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Housing system and laying hen strain impacts on egg microbiology
PRODUCTION, MODELING, AND EDUCATION Housing system and laying hen strain impacts on egg microbiology D. R. Jones*1 and K. E. Anderson† *Egg Safety and Quality Research Unit, USDA Agricultural Research Service, Athens, GA 30605; and †Department of Poultry Science, North Carolina State University, Raleigh 27695 cages had significantly different (P < 0.05) levels of aerobic contamination in relation to hen strain with Hy-Line Silver Brown having the greatest (4.57 log cfu/ mL). Hy-Line Brown and Barred Plymouth Rock hens produced eggs with significantly different (P < 0.01) levels of Enterobacteriaceae among housing systems with conventional caged eggs having the lowest level of contamination for the hen strains. There were no differences within each strain among housing systems for yeast and mold contamination. The study shows that hen strain has an effect on egg microbial levels for various housing systems, and egg safety should be considered when making hen strain selections for each housing system.
Key words: housing system, egg microbiology, laying strain, alternative production 2013 Poultry Science 92:2221–2225 http://dx.doi.org/10.3382/ps.2012-02799
INTRODUCTION Consumers, egg producers, legislatures, consumer groups, as well as animal welfare organizations have a vested interest in how retail eggs are produced. The egg industry has always embraced the request for consumer choice and provides many animal husbandry and nutritional enhancement options for retail eggs. Legislation in Europe went into effect in 2012 banning the use of conventional cages (European Commission, 1999). In the United States, California voters passed a law defining hen housing conditions in that state which goes into effect in 2015 (California Health and Safety Code, 2009). Additionally, a US federal regulation defining minimum hen housing conditions has been proposed and is currently in committee for consideration (US House of Representatives, 2012). Although the various laws and legislations have been proposed and enacted, the impact of the housing requirements on egg safety is not completely understood.
©2013 Poultry Science Association Inc. Received September 25, 2012. Accepted March 31, 2013. 1 Corresponding author: [email protected]
In preparation for the EU hen housing transition, researchers have explored various aspects of the law in member countries on egg microbiology (De Reu et al., 2005, 2006, 2008, 2009; Mallet et al., 2006; Schwaiger et al., 2008; Huneau-Salaün et al., 2009), often presenting conflicting results. Unfortunately, due to the dynamic nature of microbial growth and the differences in husbandry practices and laying hen genetics between the European Union and United States, much of the egg microbial information from the European Union cannot be directly applied to US conditions (Holt et al., 2011). Environmental and egg microbiology affect not only consumer safety, but also the ability of the US egg industry to meet the requirements of the federal law governing egg production and transportation (FDA, 2009). Surveys of the microbial diversity of a variety of commercial egg production systems in Europe have been conducted (De Reu et al., 2006, 2009; Schwaiger et al., 2008; Huneau-Salaün et al., 2010), but very few controlled commercial-style studies have occurred. In the United States, researchers have begun to examine the effects of commercial-scale alternative egg production systems on environmental and egg microbiology (Jones et al., 2011, 2012). The current study was conducted to examine the impact of alternative housing systems and laying hen strain on egg microbiology, thus expanding
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ABSTRACT Alternative hen housing is becoming more commonplace in the egg market. However, a complete understanding of the implications for alternative housing systems on egg safety has not been achieved. The current study examines the impact of housing Hy-Line Brown, Hy-Line Silver Brown, and Barred Plymouth Rock hens in conventional cage, cage-free, and free range egg production systems on shell microbiology. Eggs were collected at 4 sampling periods. Egg shell emulsion pools were formed and enumerated for total aerobic organisms, Enterobacteriaceae, and yeast and mold counts. Hy-Line Brown and Hy-Line Silver Brown hens produced eggs with significantly (P < 0.05 and 0.001, respectively) different levels of aerobic organisms dependent on housing system. Eggs from conventional
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the understanding of US commercial egg production systems on egg safety.
MATERIALS AND METHODS Hen Management
Egg Sample Collection Egg samples were collected according to the methods outlined in Jones et al. (2011) at hen age 23, 32, 41, and 48 wk. For each strain of laying hens, 18 eggs were aseptically collected from conventional cage (CC), cage-free nest box (CFNB), cage-free floor (CFF), and free range nest box (FRNB). Not enough eggs were laid out of the nest boxes in the free range environment by any of the strains for analysis at any sampling period. Three shell emulsion pools of 3 shells were formed in sterile specimen cups for each hen strain/ housing system combination during a sampling period by adding 50 mL of 42°C sterile PBS according to the methods of Musgrove et al. (2005). Cracked eggs were excluded from analysis.
