The effect of blue and red LED light on the growth, egg production, egg quality, behavior, and hormone concentration of Hy-Line® W-36 laying hens


A study was conducted to investigate the effect of blue and red light-emitting diode (LED) on performance, behavior, egg quality, and hormonal concentration of pullets and laying hens. A total of 1,000 Hy-Line® W-36 birds were raised in a cage-free housing system consisting of two identical rooms, one equipped with blue-LED from 1 to 18 wk of age and red-LED from 19 to 31 wk of age, and the other with normal-LED bulbs throughout the study. The study was repeated the following year. Body weight was recorded monthly, and egg production was recorded daily. Egg quality, organ weight, tonic immobility (TI), heterophil-lymphocyte ratio, serum corticosterone, luteinizing hormone and melatonin were analyzed monthly after the egg production started. The results showed that birds raised in blue-LED had a significantly higher body weight during the pullet phase from 1 to 18 wk of age (P = 0.049). A higher relative egg yolk percentage was observed in the red-LED (P = 0.043) with a lower relative albumen percentage (P = 0.015) compared to the normal-LED light. Lower relative spleen weight was observed in the red-LED compared to the normal-LED (P = 0.027). There was no difference between the light treatments with respect to hen day egg production, brain weight, tonic immobility and hormone concentration. The effects of these lights on the overall production, health, and immune system are not completely understood. Hence, it is difficult to conclude that exposing birds to blue-LED in the pullet phase for improved growth and red-LED in the laying phase for earlier and greater egg production would benefit overall egg production.


Laying hen


Light is necessary for the growth, development, and production performance of the laying hen. In the early 1930s, the main focus of research in poultry lighting was photoperiod and light intensity (Lewis and Morris, 2000). With the discovery of new efficient lighting technologies, the wavelength of light became an interest to many researchers. Light-emitting-diode (LED) has improved light delivery technology by providing better energy efficiency, a longer lifespan of bulbs, and the ability to select a spectrum of light. It has been shown that the wavelength of light can affect the behavior and performance of layers (Manser, 1996). Thus, LED has been extensively researched to enhance the performance of laying hens (Archer, 2019).

In the past, experiments have exposed poultry to various kinds of light sources and studied their effects on growth and production. Different wavelengths of light also play a role in regulating the physiology of the bird. Wavelength has the potential to promote growth and egg production and reduce stress, increase fertility, and improve the quality of eggs (Li et al., 2014). Longer wavelengths in red light can penetrate the skull and stimulate hypothalamic photoreceptors and reproduction in birds (Lewis and Morris, 2000). Laying hens kept under red light not only started to lay eggs earlier but also had higher egg production than birds kept in white or green light (Huber-Eicher et al., 2013Baxter et al., 2014). Exposing birds to the red light resulted in significantly higher egg production in the first cycle (Pyrzak et al., 1987) as well as in the second cycle (Pyrzak et al., 1987Reddy et al., 2012). In addition, the wavelength has been shown to affect the behavior of the hen (Sultana et al., 2013Archer, 2019Archer (2019). found that caged White Leghorns, exposed to red-LED from 42 to 72 wk of age, had lower plasma corticosterone levels, lower heterophil to lymphocyte ratio and lower composite asymmetry scores compared to white-LED, indicating that red-LED can lower stress in commercial laying hens.

On the other hand, shorter wavelengths like blue light can promote growth and minimize locomotory activity in birds (Prayitno and Phillips, 1997Sultana et al., 2013Hassan et al., 2014). Thus, several previous studies have shown that blue light can promote growth, improve immunity and reduce mortality, while red light can enhance egg production. While several studies have investigated the activity of various colors of LED lights on body weight, reproductive performance and behavior of laying hens, those studies have been conducted using only individual lights, meaning either only blue or only red in their lifetime. Research with the combination of blue-LED during the pullet phase and then switching to red-LED in the laying phase has not been carried out. Therefore, to fulfill the gap, this study has investigated the possibility of utilizing a blue spectrum of light during the pullet phase and the red spectrum in the production phase enhance the growth and production of Hy-Line® W-36 laying hens. We hypothesized that hens reared under experimental light (first blue-LED and then red-LED) would increase body weight gain, egg production and lower stress and fear responses as compared to a normal-LED light used in poultry farms. Therefore, the major objectives of this study were 1) to determine the effect of blue-LED light in the pullet phase and red-LED light in the layer phase on growth, reproductive hormonal concentration, production performance (hen-day egg production), and egg quality of Hy-Line® W-36 laying hens compared with normal-LED and 2) to determine the effect of the blue/red LED lighting regimen on behavior and stress responses of Hy-Line® W-36 laying hens.


