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Different Effects of Individual Identification Systems on Chicken Well-Being¹

R. L. Dennis^*,, A. G. Fahey^*, and H. W. Cheng^*,2

^* Livestock Behavior Research Unit, USDA-ARS, West Lafayette, IN 47907; and Purdue University, Department of Animal Science, West Lafayette, IN 47907

² Corresponding author: Heng-wei.Cheng{at}ars.usda.gov

	ABSTRACT

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Individual identification is a common method used in animal research. This study was designed to examine if commonly used identification systems (i.e., leg bands, wing bands, neck tags, and livestock markers) have different effects on hens’ behavioral and physiological homeostasis. At 18 wk of age, hens were paired in all combinations of treatments and control (unmarked hens; n = 10) in a novel cage for 5 trials of 1 h each to test the effects of identification markers on social behaviors. Wing-banded hens tended to exhibit increased feather pecking compared with control hens (P < 0.10), suggesting a slight increase in social stress. No effect of identification treatment was evident on frequency of aggressive behaviors (P > 0.10). At 20 wk of age, absolute fluctuating asymmetry (FA), but not relative FA, of shank length and width was more significant in leg-banded hens (P < 0.05) and tended to be significant in wing-banded hens (P < 0.10), compared with control hens. Asymmetry of the shank is often a result of high stress levels, including social stress. Body weight measured at 20 wk of age showed that hens with leg bands were significantly lighter than control hens (P < 0.05), possibly as a result of decreased access to resources, increased metabolism, or decreased appetite due to elevated stress. Increased absolute FA and decreased BW could be evidence of a disruption of the hens’ physiological homeostasis due to increased stress. Hens with leg bands also tended to have lower percentage of heterophils (P < 0.10), indicative of increased stress and reduced immunocompetence. Our findings provide clear evidence of the negative effects of wing and leg band identification systems on hens’ well-being, altering both physiological and behavioral homeostasis. Without knowledge of the effects, the use of individual identification systems could lead to misinterpretation of experimental results.

Key Words: chicken • identification system • wing band • leg band • livestock marker • swiftack tag

	INTRODUCTION

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Evolutionary scientists have shown that selection, including kin and sexual selection, is often based on physical characteristics. Hamilton and Zuk’s (1982) parasite hypothesis suggests that plumage color and condition can be indicative of parasite resistance or susceptibility. Similarly, Pagel and Dawkins’ (1997) status badge hypothesis supports that birds confer information regarding their social status through distinct physical features such as the presence of extremely large combs or other distinguishing markings. Easily interpretable status badges may serve to conserve energy and reduce injury because birds may use them in place of antagonistic interactions to assess the relative social dominance of competitors.

Identification marking has become an integral part of experimental research incorporating animals and birds. Artificial markings are described in detail in the legislation of the United Kingdom’s Animal Act and the United States Animal Welfare Act. Marks are employed in numerous disciplines, including behavior (Estevez et al., 2003; Dennis, 2004), wildlife management (Mellor et al., 2004), and reproduction studies (Broderick and Godley, 1999). However, the implications of using different identification systems are often omitted from methodological sections in the literature. An in-depth analysis of the effects of marks on chickens determined that livestock markers on the head and neck areas of the bird caused marked birds to meet with increased aggression from conspecifics (Dennis, 2004). Marking birds also decreases the amount of aggression the marked bird will be likely to exhibit and creates an archetype of increased stress alongside depressed sympathetic-adrenal-medullary responsiveness, especially when marked birds make up only a small portion of the group (Dennis, 2004). Additionally, modifications to the bird’s appearance, such as alterations to the feathers and comb, have been shown to affect its behavior and the manner in which conspecifics behave toward it (Guhl, 1953; Guhl and Ortman, 1953). Following physical alteration by dubbing (removal of wattle and comb), altered roosters were shown to be the recipients of increased aggression (Marks et al., 1960; Siegel and Hurst, 1962). In addition, leg banding, a common identification system used in song birds, has been shown to alter mate choice and mate guarding behavior (Burley, 1988; Johnsen et al., 1997, 2000). Zebra finches have shown color and symmetry preferences for leg bands, affecting female-male associations (Burley, 1988; Jennions, 1998).

Increased aggression has been linked to lowered coping ability to stress and reduced immunocompetence (Cheng et al., 2001; Dennis et al., 2004). Some marks may be suggestive of the presence of parasites (Hamilton and Zuk, 1982), resulting in the marked individual receiving increased aggression from conspecifics. Marks and tags may also mimic status badges, thereby communicating incorrect information regarding dominance status, resulting in the individual facing different social challenges (Dennis, 2004). The current study was designed to investigate the effects of various marking systems to determine which system has the mildest effect on the bird itself and the way in which conspecifics behave toward it.