Microbial Assessments Total aerobic populations were determined by duplicate spread plating 100 µL of appropriate dilutions onto standard method agar (Acumedia Manufacturers, Lansing, MI). The plates were incubated at 35°C for 48 h before enumeration. Levels of yeasts and molds were
Statistical Analysis Microbial counts were subjected to log-transformation (SAS Institute, 2002) before analysis. Plate counts with no growth were converted to zero after log-transformation. The Barred Plymouth Rock hens in cagefree production produced floor eggs exclusively in the first sampling period. These data were therefore excluded from analysis. Additionally, Barred Plymouth Rock hens in CC production did not produce eggs at 23 wk of age and Hy-Line Brown hens in cage-free production did not produce floor eggs at 48 wk of age. A mixed model was therefore used for initial statistical analyses with hen strain as the fixed and housing system as the random variables. Due to the biological nature of the study, numerous main effect interactions were found. The data were then sorted for hen strain and analyzed to determine differences between housing systems within a single laying strain for each microbial populations monitored using the GLM. Subsequently, data were analyzed after sorting for housing system to determine differences between hen strains within a single housing system for the monitored microbial populations via the GLM. Means were separated by the least squares method.
RESULTS AND DISCUSSION The concentration of total aerobic organisms associated with the shells of eggs from each strain and housing system is presented in Table 1. When considering each housing system, CC was significantly different among the hen strains (P < 0.05) with the Hy-Line Silver Brown having the greatest level of aerobic bacteria associated with the shell and shell membranes (4.57 log cfu/mL). Although a statistically significant difference was found, biologically, the 0.5 log cfu/mL difference was not of practical commercial importance. When comparing housing systems within a single strain of laying hens, Hy-Line Brown and Hy-Line Silver Brown produced eggs with significantly different (P < 0.05 and P < 0.001, respectively) levels of aerobic organisms in the various production environments. HyLine Brown had significantly different levels of aerobic organisms associated with the shell in the CFF (4.49 log cfu/mL) and CFNB (3.31 log cfu/mL). Hy-Line Silver Brown produced eggs with the greatest level of aer-
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on June 26, 2015
Three strains of laying hens (Hy-Line Brown, HyLine Silver Brown, and Barred Plymouth Rock) were hatched together at the Piedmont Research Station, North Carolina Department of Agriculture and Consumer Services, Salisbury. At the appropriate age, each strain was housed in conventional cage, cage-free, and free range production systems. The full description of rearing and production management can be found in Anderson (2011). Briefly, conventional cage hens were reared in a brood/grow pullet house with quad-deck cage systems (13 birds/cage; 310 cm2/bird). At 17 wk of age, the conventional cage-reared hens were moved to a quad-deck laying house with 4 cage replicates of 6 hens/cage (413 cm2/hen). Cage-free and free range pullets were reared comingled and brooded on litter at a density of 638 cm2/bird. All birds had access to roosts to encourage roosting behavior and usage of nest boxes. At 12 wk of age pens were split, with the range pullets being moved to the range hut at (929 cm2/pullet) with full access to range paddocks (7.91 m2/hen). Nest boxes were available at a rate of 1 nest box/7.5 hens and 13 cm of roosting space/hen. The cage-free house was a litter/slat facility consisting of 2/3 slates and 1/3 litter with 216 hens/ pen (929 cm2/hen), 1 nest box/6 hens, and 13 cm of roosting space/hen.
determined by duplicate spread plating 100 uL of appropriate dilutions onto Dichloran Rose Bengal Chloramphenicol agar (Acumedia Manufacturers). Plates were incubated, right side up, for 6 d at 25 to 26°C before enumeration. Enterobacteriaceae were enumerated by dispensing 1 mL of appropriate dilutions into violet red bile glucose agar (Acumedia Manufacturers) pour plates with overlay. Duplicate plates per sample were incubated at 37°C for 18 to 20 h before typical colonies were counted.
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HOUSING SYSTEM AND EGG MICROBIOLOGY Table 1. Laying strain and housing system effects on egg shell total aerobic organism counts (log Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value
Conventional cage 4.01b,xy 4.57a,x 4.05b 0.19 0.05
Cage-free nest box
Cage-free floor eggs
3.31y 3.68y 3.47 0.22 NS
4.49x 4.32x 0.15 NS
cfu/mL)1
Free range nest box 4.17x 2.92z 3.86 0.37 NS
SE
0.29 0.22 0.35
P-value
0.05 0.001 NS
a,bMeans