The study was conducted at Mississippi State University’s Poultry Research Unit, Mississippi State, Mississippi, and all procedures complied with the Institutional Animal Care and Use Committee authorization number, IACUC-18-372.

Bird Husbandry and Room Configuration

Hy-Line® W-36 (N=1000) were obtained as day-old chicks from Hy-Line hatchery (Hy-Line North America, LLC, GA) and wing-banded on placement. The study was conducted in two identical rooms measuring 8.83 m × 4.26 m (length × width). The study was repeated twice. Feed and water were provided ad libitum and feed was formulated according to Hy-Line® Management Guide (2016), as shown in Table 1. Forty nest boxes with dimensions 30.48 cm × 30.48 × cm 33.03 cm (length × width × height) were placed in the center of each room. Scratchpads were placed in all nest boxes. Eight wooden perches of were placed in each room, four on the sides of the wall and four on the nest boxes. Each room had a perching area of 8.2 cm per bird. The stocking density at the beginning of the trial was 0.15 birds per meter square. Care was taken to prevent outside light from entering the room, using black plastic curtains on the open side to minimize external light. A ventilation fan was used to circulate the air. Air temperature and humidity were measured daily to ensure that optimum conditions were provided. To ensure that external factors were similar for the replicates, the experiment was started at the same time of the year. Tissue sampling and other variables were also measured a year later for the replicate during the same time of the year.

Table 1. Composition and nutritional values of Hy-Line® W-36 grower diet.

Ingredient Amount (%)
Grower diet Layer 1 diet
Corn 59.32 61.20
Soybean meal 31.34 20.49
Limestone 1.34 10.40
Distiller dried grain with soluble 3.00 3.00
Poultry fat 2.50 2.50
Dicalcium phosphate 1.70 1.72
NaCl 0.30 0.30
Vitamin mineral premix1 0.26 0.26
DL-Methionine 0.09 0.12
L-Lysine 0.05 0.02
Calculated values
ME (Kcal/kg) 2,980.00 2,850.00
Total protein (%) 17.50 16.15
Total Calcium2 (%) 1.00 4.48
Available P (%) 0.47 0.49
dLys (%) 0.90 0.86
dMet (%) 0.42 0.41
dCys (%) 0.25 0.30

Provided as: Vitamin A, 1,400,000 IU/lb.; Vitamin D3, 500,000 ICU/lb.; Vitamin E, 3,000IU/lb.; Vitamin B1, 2mg/lb.; menadione, 150mg/lb.; riboflavin, 1,200mg/lb., D-pantothenic acid, 1,200mg/lb.; niacin, 5,000mg/lb.; choline, 70,000 mg/lb.; folic acid, 125mg/lb.; pyridoxine, 250 mg/lb.; thiamine, 200 mg/lb.; biotin, 6 mg/lb., Manganese, 4%; zinc, 4%; Iron, 2%; copper, 4,500ppm; iodine, 600ppm; selenium, 60ppm.


The ratio of fine and coarse calcium ratio was changed according to the Hy-Line® management guidelines (2016).

Lighting Treatment

One of the rooms was fitted with four normal-LED bulbs (Overdrive®, 6 W, 3000K, 120 V, 60Hz, 55mA, 520 lumens) and was given the light schedule as per the Hy-Line® Management Guide. The other room was fitted with four proprietary experimental bulbs which supplied blue light from 1-18 wk of age and red light from 19-31 wk of age according to the manufacturer’s proprietary guidelines for 24-h photoperiod. The light intensity in each room was measured with a LM-200 LED Light MeterTM (Amprobe®). The light intensity during bird placement at bird’s eye level was 2.0 and 30 lux in the experimental LED and normal-LED rooms, respectively; attenuation was not measured, and intensity was not converted to clux. As intensity per se was not an experimental treatment, the relative difference in intensity between the two rooms was reported. The lower intensity of experimental LED was used as per the manufacture’s proprietary guidelines. To rule out room effect, the experiment was repeated in the second year, with all other experimental parameters remaining the same.

