.1093/ps/86.1.63. PMID:
17179417

64. Kyung-Woo Lee, Hyun S. Lillehoj, Seung I. Jang, Guangxing Li, Sung-Hyen Lee, Erik P. Lillehoj, Gregory
R. Siragusa, Effect of Bacillus-based direct-fed microbials on Eimeria maxima infection in broiler chickens,
Comparative Immunology, Microbiology and Infectious Diseases,Volume 33, Issue 6, 2010, Pages e105-
e110,ISSN 0147-9571, https://doi.org/10.1016/j.cimid.2010.06.001.

65. Giannenas I, Tsalie E, Triantafillou E, Hessenberger S, Teichmann K, Mohnl M, Tontis D. Assessment of

probiotics supplementation via feed or water on the growth performance, intestinal morphology and
microflora of chickens after experimental infection with Eimeria acervulina, Eimeria maxima and Eimeria
tenella. Avian Pathol. 2014;43(3):209-16. doi: 10.1080/03079457.2014.899430. Epub 2014 Apr 4. PMID:
24601749.

66. Wang, Y., Cho, J., Chen, Y., Yoo, J., Huang, Y., Kim, H., Kim, I., 2009. The effect of probiotic BioPlus 2B
Ⓡ on growth performance, dry matter and nitrogen digestibility and slurry noxious gas emission in growing
pigs. Livest. Sci. 120, 35–42.

67. Ahmed, S.T., Islam, M.M., Mun, H.-S., Sim, H.-J., Kim, Y.-J., Yang, C.-J., 2014. Effects of Bacillus
amyloliquefaciens as a probiotic strain on growth performance, cecal microflora, and fecal noxious gas
emissions of broiler chickens. Poult. Sci. 93, 1963–1971.

68. Pezzuolo, Andrea & Sartori, Cristina & Vigato, Evelin & Guercini, Stefano. (2019). Effect of litter treatment
with probiotic bacteria on ammonia reduction in commercial broiler farm. 10.22616/ERDev2019.18.N390.

16
69. IU Sheikh, SS Nissa, Bushra Zaffer, KH Bulbul, AH Akand, HA Ahmed, Dilruba Hasin, Isfaqul Hussain,
SA Hussain. Ammonia production in the poultry houses and its harmful effects. Int J Vet Sci Anim
Husbandry 2018;3(4):30-33.

70. HOSSEIN, Ahmed & Gurbuz, Yavuz. (2016). Aflatoxins in Poultry Nutrition. Kahramanmaraş Sütçü İmam
Üniversitesi Doğa Bilimleri Dergisi. 18. 1. 10.18016/ksujns.98227.

71. Fan, Y., Zhao, L., Ji, C., Li, X., Jia, R., Xi, L., Zhang, J., Ma, Q., 2015. Protective effects of
Bacillus subtilis ANSB060 on serum biochemistry, histopathological changes and antioxidant enzyme
activities of broilers fed moldy peanut meal naturally contaminated with aflatoxins. Toxins 7, 3330–3343.

72. Bagherzadeh Kasmani F, Karimi Torshizi MA, Allameh A, Shariatmadari F. A novel aflatoxin-binding
Bacillus probiotic: Performance, serum biochemistry, and immunological parameters in Japanese quail.
Poult Sci. 2012 Aug;91(8):1846-53. doi: 10.3382/ps.2011-01830. PMID: 22802177.

73. Sakthivelan, Sigamani & Rao, Ganne. (2010). Effect of Ochratoxin A on Body Weight, Feed Intake and
Feed Conversion in Broiler Chicken. Veterinary medicine international. 2010. 590432.
10.4061/2010/590432.

74. Śliżewska K, Piotrowska M. Reduction of ochratoxin A in chicken feed using probiotic. Ann Agric Environ
Med. 2014;21(4):676-80. doi: 10.5604/12321966.1129913. PMID: 25528900.

75. Piotrowska M, Zakowska Z. The elimination of ochratoxin A by lactic acid bacteria strains. Pol J Microbiol.
2005;54(4):279-86. PMID: 16599298.

76. Piotrowska M. The adsorption of ochratoxin a by lactobacillus species. Toxins (Basel). 2014 Sep
22;6(9):2826-39. doi: 10.3390/toxins6092826. PMID: 25247265; PMCID: PMC4179162.

77. Sugiharto, Sugiharto & Yudiarti, Turrini & Isroli, Isroli & Widiastuti, Endang & Kusumanti, Endang.
(2016). Dietary supplementation of probiotics in poultry exposed to heat stress - A review. Annals of Animal
Science. 17. 10.1515/aoas-2016-0062.

78. Abdur-Rahman Al-Fataftah, Anas Abdelqader, Effects of dietary Bacillus subtilis on heat-stressed broilers
performance, intestinal morphology and microflora composition, Animal Feed Science and Technology,
Volume 198, 2014, Pages 279-285, ISSN 0377-8401 https://doi.org/10.1016/j.anifeedsci.2014.10.012.

79. M.U. Sohail, A. Ijaz, M.S. Yousaf, K. Ashraf, H. Zaneb, M. Aleem, H. Rehman, Alleviation of cyclic heat
stress in broilers by dietary supplementation of mannan-oligosaccharide and Lactobacillus-based probiotic:
Dynamics of cortisol, thyroid hormones, cholesterol, C-reactive protein, and humoral immunity, Poultry
Science, Volume 89, Issue 9, 2010, Pages 1934-1938, ISSN 0032-5791, https://doi.org/10.3382/ps.2010-
00751.

17
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