within a column with different superscripts are significantly different. within a row with different superscripts are significantly different. 1Lowest n = 9 pools. x–zMeans
The levels of Enterobacteriaceae detected during the study are found in Table 2. Within each housing system, there were no significant differences among the laying hen strains for Enterobacteriaceae detection. The Hy-Line Brown and Barred Plymouth Rock hens produced eggs with significantly (P < 0.01) different levels of Enterobacteriaceae associated with the shells for the various housing systems. Hy-Line Brown hens had the greatest concentration of Enterobacteriaceae associated with FRNB and CFF (1.81 and 1.63 log cfu/ mL, respectively) and similar low levels for CC and CFNB (0.36 log cfu/mL). In the Barred Plymouth Rock strain, similar levels of Enterobacteriaceae were detected FRNB and CFNB (1.86 and 1.12 log cfu/ mL, respectively), which were different from CC where no Enterobacteriaceae were detected during the study. Singh et al. (2009) and Jones et al. (2011) reported the lowest coliform levels associated with conventional cage eggs. The current study also finds the lowest shellrelated Enterobacteriaceae levels in CC eggs. De Reu et al. (2009) found statistical, but not biologically important, differences in Enterobacteriaceae levels on shells from furnished and noncage production systems. In the current study, for the 2 strains exhibiting statistically different levels of Enterobacteriaceae among housing systems, the differences were of a magnitude to be biologically important (>1 log cfu/mL). The impact of laying hen strain and housing system on yeast and mold levels is presented in Table 3. Each of the 3 laying hen strains produced eggs with similar yeast and mold contamination associated with the shell for the various housing systems. There was a significant difference (P < 0.05) in the concentra-
Table 2. Laying strain and housing system effects on egg shell Enterobacteriaceae counts (log cfu/mL)1 Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value x,yMeans
Conventional cage 0.36y 0.23 ND2, y 0.10 NS
Cage-free nest box 0.36y 0.95 1.12x 0.28 NS
within a row with different superscripts are significantly different. n = 9 pools. 2ND = none detected. 1Lowest
Cage-free floor eggs 1.63x 0.94 0.34 NS
Free range nest box 1.81x 0.89 1.86x 0.46 NS
SE
0.38 0.32 0.37
P-value
0.01 NS 0.01
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obic contamination in the CC and CFF (4.57 and 4.32 log cfu/mL, respectively). The lowest level of aerobic contamination for this laying hen strain was detected in FRNB (2.92 log cfu/mL). Barred Plymouth Rock hens produced eggs with similar levels of aerobic contamination across FRNB, CC, and CFNB. The levels of shell aerobic organism contamination detected in the CC for the current study is similar to the average levels reported by Jones et al. (2011). Average FRNB shell aerobic counts for Hy-Line Silver Brown are lower for the current study compared with Jones et al. (2011). Mallet et al. (2006) found greater bacterial loads on eggs laid outside of the nests in furnished cages. In the current study, when significant differences between production systems occurred within a hen strain, CFF eggs were always significantly greater than other production systems. De Reu et al. (2006) also found aerobic levels to be greatest on floor eggs, surmising floor eggs should not be consumed. The authors also reported aerobic levels to be greater for nonconventional cage eggs, which was not the case for the Hy-Line Silver Brown hens in the current study. De Reu et al. (2008) determined aerobic plate counts to be greater associated with eggs from nests of noncage systems compared with nests from furnished or conventional cage production systems. This was not seen for all the hen strains in the current study. De Reu et al. (2008) determined that differences between housing systems are less pronounced under commercial conditions. The current study was conducted on a research farm equipped with commercial style hen housing systems, which could account for the differences in microbial results compared with previously published studies.
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Table 3. Laying strain and housing system effects on egg shell yeast and mold counts (log cfu/mL)1 Conventional cage
Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value
1.31 1.37 0.64 0.28 NS
Cage-free nest box
Cage-free floor eggs
0.99 0.81 0.86 0.21 NS
1.73 1.37 0.15 NS
Free range nest box 1.99a 0.95b 0.84b 0.29 0.05
SE
P-value
0.31 0.22 0.24
NS NS NS
a,bMeans 1Lowest
within a column with different superscripts are significantly different. n = 9 pools.
ACKNOWLEDGMENTS The authors appreciate the laboratory efforts of Patsy Mason, Victoria Broussard, Otis Freeman, and Jerrie Barnett of USDA-ARS, Athens, Georgia. Furthermore, the authors appreciate the flock maintenance efforts of the Piedmont Research Station, Poultry Unit staff in Salisbury, North Carolina.