Blood Sampling Procedures

Blood was collected by brachial venipuncture using a 3-mL disposable Luer-LokTM Tip syringe (BD, Franklin Lakes, NJ) with a 22G disposable hypodermic needle (Exel International, Quebec, CA) from 6 birds randomly selected from each room at intervals of four wk from 18 to 30 wk of age. All blood samples for the trial were collected from 8:00 AM to 12:00 Noon. To minimize stress to the birds during the blood collection. All birds were handled similarly and quickly by two persons, one person holding the bird and another person carrying out the venipuncture. One drop of blood was immediately placed on each of two clean glass slides per bird for subsequent H:L ratio determinations (described below). The remaining blood was then transferred to a 5-ml vacutainer (BD, Franklin Lakes, NJ) and was kept for 30 min at room temperature to coagulate. The vacutainers were then centrifuged at 2000 × g at 4°C for 10 mins to obtain serum placed in a sterile cryovial and stored at -20°C until analyzed.


Body weights of 20 random birds per room were recorded every 2wk during the pullet phase (1 to 18 wk). Eggs were counted and collected daily at 8:30 AM and 3:30 PM. All the eggs were collected within a half-hour window. Mortality records were maintained throughout the experimental period.

Egg Quality

Egg quality and eggshell breaking strength (EBS) were measured at 22, 26 and 30 wk of age. For each wk, 30 eggs per room were collected for egg quality measurements and the following day, 30 additional eggs per room were collected for EBS. Egg quality measurements included egg weight (EW), specific gravity (SG), eggshell thickness (EST), shell weight (SW), percent shell weight (PSW), albumen height (AH), Haugh units (HU), yolk weight (YW), and yolk albumen ratio (YAR). For SG, eggs were submerged in ascending order of saline solutions with a predetermined specific gravity (Peebles and McDaniel, 2013). Haugh units were calculated by breaking a fresh egg on a level flat surface, calculating the albumen height using the TSS QCD apparatus (Technical Services and Supplies Ltd, York, England) and corresponding it with egg weight. Immediately after breaking the eggs for internal egg quality, the eggshells were collected and washed with clean water to remove all the adhering albumen. The cleaned eggshells were dried at room temperature for two consecutive days, and eggshell weight and eggshell thickness were recorded. The shell weight and shell thickness were measured with intact shell membranes. To calculate the percent shell weight, SW was divided by EW and multiplied by 100. Eggshell thickness was measured at three different sites on the eggshell equator using an Ames micrometer™ (B.C. Ames Incorporated, MA), after which the average reading of the three measurements was recorded.

To evaluate EBS, 30 eggs from each room were weighed and EBS was measured using an Instron Universal Testing Machine model 3345 (Instron Inc., Norwood, MA) with a crosshead speed of 20 mm/min using a 100 N load cell and 35 mm compression device as a probe (Clerici et al., 2006). The breaking force was analyzed in kilogram-force (KgF) using Bluehill software (Instron Inc., Norwood, MA) and then converted to Newtons (Thompson and Taylor, 2008).

Hormonal Analysis

Serum hormone concentrations of corticosterone, melatonin and luteinizing hormone (LH) were determined using an Enzyme-Linked Immunosorbent Assay (ELISA) assay for samples collected at 18, 22, 26, and 30 wk of age according to the manufacturer’s protocol and validated for avian serum (Chicken Cortisone: MBS7264609; Chicken Luteinizing Hormone: MBS008505; Chicken Melatonin: MBS262913; My BioSource, San Diego, CA). Duplicate 100 μl volumes of standard and experimental samples were loaded into specific ELISA plate wells pre-coated with capture antibody and were then incubated at 37°C for 90 min. Plates were washed 2 times with wash solution (50 mM Tris-buffered saline, 0.14 M NaCl, 0.05% Tween 20, pH 8.0); 100 μl of biotinylated chicken hormone antibody were added to each well and plates were incubated at 37°C for 60 min. Plates were washed 3 times with wash solution and 100 μl of enzyme-conjugate liquid were added to each well to initiate a Tetramethylbenzidine substrate color reaction. The reactant was thoroughly washed off using Phosphate-buffered saline or Tris-buffered saline. Plates were incubated at 37°C for 30 min and washed 5 times with wash solution. A 100 μl volume of color reagent was added to each well to produce a blue color in each sample and samples were then incubated in a dark incubator at 37°C for 30 min. Finally, the color of each sample changed from blue to yellow under the action of a 100 μl volume of stop solution. Optical density (OD) at 450 nm (OD450) for melatonin, corticosterone, and LH was measured with a SpectraMax M5 Microplate Reader (Molecular Devices, San Jose, CA).

