The relationship between the duration of fecal shedding and the production of contaminated eggs by laying hens infected with strains of Salmonella enteritidis and Salmonella Heidelberg

Egg contamination by Salmonella Enteritidis has remained a significant public health problem for nearly two decades, and Salmonella Heidelberg has also been recently implicated in egg-transmitted human illness. Colonization of the intestinal tract is a necessary precursor to the invasion of reproductive organs and subsequent deposition inside eggs laid by infected hens, but the relationship between the persistence of Salmonella in the intestinal tract and the likelihood of egg contamination has been uncertain. In this study, groups of laying hens were inoculated with large oral doses of strains of Salmonella Enteritidis and Salmonella Heidelberg, including variants of the original parent strains that had been reisolated from eggs laid by infected hens in a prior study. The shedding of Salmonella in voided feces was monitored for 6 wk postinoculation, and all eggs laid by infected hens between 5 and 22 days postinoculation were cultured for Salmonella in their contents. The mean duration of fecal shedding was significantly longer for the previously passaged Salmonella strains (26.7 days) than for the original parent strains (17.5 days), and the passaged strains caused a significantly higher frequency of egg contamination (6.4%) than did the parent strains (3.3%). However, the duration of fecal shedding and the frequency of egg contamination were not correlated for any of the Salmonella Enteritidis or Salmonella Heidelberg strains.     

Colonization of specific regions of the reproductive tract and deposition at different locations inside eggs laid by hens infected with Salmonella enteritidis or Salmonella heidelberg

Internal contamination of eggs by Salmonella Enteritidis has been a significant source of human illness for several decades and is the focus of a recently proposed U.S. Food and Drug Administration regulatory plan. Salmonella Heidelberg has also been identified as an egg-transmitted human pathogen. The deposition of Salmonella strains inside eggs is apparently a consequence of reproductive tissue colonization in infected laying hens, but the relationship between colonization of specific regions of the reproductive tract and deposition in different locations within eggs is not well documented. In the present study, groups of laying hens were experimentally infected with large oral doses of Salmonella Heidelberg, Salmonella Enteritidis phage type 13a, or Salmonella Enteritidis phage type 14b. For all of these isolates, the overall frequency of ovarian colonization (34.0%) was significantly higher than the frequency of recovery from either the upper (22.9%) or lower (18.1%) regions of the oviduct. No significant differences were observed between the frequencies of Salmonella isolation from egg yolk and albumen (4.0% and 3.3%, respectively). Some significant differences between Salmonella isolates were observed in the frequency of recovery from eggs, but not in the frequency or patterns of recovery from reproductive organs. Accordingly, although the ability of these Salmonella isolates to colonize different regions of the reproductive tract in laying hens was reflected in deposition in both yolk and albumen, there was no indication that any specific affinity of individual isolates for particular regions of this tract produced distinctive patterns of deposition in eggs.     

Production of Salmonella enteritidis-contaminated eggs by experimentally infected hens

Laying hens of three different ages were experimentally infected with a strain of Salmonella enteritidis by either oral inoculation or contact transmission. Total egg production was depressed in exposed hens of all three age groups. Persistent intestinal shedding was observed in a small number of hens. Eggs with contents contaminated by S. enteritidis were produced by exposed hens at a high frequency, but only during a fairly short period of time that extended through approximately 1 week postinoculation for older hens and through 2 weeks for younger hens. S. enteritidis was recovered from whole yolks and albumen of these eggs at similar frequencies, but not from the content of yolks. Eggs with contaminated shells were also produced, but at a lower frequency. Contaminated eggs were produced by orally inoculated and contact-exposed hens at similar frequencies. S. enteritidis was not isolated from the contents of eggs laid by hens infected with other S. enteritidis strains.     

Effect of prior serial in vivo passage on the frequency of Salmonella enteritidis contamination in eggs from experimentally infected laying hens

Experimental infection models are valuable tools for understanding and preventing the deposition of Salmonella enteritidis inside eggs. Oral inoculation is believed to closely simulate naturally occurring S. enteritidis infections of chickens, but oral infection studies have often generated relatively low frequencies of egg contamination. The present study assessed whether repeated in vivo passage of an S. enteritidis strain could affect its ability to cause egg contamination in experimentally infected hens. The incidence of egg contamination was determined in groups of hens inoculated orally with either a phage type 13a S. enteritidis strain or derivatives of this parent strain that were obtained by three successive rounds of passage and reisolation from tissues of infected hens. Passaged S. enteritidis isolates recovered from ovaries and oviducts induced a significantly higher incidence of egg contamination (16.97%) than was attributed to the parent strain (8.27%). However, passaged S. enteritidis isolates recovered from livers and spleens were not associated with a significantly increased frequency of deposition in eggs. By either inducing or selecting for the expression of relevant microbial properties, passage of S. enteritidis through reproductive tissues of chickens may be useful for improving the efficiency at which experimental infection models produce egg contamination.     