REFERENCES Anderson, K. E. 2011. Single production cycle report of the thirty eighth North Carolina layer performance and management test: Alternative production environments. Vol. 38, No. 4. North Carolina State University, Cooperative Extension, Raleigh, NC. Accessed Aug. 30, 2012. http://www.ces.ncsu.edu/depts/poulsci/ tech_manuals/layer_reports/38_single_cycle_report.pdf. California Health and Safety Code. 2009. Division 20, Chapter 13.8, Sections 25990–25994. De Reu, K., K. Grijspeerdt, M. Heyndrickx, M. Uyttendaele, J. Debevere, and L. Herman. 2006. Bacterial shell contamination in the egg collection chains of different housing systems for laying hens. Br. Poult. Sci. 47:163–172. De Reu, K., K. Grijspeerdt, M. Heyndrickx, J. Zoons, K. De Baere, M. Uyttendaele, J. Debevere, and L. Herman. 2005. Bacterial eggshell contamination in conventional cages, furnished cages and aviary housing systems for laying hens. Br. Poult. Sci. 46:149–155. De Reu, K., W. Messens, M. Heyndrickx, T. B. Rodenburg, M. Uyttendaele, and L. Herman. 2008. Bacterial contamination of table eggs and the influence of housing systems. World’s Poult. Sci. J. 64:5–19. De Reu, K., T. B. Rodenburg, K. Grijspeerdt, W. Messens, M. Heyndrickx, F. A. M. Tuyttens, B. Sonck, J. Zoons, and L. Herman. 2009. Bacteriological contamination, dirt, and cracks of eggshells in furnished cages and noncage systems for laying hens: An international on-farm comparison. Poult. Sci. 88:2442–2448. European Commission. 1999. Council Directive 1999/74/EC of 19 July 1999: Minimum standards for the protection of laying hens. Off. J. Eur. Communities L203:53–57. FDA. 2009. Prevention of Salmonella Enteritidis in Shell Eggs During Production, Storage, and Transportation; Final Rule. Accessed Aug. 30, 2012. http://edocket.access.gpo.gov/2009/pdf/ E9-16119.pdf. Holt, P. S., R. H. Davies, J. Dewulf, R. K. Gast, J. K. Huwe, D. R. Jones, D. Waltman, and K. R. Willian. 2011. The impact of different housing systems on egg safety and quality. Poult. Sci. 90:251–262. Huneau-Salaün, A., M. Chemaly, S. Le Bouquin, F. Lalande, I. Petetin, S. Rouxel, V. Michel, P. Fravalo, and N. Rose. 2009. Risk factors for Salmonella enterica ssp. enterica contamination in 519 French laying hen flocks at the end of the laying period. Prev. Vet. Med. 89:51–58. Huneau-Salaün, A., V. Michel, D. Huonnic, L. Balaine, and S. Le Bouquin. 2010. Factors influencing bacterial eggshell contamination in conventional cages, furnished cages, and free-range sys-
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tion of yeast and mold detected from FRNB among the laying hen strains. Hy-Line Brown hens produced eggs with a much greater level of yeast and mold than Hy-Line Silver Brown and Barred Plymouth Rock hens (1.99, 0.95, and 0.84 log cfu/mL, respectively). Hy-Line Brown hens produced eggs with a higher average yeast and mold level than Hy-Line Silver Brown and Barred Plymouth Rock hens. The Barred Plymouth Rock hens produced eggs with the lowest average yeast and mold levels across the various housing systems. Jones et al. (2011) reported yeast and mold levels to be significantly different among free range nest box, free range floor, and conventional cage shells. The current study does not find any significant difference in yeast and mold levels among production systems for any of the hen strains. The strain of laying hen appears to affect the populations of indicator organisms associated with the shell in conventional cage, cage-free, and free range production systems. During the current study, none of the hen strains produced enough free range floor eggs to create a sampling pool, indicating all hen strains responded well to the nest boxes in the free range production system. Barred Plymouth Rock hens produced the fewest floor eggs in cage-free production systems. As the study progressed, Hy-Line Brown hens ceased producing floor eggs in the cage-free production system. Singh et al. (2009) found strain of laying hen can affect adaptability to laying environment and nest box usage. Hy-Line Silver Brown hens produced FRNB eggs with significantly lower aerobic bacteria levels compared with other housing systems. All 3 strains of hens produced eggs with the lowest levels of Enterobacteriaceae in CC. Hy-Line Silver Brown hens in all housing systems produced eggs with similar levels of Enterobacteriaceae. Yeast and mold levels were similar across all housing systems for each of the 3 strains of laying hens. Hy-Line Brown hens produced FRNB eggs with significantly greater yeast and mold levels compared with Hy-Line Silver Brown and Barred Plymouth Rock FRNB eggs. More research is needed to fully understand the differences in egg microbial contamination among strains of laying hens for various housing systems. When selecting hen strains for various egg production systems, producers should consider egg microbial integrity to enhance the safety of eggs produced.
HOUSING SYSTEM AND EGG MICROBIOLOGY tems for laying hens under commercial conditions. Br. Poult. Sci. 51:163–169. Jones, D. R., K. E. Anderson, and J. Y. Guard. 2012. Prevalence of coliforms, Salmonella, Listeria, and Campylobacter associated with eggs and the environment of conventional cage and freerange egg production. Poult. Sci. 91:1195–1202. Jones, D. R., K. E. Anderson, and M. T. Musgrove. 2011. Comparison of environmental and egg microbiology associated with conventional and free-range laying hen management. Poult. Sci. 90:2063–2068. Mallet, S., V. Guesdon, A. M. H. Ahmed, and Y. Nys. 2006. Comparison of eggshell hygiene in two housing systems: Standard and furnished cages. Br. Poult. Sci. 47:30–35. Musgrove, M. T., D. R. Jones, J. K. Northcutt, M. A. Harrison, N. A. Cox, K. D. Ingram, and A. J. Hinton Jr. 2005. Recovery of
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Salmonella from commercial shell eggs by shell rinse and shell crush methodologies. Poult. Sci. 84:1955–1958. SAS Institute. 2002. User’s Guide to SAS, Version 9.3. SAS Institute Inc., Cary, NC. Schwaiger, K., E.-M. V. Schmied, and J. Bauer. 2008. Comparative analysis of antibiotic resistance characteristics of gram-negative bacteria isolated from laying hens and eggs in conventional and organic keeping systems in Bavaria, Germany. Zoonoses Public Health 55:331–341. Singh, R., K. M. Cheng, and F. G. Silversides. 2009. Production performance and egg quality of four strains of laying hens kept in conventional cages and floor pens. Poult. Sci. 88:256–264. US House of Representatives. 2012. H.R.3798 Egg Products Inspection Act Amendments of 2012. Accessed Aug. 29, 2012. http:// thomas.loc.gov/cgi-bin/query/z?c112:H.R.3798.