Tonic Immobility Measurement

Tonic immobility (TI) was measured at 18, 22, 26, and 30 wk of age. Six birds at random were utilized from each room at each wk to examine their TI response. To measure the TI, a wooden cradle measuring 32 cm length × 21 cm width × 27 cm height with a black cloth to cover the sides was used (Mahboub et al., 2004). Tonic immobility was recorded on the same day for both rooms from 14:00 to 16:00 hours each time. To induce TI, birds were placed inside the cradle and a hand was placed on the sternum to calm the birds for 15 seconds. A stopwatch was used to measure the time it took for the bird to recover from dorsal recumbency position to standing on its feet again (Archer, 2019). The minimum and maximum acceptable time scores were set at 15 and 480 seconds, respectively, to be recorded as a valid run. In case the bird stood on its feet before 10 seconds, the bird was restrained, and the procedure was repeated. On the other hand, if the bird did not stand even after 480 seconds, that bird was assigned the maximum score (Campbell et al., 2019).

Heterophil / Lymphocyte Ratio Measurement

Blood smears were made from blood samples collected from 6 randomly selected birds from each room at 18, 22, 26 and 30 wk of age immediately after blood collection. One drop of blood was placed on each of two clear Superfrost® microscope slides (Fisher Scientific, Waltham, MA) per bird. The smear was made with the canted end of another slide. The blood smears were dried for 5 min at room temperature and stained with Wright-Giemsa Stain (Fisherbrand®, Waltham, MA). Heterophils and lymphocytes were counted under a Laxco™ SeBa™ Digital microscope (Laxco Inc, Mill Creek, WA) at 40 × magnification until a total of 100 cells per slide was achieved. The H/L ratio was obtained by dividing the number of heterophils by the number of lymphocytes on each slide and averaged across duplicate slides per bird.

Organ and Tissue Sampling

A total of 96 birds (six per room × two rooms × eight-time points) were randomly selected and euthanized using carbon dioxide asphyxiation at 18, 22, 26, and 30 wk of age. The entire brain, including the hypothalamus and pituitary, was excised by cutting through the supraoccipital bone and exposing the cranial cavity to access the brain. The brain was weighed after it was completely removed from the cranial cavity. Similarly, the spleen was also removed and weighed.

Statistical Analysis

Data were tested for normality with PROC UNIVARIATE procedure of SAS 9.4 (SAS Inc., Cary, NC). Log transformation was performed on tonic immobility score and hormonal concentration to normalize the data. The data was analyzed using PROC GLM procedure of SAS 9.4 (SAS Inc., Cary, NC). The data were analyzed as a randomized complete block with split-plot in time design, where year was a blocking factor, room was considered a main plot factor, and wk was considered a split plot factor. Body weight, relative organ weight, internal egg quality, eggshell quality, hormonal concentration, H/L ratio, and tonic immobility were analyzed using the following statistical model:Yijkl=μ+αi+δk+εik+τj+(ατ)ij+eijk+θijkl

Here, Yijkl denotes the independent observation for light type i in two year j at k wk of age during sampling. Each room was considered as an experimental unit. Where μ is the overall mean; αi is the effect of light, δk is the effect of year and δk∼N(0,σδ2)εik is the error term between light type and year such that εik∼N(0,σε2); τjis the age effect. Similarly, (ατ)ij is the interaction effect of light and age; eijk is the sample error with eijk∼N(0,σ2)θijklis the sampling error within groups, such that θijkl∼N(0,σ2). To compare the HDEP, the same model was used except for the sampling error term since the entire room (250 birds) was used to calculate HDEP. Light treatment was selected as a main plot factor, wk as a split plot factor, and year as a random factor. A P-value ≤ 0.05 was considered significant. Fisher’s protected LSD was used to separate the means amongst the treatments.


