Table 8.

In ovo alternatives to control Salmonella infection in poultry.

Target Species	Delivered Compound	Experimental Procedure	Main Results	Ref.
SPF Ross 308 broilers	Vaccine	Vaccination Salmonella flagellin to 18 day old embryonated eggs	Elevated pro-inflammatory chIL-6 and chIL-8 cytokine transcript levels 24 h post-vaccination. High titers of FliC-specific antibodies 21 day post-hatch	[173]
Cobb 500 embryonated eggs	Probiotics	Inoculation with a 3 × 1011 CFU/mL suspension of Lactobacillus acidophilus, L. fermentum, and L. salivarius in the air cell of 18 d embryonated eggs. S. Enteritidis inoculation 2 day after hatching	No decrease (p > 0.05) in S. Enteritidis colonization of chick ceca.	[174]
Coob 500 broiler fertil eggs	Prebiotics	In ovo injection of Raffinose (1.5, 3.0, and 4.5 mg in 0.2 mL of aqueous diluents) into the air sac of 12 day embryonated eggs	Increase of the villus height, the villus height–crypt depth ratio (p < 0.05), and the expression levels of CD3 and chB6	[175]
Ross 308 hatching eggs	Prebiotics and synbiotics	Administration, in the air chamber at 12 day of incubation, of inulin, Bi2tos, inulin, and Lactococcus lactis subs. lactis or Bi2tos and Lactococcus lactis subs. lactis	Modulation of central and peripheral lymphatic organ development (cortex/medulla ratio in the thymus, development of cortex in bursal follicles, and germinal center’s formation in the spleens), especially through the use of symbiotics	[176]
Broilers’ embryonated eggs	Prebiotic	Commercial egg injector system (InovojectTM) to apply a dextrin solution (18% maltodextrin, 10% potato extract dextrin) containing iodinated casein (80, 240, 720, or 2160 µg/)mL	Improvement in hatchability and early growth attributable to iodinated casein in combination with Dextrin. No differences in Salmonella colonization after chicks were challenged	[177]
Ross 308 Salmonella free hatching eggs	Probiotic	In ovo injection, into the air cell, at 18 day incubation with 0.1 mL of a commercial probiotic suspension (7 × 107 CFU/mL in PBS). After hatching, chicks were challenged against S. Enteritidis (Se) (8 log CFU)	Reduction in the number of Se colonized chicks since day 1 post hatching. Reduction of Se colonization in the alimentary tract of chicks	[178]
Fertile broiler eggs	Probiotic	Probiotic administration, at 18 day of incubation, of Marek’s vaccine + one of three Bacillus subtilis strains (ATCC 6051, ATCC 8473, ATCC 9466)	Results regarding hatching were strain-dependent; however, probiotic strains reduced the bacterial counts (total aerobes and coliforms) in the ileum and ceca	[172]
White Leghorn hens	Immune lymphokines	On day 18 of embryogenesis, eggs were injected into the amnion with Immune (ILK) and nonimmune (NILK) Lymphokines. Post-hatch, chicks were orally challenged against S. Enteritidis (5 × 104 CFU)	In vitro bactericidal activity was higher, and organ invasion with S. Enteritidis decreased in ILK-treated chicks. Hatchability was not affected, although ILK-treated chicks were 1 g lighter than NILK-treated ones (p < 0.05)	[179]
Broiler embryonated eggs	Probiotic	Injection, at 18 day of incubation, of an undefined and anaerobically grown competitive exclusion culture into the air cells or beneath the inner air cell membrane	Evident resistance to S. Typhimorium of chicks challenged at day 7 post hatch	[180]
Broiler embryonated eggs	Probiotic	A competitive exclusion culture consisting of several species of unrevealed bacteria injected either the air cell or body of the 18 day of incubation embryos	Injection in the body proper resulted in losing almost all the hatchability. Hatchability was reduced and mortality during the first week increased in air cell injected embryos. No effects on Salmonella infection were observed when chicks were challenged 1 day after hatching	[181]
Ross embryonated eggs	Probiotic	18 d incubating eggs were inoculated using cecal microbiota (total or diluted) and Lactobacillus salivarius into the inner air sac	Maximum hatchability observed was 65%. 2-d chicks were challenged against S. Enteritidis. Liver and cecum colonization was not reduced in the in ovo inoculated chicks	[182]
18-d White Leghorn 15I5 × 71 embryonated eggs	Probiotic	Eggs were inoculated after 18 d of incubation with a commercial probiotic (FloraMax®-B11) through injection into the amnion. After hatching, chickens were orally infected with S. Enteritidis	Probiotic administration did not affect hatchability but increased body weight during first 7 d, increased the villi surface area in the ileum and reduced the presence of lactose-positive Gram-negative bacteria, as well as reduced the incidence of S. Enteritidis	[183]
Broiler embryonated eggs	Antibiotic	Gentamicin was administered at 18 d of incubation into the amnion. At hatching, chicks were gavaged with a commercial Competitive Exclusion Culture (MSC®. 0.2ml, 1 × 108 UFC/)mL and challenged against S. Typhimurium	A cumulative effect was observed by the in ovo application of Gentamicin and the supplementation with the Competitive Exclusion Culture at hatching in ceca colonization with S. Typhimurium	[184]
Ross × Ross 708 fertile eggs	Probiotic	18 d of incubation eggs were inoculated with an Enterococcus faecium-based commercial probiotic (Galli-Pro Hatch) at three concentrations	Hatchability was not affected, and live performance in the first 21 days were improved as well as yolk absorption and intestinal and spleen morphology	[185]
Ross 308 broiler embryonated eggs	Probiotic	Injection was performed at 17.5 d of incubation for the inoculation of two probiotic strains (Enterococcus faecium and Bacillus subtilis). Chicks were orally challenged against S. Enteritidis 4 days post hatching	Probiotic administration, at a dose up to 109 CFU/egg) not only reduces but also eliminates the presence of Salmonella in broilers	[186]
Cobb 500 fertile eggs	Probiotic	On day 18 embryonic day, eggs were injected into the air cell with a commercial probiotic (Primalac W/S. Lactobacillus acidophilus, Lactobacillus casei, Enterococcus faecium, and Bifidobacterium bifidum) using three different concentrations	Hatchability, feed intake, and feed conversion ratio were not affected by the probiotic administration. The expression of immune-related genes in the ileum and cecal tonsils were modulated	[187]
Broiler fertilized eggs from a commercial breeder	Immune response stimulation	18 d old embryos were injected in the amnion with CpG oliodeoxynucleotides (CpG-ODN) and orally infected with S. Enteritidis at day 10 post-hatch	Colonization of S. Enteritidis in ceca was reduced greater than 10-fold in comparison to placebo birds. CpG-ODN stimulates innate immune responsiveness of birds heterophils	[188]