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on June 26, 2015
Key words: housing system, egg microbiology, laying strain, alternative production 2013 Poultry Science 92:2221–2225 http://dx.doi.org/10.3382/ps.2012-02799
INTRODUCTION Consumers, egg producers, legislatures, consumer groups, as well as animal welfare organizations have a vested interest in how retail eggs are produced. The egg industry has always embraced the request for consumer choice and provides many animal husbandry and nutritional enhancement options for retail eggs. Legislation in Europe went into effect in 2012 banning the use of conventional cages (European Commission, 1999). In the United States, California voters passed a law defining hen housing conditions in that state which goes into effect in 2015 (California Health and Safety Code, 2009). Additionally, a US federal regulation defining minimum hen housing conditions has been proposed and is currently in committee for consideration (US House of Representatives, 2012). Although the various laws and legislations have been proposed and enacted, the impact of the housing requirements on egg safety is not completely understood.
©2013 Poultry Science Association Inc. Received September 25, 2012. Accepted March 31, 2013. 1 Corresponding author: [email protected]
In preparation for the EU hen housing transition, researchers have explored various aspects of the law in member countries on egg microbiology (De Reu et al., 2005, 2006, 2008, 2009; Mallet et al., 2006; Schwaiger et al., 2008; Huneau-Salaün et al., 2009), often presenting conflicting results. Unfortunately, due to the dynamic nature of microbial growth and the differences in husbandry practices and laying hen genetics between the European Union and United States, much of the egg microbial information from the European Union cannot be directly applied to US conditions (Holt et al., 2011). Environmental and egg microbiology affect not only consumer safety, but also the ability of the US egg industry to meet the requirements of the federal law governing egg production and transportation (FDA, 2009). Surveys of the microbial diversity of a variety of commercial egg production systems in Europe have been conducted (De Reu et al., 2006, 2009; Schwaiger et al., 2008; Huneau-Salaün et al., 2010), but very few controlled commercial-style studies have occurred. In the United States, researchers have begun to examine the effects of commercial-scale alternative egg production systems on environmental and egg microbiology (Jones et al., 2011, 2012). The current study was conducted to examine the impact of alternative housing systems and laying hen strain on egg microbiology, thus expanding
2221
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on June 26, 2015
ABSTRACT Alternative hen housing is becoming more commonplace in the egg market. However, a complete understanding of the implications for alternative housing systems on egg safety has not been achieved. The current study examines the impact of housing Hy-Line Brown, Hy-Line Silver Brown, and Barred Plymouth Rock hens in conventional cage, cage-free, and free range egg production systems on shell microbiology. Eggs were collected at 4 sampling periods. Egg shell emulsion pools were formed and enumerated for total aerobic organisms, Enterobacteriaceae, and yeast and mold counts. Hy-Line Brown and Hy-Line Silver Brown hens produced eggs with significantly (P < 0.05 and 0.001, respectively) different levels of aerobic organisms dependent on housing system. Eggs from conventional
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the understanding of US commercial egg production systems on egg safety.
MATERIALS AND METHODS Hen Management
Egg Sample Collection Egg samples were collected according to the methods outlined in Jones et al. (2011) at hen age 23, 32, 41, and 48 wk. For each strain of laying hens, 18 eggs were aseptically collected from conventional cage (CC), cage-free nest box (CFNB), cage-free floor (CFF), and free range nest box (FRNB). Not enough eggs were laid out of the nest boxes in the free range environment by any of the strains for analysis at any sampling period. Three shell emulsion pools of 3 shells were formed in sterile specimen cups for each hen strain/ housing system combination during a sampling period by adding 50 mL of 42°C sterile PBS according to the methods of Musgrove et al. (2005). Cracked eggs were excluded from analysis.
Microbial Assessments Total aerobic populations were determined by duplicate spread plating 100 µL of appropriate dilutions onto standard method agar (Acumedia Manufacturers, Lansing, MI). The plates were incubated at 35°C for 48 h before enumeration. Levels of yeasts and molds were
Statistical Analysis Microbial counts were subjected to log-transformation (SAS Institute, 2002) before analysis. Plate counts with no growth were converted to zero after log-transformation. The Barred Plymouth Rock hens in cagefree production produced floor eggs exclusively in the first sampling period. These data were therefore excluded from analysis. Additionally, Barred Plymouth Rock hens in CC production did not produce eggs at 23 wk of age and Hy-Line Brown hens in cage-free production did not produce floor eggs at 48 wk of age. A mixed model was therefore used for initial statistical analyses with hen strain as the fixed and housing system as the random variables. Due to the biological nature of the study, numerous main effect interactions were found. The data were then sorted for hen strain and analyzed to determine differences between housing systems within a single laying strain for each microbial populations monitored using the GLM. Subsequently, data were analyzed after sorting for housing system to determine differences between hen strains within a single housing system for the monitored microbial populations via the GLM. Means were separated by the least squares method.