	MATERIALS AND METHODS

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Experimental Design

At 16 wk of age W-36 (Hy-Line Farms Inc.), White Leghorn chickens were transported from a grower facility at Hy-Line Farms to a layer facility at Purdue University. Hens were pair housed in standard layer cages (542 cm²/bird). One bird per cage was randomly selected and was assigned to 1 of 4 artificial marking treatments: leg bands, wing bands, neck tags from the Swiftack system (Heartland Animal Health Inc., Fair Play, MO), and livestock marker applied to the tail area or unmarked control (10 cages with 2 birds per cage were used for each of the 4 treatments and the unmarked control group, and cage was considered as the experimental unit). The control birds remained unmarked and were identified visually throughout the experiment by digital photographs taken on the day of treatment assignment, and these images were used as an identification reference so that behavioral and physiological measures could be taken from the same bird. The Swiftack neck tag, leg band, and livestock marker identification systems used were all blue, whereas the wing bands used were silver colored. The second, unmarked bird per cage was a companion bird to the experimental bird only and not used in behavioral or physiological analysis. Birds were given 2 wk to become acclimated to their identification system before behavioral analysis.

Behavior Analysis—Novel Cage Testing

Behavior testing in a novel cage environment was carried out at 18 and 19 wk of age. Birds of each artificial marking treatment and control birds were paired sequentially, and behavior was recorded to DVD for a 30-min period beginning when the cage door was shut after the second bird entered the cage. Frequency of aggressive behaviors and feather pecking behaviors were recorded using the ethogram in Table 1.

Blood samples were collected from chickens at 20 wk of age from the bronchial vein within 2 min of being removed from the cage. Blood samples were collected in EDTA coated tubes using a 25-gauge needle, centrifuged for 15 min at 700 x g to extract plasma, and plasma was stored at –80 C until the radioimmunoassay. Total plasma corticosterone was measured using a commercially available ¹²⁵I corticosterone radioimmunoassay kit (MP Biomedicals, Catalog No. 07–120122) as outlined by Cheng et al. (2001).

Heterophil to Lymphocyte Ratio

Unheparinized blood samples were smeared on a slide using a cover glass and then stained using Wright’s stain. Using a 2,000x magnification 100 leukocytes were counted. Eosinophils, basophils, monocytes, heterophils, and lymphocytes were identified using discriptions by Campbell (1988). Percent heterophil and lymphocyte were calculated and used in statistical analyses.

Body Weight and Fluctuating Asymmetry

Body weight was assessed before marking at 16 wk of age and again following testing at 20 wk of age. Shank length and width were measured using dial calipers. Fluctuating asymmetry (FA) was assessed as the absolute difference in shank width plus absolute difference in shank length, using the equation below:

To minimize disturbance to the birds’ final BW, FA and blood sampling were taken at the same time.

Statistical Analysis

Data were analyzed using a mixed model ANOVA. A random block design was used to control for the location within the house. For behavioral data analysis, a Latin square design was used to control for day tested. Log-transformation was used in analysis of white blood cell count and corticosterone. The experimental unit used for the split plot analysis was the marking treatment by cage, where cage was considered the experimental whole plot unit. Least squares means and SEM were reported for all groups. Contrasts were used to determine significance using the Sidak adjustment to maintain an experimental of 0.05. Trends were assigned if was less than 0.10, above which we assigned no difference. Data were analyzed using PROC MIXED of SAS 8.2 software (SAS Institute Inc., Cary, NC) main effects marked treatment, all interactions between main effects were considered.

	RESULTS

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Aggressive Behaviors

No significant effect of marking system was observed in any of the aggressive behaviors given or received between the treatments (P > 0.10; n = 10). There was also no difference found in the total aggression among the tested birds of all marked treatments, given or received (P > 0.10; n = 10).

Feather Pecking

Feather pecking tended to increase in wing-banded birds compared with control birds (21 ± 3.6 compared with 13 ± 3.8 feather pecks per bird per hour; P < 0.10; n = 10). No difference in feather pecking was detected in birds bearing leg bands, markers, or tags (P > 0.10). There was no difference in feather pecking given by the treatments (P > 0.10).