Body Weight and Egg Production

During the pullet phase (1-18 wk) there was a main effect of light (P = 0.049) on body weight (805 g under blue-LED; 761 g under normal-LED) (Table 2 and Figure 1). In a previous study, broiler raised under blue and green light showed higher body weight gain than white and red light (Wabeck and Skoglund, 1974). The reason for higher body weight gain may be the reduced degree of locomotion. Broilers raised under single monochromatic light and a combination of lights (blue and green) have been reported to spend more time standing and resting with a lesser degree of locomotion compared to birds that were exposed to the longer wavelength of light (red) (Sultana et al., 2013). Similar results were observed by Prayitno et al. (1997), where birds exposed to blue and green light were more sedentary than compared to red light. The lesser extent of locomotive behavior might be a factor that contributes to higher weight gain (Sultana et al., 2013). In one study, Hy-Line® W-36 hens were exposed to blue-LED, red-LED and CFL bulbs from 5 to 14 wk and locomotion was recorded every wk using a computerized program to track the movement of the bird throughout the pen for one entire day (Liu, 2018b). The locomotory behavior was quantified as a movement index, defined as the cumulative displacement in the area caused by the locomotion and calculated in 1-second intervals. The results showed that birds exposed to blue-LED had a higher movement index than CFL or red-LED (Lui, 2018b). Birds exposed to blue-LED came into lay at a mean age of 17.35 wk compared to 19.07 wk for the normal-LED (P = 0.117).

Table 2. Effect of LED type, age and their interaction on body weight (g) recorded from 1 to 18 wk, 18 to 30 wk and overall from 1 to 30 wk of age

Light Age (wk) 1 to 18 wk 18 to 30 wk 1 to 30 wk
Blue-Red-LED 805a 1463 1052
Normal-LED 761b 1448 1021
SEM 9.7 15.8 9.1
1 37e 37g
7 541d 541f
10 779c 779e
12 988b 988d
14 1150a 1150c
18 1202a 1202c 1202c
22 1476b 1476b
26 1548ab 1548a
30 1605a 1605a
SEM 16.9 27.3 19.4
Source of Variation P-value
Year 0.0929 0.9323 0.7097
Light 0.0494 0.8287 0.4333
Age <0.001 <0.001 <0.001
Light × Age 0.6378 0.3287 0.2809

a-fvalues within columns with different superscripts are significantly different at p ≤ 0.05

Figure 1

Figure 1. Effect of LED type on body weight of pullets from 1 to 18 wk of age. The P-value indicates that there was a difference in the weight of pullets at that age. * Statistically significant difference between the treatment

Photoperiod and color of light when combined, are another factor that can influence the growth of birds. During our experiment, we observed that birds exposed to blue-LED had significantly higher body weight than normal-LED birds even when 24 hours of photoperiod was maintained. The increase in body weight of hens that lay earlier than other hens has been attributed specifically to the increase in ovarian and oviduct weight of these birds (Dunnington and Siegel, 1984). Although we did not measure the reproductive organ weight in our experiment, previous studies have reported that birds exposed to photostimulation have higher ovary weight at first lay (Robinson et al., 1996). In another experiment, White Leghorns from 72 to 82 wk of age exposed to a red-LED with 16L:8D had higher body weight than birds exposed to incandescent lights (Reddy et al., 2012). It was observed that red-light stimulation of the reproductive axis had a significant impact on ovarian shape and weight (Reddy et al., 2012). However, in our experiment, during the laying phase (19 to 31 wk of age), we found comparable body weight between red-LED at 24L:0D as and normal-LED as per Hy-Line® guideline. In broilers, higher growth has been observed from 3 to 20 days of age that were exposed to green and blue lights with 23L:1D photoperiod as compared to red and normal light with 16L:8D photoperiod (Rozenboim et al., 1999). Similarly, Rozenboim et al., (2004) found that birds reared in green and blue light with 23L:1D had higher body weight gain as compared to red or natural light with 23L:1D. This accelerated growth was attributed to the higher proliferation of muscle satellite cells in birds exposed to green and blue light (Halevy et al., 1998). There was no difference in HDEP between the red-LED (64.04) and normal-LED (65.71). (P = 0.775; Table 3 and Figure 2).