RESULTS AND DISCUSSION The concentration of total aerobic organisms associated with the shells of eggs from each strain and housing system is presented in Table 1. When considering each housing system, CC was significantly different among the hen strains (P < 0.05) with the Hy-Line Silver Brown having the greatest level of aerobic bacteria associated with the shell and shell membranes (4.57 log cfu/mL). Although a statistically significant difference was found, biologically, the 0.5 log cfu/mL difference was not of practical commercial importance. When comparing housing systems within a single strain of laying hens, Hy-Line Brown and Hy-Line Silver Brown produced eggs with significantly different (P < 0.05 and P < 0.001, respectively) levels of aerobic organisms in the various production environments. HyLine Brown had significantly different levels of aerobic organisms associated with the shell in the CFF (4.49 log cfu/mL) and CFNB (3.31 log cfu/mL). Hy-Line Silver Brown produced eggs with the greatest level of aer-
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on June 26, 2015
Three strains of laying hens (Hy-Line Brown, HyLine Silver Brown, and Barred Plymouth Rock) were hatched together at the Piedmont Research Station, North Carolina Department of Agriculture and Consumer Services, Salisbury. At the appropriate age, each strain was housed in conventional cage, cage-free, and free range production systems. The full description of rearing and production management can be found in Anderson (2011). Briefly, conventional cage hens were reared in a brood/grow pullet house with quad-deck cage systems (13 birds/cage; 310 cm2/bird). At 17 wk of age, the conventional cage-reared hens were moved to a quad-deck laying house with 4 cage replicates of 6 hens/cage (413 cm2/hen). Cage-free and free range pullets were reared comingled and brooded on litter at a density of 638 cm2/bird. All birds had access to roosts to encourage roosting behavior and usage of nest boxes. At 12 wk of age pens were split, with the range pullets being moved to the range hut at (929 cm2/pullet) with full access to range paddocks (7.91 m2/hen). Nest boxes were available at a rate of 1 nest box/7.5 hens and 13 cm of roosting space/hen. The cage-free house was a litter/slat facility consisting of 2/3 slates and 1/3 litter with 216 hens/ pen (929 cm2/hen), 1 nest box/6 hens, and 13 cm of roosting space/hen.
determined by duplicate spread plating 100 uL of appropriate dilutions onto Dichloran Rose Bengal Chloramphenicol agar (Acumedia Manufacturers). Plates were incubated, right side up, for 6 d at 25 to 26°C before enumeration. Enterobacteriaceae were enumerated by dispensing 1 mL of appropriate dilutions into violet red bile glucose agar (Acumedia Manufacturers) pour plates with overlay. Duplicate plates per sample were incubated at 37°C for 18 to 20 h before typical colonies were counted.
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HOUSING SYSTEM AND EGG MICROBIOLOGY Table 1. Laying strain and housing system effects on egg shell total aerobic organism counts (log Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value
Conventional cage 4.01b,xy 4.57a,x 4.05b 0.19 0.05
Cage-free nest box
Cage-free floor eggs
3.31y 3.68y 3.47 0.22 NS
4.49x 4.32x 0.15 NS
cfu/mL)1
Free range nest box 4.17x 2.92z 3.86 0.37 NS
SE
0.29 0.22 0.35
P-value
0.05 0.001 NS
a,bMeans
within a column with different superscripts are significantly different. within a row with different superscripts are significantly different. 1Lowest n = 9 pools. x–zMeans
The levels of Enterobacteriaceae detected during the study are found in Table 2. Within each housing system, there were no significant differences among the laying hen strains for Enterobacteriaceae detection. The Hy-Line Brown and Barred Plymouth Rock hens produced eggs with significantly (P < 0.01) different levels of Enterobacteriaceae associated with the shells for the various housing systems. Hy-Line Brown hens had the greatest concentration of Enterobacteriaceae associated with FRNB and CFF (1.81 and 1.63 log cfu/ mL, respectively) and similar low levels for CC and CFNB (0.36 log cfu/mL). In the Barred Plymouth Rock strain, similar levels of Enterobacteriaceae were detected FRNB and CFNB (1.86 and 1.12 log cfu/ mL, respectively), which were different from CC where no Enterobacteriaceae were detected during the study. Singh et al. (2009) and Jones et al. (2011) reported the lowest coliform levels associated with conventional cage eggs. The current study also finds the lowest shellrelated Enterobacteriaceae levels in CC eggs. De Reu et al. (2009) found statistical, but not biologically important, differences in Enterobacteriaceae levels on shells from furnished and noncage production systems. In the current study, for the 2 strains exhibiting statistically different levels of Enterobacteriaceae among housing systems, the differences were of a magnitude to be biologically important (>1 log cfu/mL). The impact of laying hen strain and housing system on yeast and mold levels is presented in Table 3. Each of the 3 laying hen strains produced eggs with similar yeast and mold contamination associated with the shell for the various housing systems. There was a significant difference (P < 0.05) in the concentra-