Body Weight

Initial BW, taken before treatments were applied, were not different between treatment groups (P > 0.10). However, leg-banded birds were significantly lighter at the final BW compared with the C birds (P < 0.05; n = 10; Figure 1). Final BW was not different from the control birds in any other treated group (P > 0.10)

View larger version (10K): [in this window] [in a new window]   Figure 1. Effects of identification systems on final BW. Data were presented as least squares mean body weight (±SEM; n = 10). **Significant difference (P < 0.05) compared with the controls. C = unmarked control, LB = leg band, LM = livestock marker, ST = swiftack tags, and WB = wing band.

Leg-banded birds had shanks that were significantly more asymmetric than the control birds (P < 0.05; n = 10; Figure 2), whereas wing-banded birds tended to be more asymmetric (P < 0.10). No difference in FA was detected in birds bearing Swiftack tags or livestock markers (P > 0.10).

View larger version (9K): [in this window] [in a new window]   Figure 2. Effects of identification systems on fluctuating asymmetry (FA). Data were presented as least squares mean fluctuating asymmetry (±SEM; n = 10). **Significant difference (P < 0.05) compared with control. *A tendency of difference (P < 0.10) compared with the controls. C = unmarked control, LB = leg band, LM = livestock marker, ST = swiftack tags, and WB = wing band.

Proportion of heterophils were significantly reduced in leg-banded chickens compared with controls in a while blood cell differentiation count (P < 0.05; n = 10; Figure 3). No difference in heterophil count was noted in the birds with wing bands, tags, or livestock markers (P > 0.10). There was also no difference in percentage of lymphocytes found in any of the marking treatments used in this study (P > 0.10).

View larger version (9K): [in this window] [in a new window]   Figure 3. Effects of identification systems on percentage of heterophils. Data were presented as least squares mean percentage of heterophils (±SEM; n = 10). **Significant difference (P < 0.05) compared with control. C = unmarked control, LB = leg band, LM = livestock marker, ST = swiftack tags, and WB = wing band.

Plasma corticosterone levels were significantly reduced in wing-banded birds compared with the unmarked control birds (P < 0.05; n = 10; Figure 4). No difference in corticosterone levels was found in birds from the other identification systems used in this study (P > 0.10).

View larger version (8K): [in this window] [in a new window]   Figure 4. Effects of identification systems on plasma corticosterone concentration (Cort. conc.). Data were presented as least squares mean corticosterone concentration (±SEM; n = 10). **Significant difference (P < 0.05) compared with the controls. C = unmarked control, LB = leg band, LM = livestock marker, ST = swiftack tags, and WB = wing band.

	DISCUSSION

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Previous studies have shown that manipulation of a bird’s appearance through application of markings may give one animal a perceived disadvantage in antagonistic confrontations (Dennis, 2004). In that study, broilers with livestock marker on the head and neck attract increased aggression from unmarked conspecifics. In the present study we show that pair-caged laying hens with livestock marker on the tail, neck tag, leg band, or wing band exhibited no difference in aggression or feather pecking given or received compared with unmarked control birds. The previous and present results may suggest that the location and color of the marking alters the aggressive response from conspecifics. In chickens, as with numerous other species of birds, status signals are often broad-cast through ornamental traits located around the head and neck such as the comb and wattles as well as specialized plumage around the head and neck (Zuk et al., 1990; Belthoff and Gauthreaux, 1991).

The color of the marking itself may alter the intensity or meaning of the signal. In this study all markings were blue, except for the wing bands that were silver in color. Neither color is naturally found in the domestic chicken skin or plumage; therefore, the signal quality of markings might be lessened. Metz and Weatherhead (1991) found that, in a natural environment, birds wearing red leg bands were attacked and displaced from their territories by conspecifics more frequently than birds with black leg bands. In contrast we found, hens with blue leg bands or other blue markings in this study did not receive any excess aggression or feather pecking from conspecifics. Birds marked with wing bands, the only silver-colored mark used in this study, showed a tendency for increased prevalence of feather pecking. Wing-banded birds also exhibited significantly reduced corticosterone levels. Reduced corticosterone response has been shown to be a result of adaptation to chronic stress (Armario et al., 1986), including social stress. The color of the marking in combination with the type of mark used may alter the effects on physiology and behavior and should therefore be tested before use in a research study.