Table 3. Effect of red-LED, normal-LED and hen age on hen day egg production (HDEP) from 17-31 wk of age

Light Age (wk) HDEP
Red-LED 64.04
Normal-LED 65.71
SEM 1.952
18 0.182f
19 2.922f
20 11.82f
21 35.94e
22 62.82d
23 76.59cd
24 77.67bcd
25 85.88abc
26 92.56ab
27 93.01a
28 83.28abc
29 95.62a
30 95.24a
31 94.77a
SEM 5.1663
Source of Variation P-value
Year 0.4550
Light 0.7751
Age <0.0001
Light × Age 0.0618

HDEP= hen day egg production

a-gvalues within columns with different superscripts are significantly different at p ≤ 0.05

Figure 2

Figure 2. Effect of LED type on hen day egg production (HDEP) from 17 to 31 wk of age.

Egg Quality

Various egg quality parameters are shown in Table 4. Percent albumen was lower (P = 0.015) under red-LED (67.05%) than under normal-LED (68.25%). Conversely, the yolk percentage was higher (P = 0.043) under red-LED (23.37%) as compared to normal-LED (22.49%) Pyrzak et al., (1987). observed a significant increase in the relative yolk weight in birds exposed to the red light and a significant decrease in the relative albumen percentage. In a previous experiment, exposing the birds to red light can increase the size of yellow yolk follicles (Reddy et al., 2012), which might be the reason for an increase in the relative yolk weight. A longer wavelength of light can stimulate the hypothalamus and the reproductive organs of the laying hen (Lewis and Morris, 2000Reddy et al., 2012). The stimulation of the hypothalamic-pituitary-gonadal (HPG) axis results in the surge of GnRH, which stimulates the release of follicle-stimulating hormone and luteinizing hormone, thereby increasing egg production, as well as egg size, when compared to those reared under white and green (Pyrzak et al., 1987Baxter et al., 2014).

Table 4. Effect of LED type, age, and their interactions with internal and external egg quality from 22 to 30 wk of age

Red-LED 56.40 1.093 67.05b 23.37a 0.34 a 106.77 0.38 5.38 9.57 37.04
Normal-LED 56.87 1.091 68.25a 22.49b 0.33 b 106.36 0.39 5.26 9.25 37.16
SEM 0.733 0.0004 0.167 0.141 0.002 0.412 0.002 0.081 0.080 0.454
22 53.95b 1.089 68.95 21.59 0.31 107.32 0.38 5.08b 9.60 38.06
26 57.68a 1.095 67.45 22.93 0.34 107.93 0.40 5.53a 9.45 37.36
30 58.30a 1.092 66.55 24.27 0.36 104.46 0.38 5.35a 9.17 36.87
SEM 0.593 0.002 0.789 0.685 0.013 2.081 0.010 0.067 0.132 0.551
Red-LED 22 52.28 1.091 68.27 21.98 0.32 108.92 0.38 5.07 9.74 37.67
Red-LED 26 58.07 1.096 67.28 23.17 0.34 108.72 0.40 5.52 9.54 37.24
Red-LED 30 58.88 1.091 65.61 24.95 0.38 102.69 0.36 5.54 9.43 35.54
Normal-LED 22 55.62 1.086 69.63 23.60 0.30 105.72 0.38 5.09 9.16 38.45
Normal-LED 26 57.28 1.094 67.63 22.68 0.33 107.15 0.40 5.54 9.67 37.49
Normal-LED 30 57.72 1.093 67.48 21.20 0.35 106.23 0.39 5.16 8.92 35.54
SEM 0.835 0.0038 1.117 0.969 0.019 2.943 0.01 0.140 0.174 0.785
Source of Variation P-value
Year 07823 0.0844 0.0448 0.7783 0.0294 0.2252 0.1143 0.5528 0.1256 0.7186
Light 0.8444 0.2422 0.0158 0.0435 0.0029 0.8559 0.2780 0.6753 0.1714 0.9544
Age 0.0126 0.3627 0.2129 0.1201 0.1344 0.5250 0.5031 0.0229 0.1734 0.1363
Light × Age 0.0954 0.6608 0.7985 0.9007 0.8567 0.5532 0.6938 0.1643 0.1686 0.7176

a-bvalues within columns with different superscripts are significantly different at p ≤ 0.05.