Table 2. Laying strain and housing system effects on egg shell Enterobacteriaceae counts (log cfu/mL)1 Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value x,yMeans
Conventional cage 0.36y 0.23 ND2, y 0.10 NS
Cage-free nest box 0.36y 0.95 1.12x 0.28 NS
within a row with different superscripts are significantly different. n = 9 pools. 2ND = none detected. 1Lowest
Cage-free floor eggs 1.63x 0.94 0.34 NS
Free range nest box 1.81x 0.89 1.86x 0.46 NS
SE
0.38 0.32 0.37
P-value
0.01 NS 0.01
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obic contamination in the CC and CFF (4.57 and 4.32 log cfu/mL, respectively). The lowest level of aerobic contamination for this laying hen strain was detected in FRNB (2.92 log cfu/mL). Barred Plymouth Rock hens produced eggs with similar levels of aerobic contamination across FRNB, CC, and CFNB. The levels of shell aerobic organism contamination detected in the CC for the current study is similar to the average levels reported by Jones et al. (2011). Average FRNB shell aerobic counts for Hy-Line Silver Brown are lower for the current study compared with Jones et al. (2011). Mallet et al. (2006) found greater bacterial loads on eggs laid outside of the nests in furnished cages. In the current study, when significant differences between production systems occurred within a hen strain, CFF eggs were always significantly greater than other production systems. De Reu et al. (2006) also found aerobic levels to be greatest on floor eggs, surmising floor eggs should not be consumed. The authors also reported aerobic levels to be greater for nonconventional cage eggs, which was not the case for the Hy-Line Silver Brown hens in the current study. De Reu et al. (2008) determined aerobic plate counts to be greater associated with eggs from nests of noncage systems compared with nests from furnished or conventional cage production systems. This was not seen for all the hen strains in the current study. De Reu et al. (2008) determined that differences between housing systems are less pronounced under commercial conditions. The current study was conducted on a research farm equipped with commercial style hen housing systems, which could account for the differences in microbial results compared with previously published studies.
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Table 3. Laying strain and housing system effects on egg shell yeast and mold counts (log cfu/mL)1 Conventional cage
Item Laying hen strain Hy-Line Brown Hy-Line Silver Brown Barred Plymouth Rock SE P-value
1.31 1.37 0.64 0.28 NS
Cage-free nest box
Cage-free floor eggs
0.99 0.81 0.86 0.21 NS
1.73 1.37 0.15 NS
Free range nest box 1.99a 0.95b 0.84b 0.29 0.05
SE
P-value
0.31 0.22 0.24
NS NS NS
a,bMeans 1Lowest
within a column with different superscripts are significantly different. n = 9 pools.
ACKNOWLEDGMENTS The authors appreciate the laboratory efforts of Patsy Mason, Victoria Broussard, Otis Freeman, and Jerrie Barnett of USDA-ARS, Athens, Georgia. Furthermore, the authors appreciate the flock maintenance efforts of the Piedmont Research Station, Poultry Unit staff in Salisbury, North Carolina.
REFERENCES Anderson, K. E. 2011. Single production cycle report of the thirty eighth North Carolina layer performance and management test: Alternative production environments. Vol. 38, No. 4. North Carolina State University, Cooperative Extension, Raleigh, NC. Accessed Aug. 30, 2012. http://www.ces.ncsu.edu/depts/poulsci/ tech_manuals/layer_reports/38_single_cycle_report.pdf. California Health and Safety Code. 2009. Division 20, Chapter 13.8, Sections 25990–25994. De Reu, K., K. Grijspeerdt, M. Heyndrickx, M. Uyttendaele, J. Debevere, and L. Herman. 2006. Bacterial shell contamination in the egg collection chains of different housing systems for laying hens. Br. Poult. Sci. 47:163–172. De Reu, K., K. Grijspeerdt, M. Heyndrickx, J. Zoons, K. De Baere, M. Uyttendaele, J. Debevere, and L. Herman. 2005. Bacterial eggshell contamination in conventional cages, furnished cages and aviary housing systems for laying hens. Br. Poult. Sci. 46:149–155. De Reu, K., W. Messens, M. Heyndrickx, T. B. Rodenburg, M. Uyttendaele, and L. Herman. 2008. Bacterial contamination of table eggs and the influence of housing systems. World’s Poult. Sci. J. 64:5–19. De Reu, K., T. B. Rodenburg, K. Grijspeerdt, W. Messens, M. Heyndrickx, F. A. M. Tuyttens, B. Sonck, J. Zoons, and L. Herman. 2009. Bacteriological contamination, dirt, and cracks of eggshells in furnished cages and noncage systems for laying hens: An international on-farm comparison. Poult. Sci. 88:2442–2448. European Commission. 