Similarly, asymmetry of shank development has been shown to result from elevated stress during development. Previous studies have shown that different colored leg bands may signal additional information to conspecifics, altering both mate guarding behavior and mate choice in song birds (Burley, 1981, 1988). In the present study, however, all leg-banded birds wore one blue band, a similar color to that used in the livestock marker and neck tagged treatments. Although no difference in aggressive or feather pecking behavior was noted in these birds, these birds did exhibit physiological signs of increased stress. Leg-banded birds had reduced BW and percentage of heterophils, along with increased asymmetry in the shank. These data may be suggestive of an increase stimulation of the sympathetic-medullary-adrenal axis and subsequent suppression of immunocompetence. One potential explanation for this is that color-related signals are location dependent, exhibiting a color x location interaction in poultry communication. Blue color on the leg and silver color on the wing may be perceived by conspecifics as a subordinate status signal.

Behavioral researchers often avoid using wing banding systems as their sole means of identification because they are difficult to see from a distance. Leg bands are also avoided in certain housing systems, such as cages with solid metal panels at the bottom, for the same reason. However, questions are often raised by the use of more conspicuous markings ideal for long-distance observation such as livestock marker and tags. Concerns have become even more prevalent since wildlife research has determined that large and bulky radio telemetry tags can alter reproductive behaviors, migratory patters, and survivability (Brubeck et al., 1981; Brua, 1998). So why are the markings that are less conspicuous to humans more conspicuous to chickens? Previous studies have shown that degree of symmetry in markings, such as leg bands, alter animal behaviors such as mate choice and guarding as well as reproductive success (Swaddle, 1996; Jennions, 1998). Similarly, FA in natural traits is also correlated with inferior reproductive quality and subordinate status (McGary et al., 2003). Both leg and wing band systems were applied in this study to only one side of the body, whereas Swiftack tags and livestock marker were applied in a more symmetric manner. The asymmetry of these markings may present a signal to conspecifics such as subordinate status or inferior reproductive quality. A signal of low status would not necessarily increase the aggression received by these birds because Pagel and Dawkins (1997) propose that status signals are used by birds to form dominance hierarchies to avoid spending energy on agonistic confrontations to determine dominance status. However, externally dictated subordinate status has been shown in previous appearance manipulation studies to cause elevated chronic stress to manipulated individuals (Guhl, 1968).

Both leg and wing bands are made of metal, and although they are lightweight, both are heavier than paper tags. In addition these bands were both positioned on a limb near a frequently used joint. The weight and the location of these bands may cause rubbing, chaffing, or general discomfort at the joint. This discomfort could alter movement causing these animals to move less or have restricted movements to adapt to the presence of the bands. Altered movement style, such as through lameness, has been shown to increase stress and alter behavioral interactions with conspecifics. Injured animals are often met with increased aggression from conspecifics, but this was not seen in the present study. The metal bands used in this study did not cause debilitating injuries. Likely, the wing bands may simply restrict the range of movement in the limbs, giving the bird an actual or perceived disadvantage in antagonistic contests, causing them to avoid excess antagonistic confrontations and relegating them to a more subordinate status.

Conclusions

Artificial identification systems are often chosen by researchers on the basis of their visibility to the researcher, invasiveness of marking, capacity to hold required information, and ability for researchers to easily distinguish between multiple individuals when required by the experimental design. However, the artificial identification systems may have a greater impact on experimental results than previously thought. Marking systems cannot simply be dictated by experimental design, but should be considered along with treatment in designing a balanced and reliable study to assure that the effects of marking are not confounded in experimental treatments. Although leg banding is a less invasive marking method than wing banding or tagging systems, here we have shown that they have the greatest effect on the physiology of the bird. Our findings also illustrate the welfare concerns with the use of wing and leg banding systems in pair-housed hens because they increase the stress of the bird. Livestock marker and neck tags provide a more well-being-friendly alternative for hens tested in pair-caged systems. Finally, our results indicate the necessity for all marking systems to be tested on the species and sex on which they are to be used to preserve the integrity of our experimental conclusions.

	ACKNOWLEDGMENTS

 
The authors thank Inma Estevez for her advice and Fred Haan and the staff at the Purdue Poultry facility as well as the graduate researchers and staff of the Livestock Behavior Research Unit for their hard work and dedication.

	FOOTNOTES

 
¹ This research was funded by a competitive grant from the Animal Welfare Institute to R. L. Dennis and A. G. Fahey. Disclaimer: The mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the US Department of Agriculture.

Received for publication June 12, 2007. Accepted for publication March 4, 2008.

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This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Dennis, R. L. Articles by Cheng, H. W. Search for Related Content PubMed PubMed Citation Articles by Dennis, R. L. Articles by Cheng, H. W.