Exp light = Experimental light, EW= Egg weight (g), SG= specific gravity, PA= percent albumen (%), PY= percent yolk (%), YAR=yolk albumen ratio; HU= haugh, ST= shell thickness (mm), SW= shell weight (g), PSW= percent shell weight (%), EBS= eggshell breaking strength (Newtons)

Egg weight was not affected by the red-LED treatment (P = 0.844). The main effect of age was observed with respect to egg weight (P = 0.012) (Table 4) with 53.95g at 22 wk of age compared with 57.68g and 58.30g at 26 and 30 wk of age, respectively. Neither light treatment nor age influenced egg specific gravity (P = 0.242) or eggshell percentage (P = 0.171). Although a numerically higher eggshell percentage was detected in the red-LED (9.57%) compared to normal-LED (9.25%), it was not influenced by the age of the birds (P = 0.173).

Organ Weight

Red-LED negatively influenced relative spleen weight in the birds (P = 0.027; Table 5). A lower relative spleen percentage of 0.12% was found in the red-LED as compared to 0.13% in normal-LED. Furthermore, relative spleen weight significantly decreased as the birds aged (P < 0.001), the relative spleen weight was highest at 18 (0.18%), followed by 22 (0.13%), 26 (0.09%) and 30 (0.07%) wk of age, respectively. These findings are consistent with Xie et al. (2008), who also observed a lower spleen weight in red-LED with 23L:1D as compared to blue-LED and white-LED of the same photoperiod. It has also been reported that lower spleen weight is associated with stress corresponding with corticosterone concentration (Donker, 1989). It can be speculated that birds in red-LED had a weaker immune system as compared to normal-LED. However, it is premature to conclude that the lower relative spleen weight observed in red-LED is due to increased stress. Therefore, other mechanisms, other than stress and serum corticosterone, may have influenced the immunity of birds maintained in red-LED.

Table 5. Effect of LED type, age and their interactions on weight of spleen and brain

Light Age (wk) SW PS BW PB
Red-LED 1.70 0.12b 2.78 0.19
Normal-LED 1.80 0.13a 2.70 0.18
SEM 0.051 0.004 0.074 0.005
18 2.23a 0.18a 2.41 0.20
22 1.99a 0.13b 2.87 0.19
26 1.53b 0.09c 2.80 0.18
30 1.25b 0.07d 2.85 0.17
SEM 0.840 0.004 0.083 0.005
Red-LED 18 2.27 0.18 2.54 0.21
Red-LED 22 1.84 0.12 2.81 0.19
Red-LED 26 1.44 0.09 2.84 0.18
Red-LED 30 1.24 0.07 2.82 0.17
Normal-LED 18 2.18 0.18 2.19 0.19
Normal-LED 22 2.14 0.14 2.92 0.20
Normal-LED 26 1.62 0.10 2.77 0.17
Normal-LED 30 1.26 0.07 2.87 0.18
SEM 0.111 0.011 0.086 0.006
Source of Variation P-value
Year 0.3956 0.0488 0.3543 0.3489
Light 0.3016 0.0274 0.7722 0.7047
Age 0.0007 0.0001 0.0656 0.1489
Light × Age 0.4523 0.3775 0.4599 0.3954

a-dvalues within columns with different superscripts are significantly different at p ≤ 0.05.

SW= spleen weight (g), PS= percent spleen (%), BW= brain weight (g). PB= percent brain (%)

Hormonal Analysis, Tonic Immobility and Heterophil/Lymphocyte Ratio

Our experiment found that serum melatonin levels were not influenced by light treatment (P = 0.565) or age (P = 0.446) from 8:00 AM to 12:00 noon as shown in Table 6. Numerically, mean serum melatonin levels were 86 pg/ml in the red-LED and 95 pg/ml in the normal-LED. In our experiment, numerically lower levels of melatonin were observed in birds raised in red-LED, and this information would be significant for us to address future research. A more elaborate experiment with multiple replicates will be needed to detect the difference in serum melatonin levels between the lights. Similarly, serum LH levels were not affected by light (P = 0.172) or age of the birds (P = 0.696). The mean serum LH concentration in the red-LED was 14 pg/ml and 12 pg/ml in the normal-LED. With the information from previous research and data obtained from this research we still do not have convincing data to conclude that providing a lighting regime of blue LED in the pullet phase and red LED during the production phase would enhance the potential for productivity in birds.