1999. Council Directive 1999/74/EC of 19 July 1999: Minimum standards for the protection of laying hens. Off. J. Eur. Communities L203:53–57. FDA. 2009. Prevention of Salmonella Enteritidis in Shell Eggs During Production, Storage, and Transportation; Final Rule. Accessed Aug. 30, 2012. http://edocket.access.gpo.gov/2009/pdf/ E9-16119.pdf. Holt, P. S., R. H. Davies, J. Dewulf, R. K. Gast, J. K. Huwe, D. R. Jones, D. Waltman, and K. R. Willian. 2011. The impact of different housing systems on egg safety and quality. Poult. Sci. 90:251–262. Huneau-Salaün, A., M. Chemaly, S. Le Bouquin, F. Lalande, I. Petetin, S. Rouxel, V. Michel, P. Fravalo, and N. Rose. 2009. Risk factors for Salmonella enterica ssp. enterica contamination in 519 French laying hen flocks at the end of the laying period. Prev. Vet. Med. 89:51–58. Huneau-Salaün, A., V. Michel, D. Huonnic, L. Balaine, and S. Le Bouquin. 2010. Factors influencing bacterial eggshell contamination in conventional cages, furnished cages, and free-range sys-
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tion of yeast and mold detected from FRNB among the laying hen strains. Hy-Line Brown hens produced eggs with a much greater level of yeast and mold than Hy-Line Silver Brown and Barred Plymouth Rock hens (1.99, 0.95, and 0.84 log cfu/mL, respectively). Hy-Line Brown hens produced eggs with a higher average yeast and mold level than Hy-Line Silver Brown and Barred Plymouth Rock hens. The Barred Plymouth Rock hens produced eggs with the lowest average yeast and mold levels across the various housing systems. Jones et al. (2011) reported yeast and mold levels to be significantly different among free range nest box, free range floor, and conventional cage shells. The current study does not find any significant difference in yeast and mold levels among production systems for any of the hen strains. The strain of laying hen appears to affect the populations of indicator organisms associated with the shell in conventional cage, cage-free, and free range production systems. During the current study, none of the hen strains produced enough free range floor eggs to create a sampling pool, indicating all hen strains responded well to the nest boxes in the free range production system. Barred Plymouth Rock hens produced the fewest floor eggs in cage-free production systems. As the study progressed, Hy-Line Brown hens ceased producing floor eggs in the cage-free production system. Singh et al. (2009) found strain of laying hen can affect adaptability to laying environment and nest box usage. Hy-Line Silver Brown hens produced FRNB eggs with significantly lower aerobic bacteria levels compared with other housing systems. All 3 strains of hens produced eggs with the lowest levels of Enterobacteriaceae in CC. Hy-Line Silver Brown hens in all housing systems produced eggs with similar levels of Enterobacteriaceae. Yeast and mold levels were similar across all housing systems for each of the 3 strains of laying hens. Hy-Line Brown hens produced FRNB eggs with significantly greater yeast and mold levels compared with Hy-Line Silver Brown and Barred Plymouth Rock FRNB eggs. More research is needed to fully understand the differences in egg microbial contamination among strains of laying hens for various housing systems. When selecting hen strains for various egg production systems, producers should consider egg microbial integrity to enhance the safety of eggs produced.
HOUSING SYSTEM AND EGG MICROBIOLOGY tems for laying hens under commercial conditions. Br. Poult. Sci. 51:163–169. Jones, D. R., K. E. Anderson, and J. Y. Guard. 2012. Prevalence of coliforms, Salmonella, Listeria, and Campylobacter associated with eggs and the environment of conventional cage and freerange egg production. Poult. Sci. 91:1195–1202. Jones, D. R., K. E. Anderson, and M. T. Musgrove. 2011. Comparison of environmental and egg microbiology associated with conventional and free-range laying hen management. Poult. Sci. 90:2063–2068. Mallet, S., V. Guesdon, A. M. H. Ahmed, and Y. Nys. 2006. Comparison of eggshell hygiene in two housing systems: Standard and furnished cages. Br. Poult. Sci. 47:30–35. Musgrove, M. T., D. R. Jones, J. K. Northcutt, M. A. Harrison, N. A. Cox, K. D. Ingram, and A. J. Hinton Jr. 2005. Recovery of
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Salmonella from commercial shell eggs by shell rinse and shell crush methodologies. Poult. Sci. 84:1955–1958. SAS Institute. 2002. User’s Guide to SAS, Version 9.3. SAS Institute Inc., Cary, NC. Schwaiger, K., E.-M. V. Schmied, and J. Bauer. 2008. Comparative analysis of antibiotic resistance characteristics of gram-negative bacteria isolated from laying hens and eggs in conventional and organic keeping systems in Bavaria, Germany. Zoonoses Public Health 55:331–341. Singh, R., K. M. Cheng, and F. G. Silversides. 2009. Production performance and egg quality of four strains of laying hens kept in conventional cages and floor pens. Poult. Sci. 88:256–264. US House of Representatives. 2012. H.R.3798 Egg Products Inspection Act Amendments of 2012. Accessed Aug. 29, 2012. http:// thomas.loc.gov/cgi-bin/query/z?c112:H.R.3798.
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