Table 6. Effect of LED type, age and their interactions on concentration of corticosterone, melatonin and luteinizing hormone from 18-30 wk of age

Light Age (wk) CS1 ML1 LH1
Red-LED 102 86 14
Normal-LED 126 95 12
SEM 10.0 9.3 1.4
18 82 120 12
22 99 67 12
26 149 88 13
30 125 588 15
SEM 25.7 18.5 2.2
Red-LED 18 75 103 11
Red-LED 22 84 70 12
Red-LED 26 144 96 14
Red-LED 30 104 76 20
Normal-LED 18 90 138 13
Normal-LED 22 113 64 11
Normal-LED 26 154 80 11
Normal-LED 30 146 100 11
SEM 32.5 27.6 3.1
Source of Variation P-value
Year 0.4844 0.2448 0.0553
Light 0.6467 0.5655 0.1721
Age 0.1836 0.4468 0.6966
Light × Age 0.7857 0.5558 0.3275
Data were log transformed for analysis, but raw values are presented in this table.

CS= Corticosterone (ng/mL), ML= melatonin (pg/mL), LH= luteinizing hormone (mIU/mL)

Mean serum corticosterone levels were not affected by light (P = 0.646) or age (P = 0.183). Birds in the normal LED room had a mean serum corticosterone concentration of 126 ng/ml compared to 102 ng/ml in the experimental LED room. Tonic immobility and H/L ratio data are shown in Table 7. There was no effect of light treatment (P = 0.115) or age (P = 0.098) on TI. The tonic immobility score of 160.98 s was observed in the red-LED as compared to 133.58 s in normal-LED light. There was no significant effect of light treatment (P = 0.544), age of birds (P = 0.615) or their interaction (P = 0.683) on H/L ratio. The H/L ratio in the red-LED was 0.51 and normal-LED was 0.50. Although we did not analyze the bird behavior data, it was observed that birds in the experimental LED (both blue-LED and red-LED) did not follow perching behavior as in the normal-LED room in both years.

Table 7. Effect of LED type, age and their interactions on heterophil (H) and lymphocyte (L) ratio and tonic immobility (TI) from 18-30 wk of age

Light Age (wk) H/L ratio TI1
Red-LED 0.51 160.98
Normal-LED 0.50 133.58
SEM 0.013 17.343
18 0.50 213.04
22 0.48 158.01
26 0.55 114.20
30 0.49 103.87
SEM 0.018 22.872
Red-LED 18 0.47 224.25
Red-LED 22 0.49 139.20
Red-LED 26 0.57 163.16
Red-LED 30 0.53 117.33
Normal-LED 18 0.53 201.83
Normal-LED 22 0.47 176.83
Normal-LED 26 0.54 65.25
Normal-LED 30 0.45 90.41
SEM 0.053 32.346
Source of Variation P-value
Year 0.3413 0.8754
Light 0.5445 0.1159
Age 0.6152 0.0989
Light × Age 0.6832 0.4814
Data was log transformed for analysis, but raw values are presented in this table.

H&L= heterophil and lymphocyte ratio, TI= tonic immobility (s).


The overall egg production was not influenced by the experimental-LED. However, blue-LED has shown an improvement in the pullet body weight as well as stimulated the onset of lay.

It was observed that birds exposed to red-LED had higher relative yolk weight with lower relative albumen weight. A higher yolk albumen ratio could be beneficial for the food industry by targeting production of higher yolk percentage to produce eggs for products that require egg yolk as the main ingredient like mayonnaise, custard, crème brûlée, and eggnog. The yolk also has cosmetic, nutritional, and medical uses. Other egg quality parameters are comparable between the red-LED and normal-LED.

A relatively lower spleen percentage was observed in the red-LED as compared to the normal-LED, which might suggest a weaker immune system, although this is not substantiated by the stress response indicator. Exposing birds to blue-LED in the pullet phase can increase the body weight and initiate lay early.

Heavier body weight at the start of production has been associated with heavier egg throughout the lifespan of birds, more feed consumption and decreased longevity. Therefore, it is difficult to conclude that exposing birds to blue light for improved growth would have a beneficial effect on overall egg production.

Uncited References

Hy-Line 2019Liu et al., 2018aRodriguez and Lea, 1994SAS 9.4 2019

taken from science direct

Journal of Applied Poultry Research
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