Specific adhesion and invasion of Salmonella Enteritidis in the vagina of laying hens

Salmonella Enteritidis is the predominant serovar associated with egg-borne salmonellosis in humans. The colonization of S. Enteritidis in the vagina may play a role in the production of S. Enteritidis-contaminated eggs. In the first experiment, the in vitro adhesion of S. Enteritidis in vaginal and follicular explants was compared with that of S. Typhimurium by bacteriological isolation methods. The mean number of S. Enteritidis associated with vaginal explants was significantly (P < 0.05) higher than S. Typhimurium associated with vaginal explants and both serovars associated with follicular explants. In the second experiment, the in vitro adhesion and invasion of S. Enteritidis strains in the vaginal epithelium was compared with that of several strains of S. Agona, S. Infantis, S. Hadar, S. Heidelberg, S. Montevideo and S. Typhimurium, by immunohistochemical methods. The mean number of Salmonella in the vaginal epithelium depended on their lipopolysaccharide (LPS) type, with the rank order as follows: LPS type O9 (S. Enteritidis) > LPS type O4 (S. Agona, S. Typhimurium and S. Heidelberg) > LPS type O7 (S. Montevideo and S. Infantis) and LPS type O8 (S. Hadar). This rank order of Salmonella invasiveness is in accordance with the frequency of Salmonella outbreaks involving contaminated eggs. These findings suggest that S. Enteritidis has a higher ability to colonize the vaginal epithelium than other serovars, and the Salmonella LPS type may play an essential role in tropism of the reproductive tract.     

Selenium deposition in tissues and eggs of laying hens given surplus of selenium as selenomethionine

Forty-eight Norwegian bred White Leghorn chickens were divided into 6 groups and fed a basal diet containing 0.30 mig Se/kg supplemented with 0, 0.1, 0.5, 1.0, 3.0 or 6.0 mg Se/ kg in the form of selenomethionine for 18 weeks. A supplement of only 0.1 mg Se/kg induced significantly higher selenium concentrations in breast muscle and eggs, particularly in the egg white. The increase of selenium in the tissue and egg was proportional to the amounts of selenomethionine added to the feed. In the group given 6.0 mg Se/kg, the selenium concentrations in all tissues and eggs analysed ranged from 4.8 to 7.3 μg Se/g. No signs of toxic effects were observed even at the highest intake of selenium. Excess supply of selenium as selenomethionine to chickens was shown to be more potent than sodium selenite in raising the selenium concentration in tissues and eggs. A supplementation up to 10 times the requirement did not increase the levels of selenium in poultry products to such a degree that they could be considered as a potential risk for human consumption.Key words: dietary selenium, laying hens, selenium concentrations, tissues, eggs     

A comparison of transmission characteristics of Salmonella enterica serovar Enteritidis between pair-housed and group-housed laying hens

ABSTRACT: Human cases of bacterial gastro-enteritis are often caused by the consumption of eggs contaminated with Salmonella species, mainly Salmonella enterica serovar Enteriditis (Salmonella Enteritidis). To reduce human exposure, in several countries worldwide surveillance programmes are implemented to detect colonized layer flocks. The sampling schemes are based on the within-flock prevalence, and, as this changes over time, knowledge of the within-flock dynamics of Salmonella Enteritidis is required. Transmission of Salmonella Enteritidis has been quantified in pairs of layers, but the question is whether the dynamics in pairs is comparable to transmission in large groups, which are more representative for commercial layer flocks. The aim of this study was to compare results of transmission experiments between pairs and groups of laying hens. Experimental groups of either 2 or 200 hens were housed at similar densities, and 1 or 4 hens were inoculated with Salmonella Enteritidis, respectively. Excretion was monitored by regularly testing of fecal samples for the presence of Salmonella Enteritidis. Using mathematical modeling, the group experiments were simulated with transmission parameter estimates from the pairwise experiments. Transmission of the bacteria did not differ significantly between pairs or groups. This finding suggests that the transmission parameter estimates from small-scale experiments might be extrapolated to the field situation.     

Pathologic and bacteriologic findings in 27-week-old commercial laying hens experimentally infected with Salmonella enteritidis, phage type 4

Two strains of 27-wk-old commercial laying chickens (strain A, brown-egg-laying type and strain B, white-egg-laying type) were inoculated either orally (PO) or intravenously (IV) with a field isolate of Salmonella enteritidis phage type 4. Chickens were sequentially necropsied at regular intervals throughout the 17-wk observation period. Gross and microscopic lesions were most evident between 1 and 14 days postinoculation (DPI). Gross lesions consisted of enlarged livers with white foci, enlarged and mottled white spleens, fibrinous exudate in the peritoneum, and atretic, misshapen ovarian follicles. Microscopic lesions included multifocal coagulative necrosis of hepatocytes and inflammation, fibrinous exudation in vascular sinuses of the spleen, and fibrinosuppurative inflammation of the peritoneum and ovarian follicles. The proportion of reproductive organ infections (ovary and oviduct) in the IV group, 83% (20/24, P = 0.007; 50% and 33% for strains A and strain B birds, respectively), was higher than that of the PO group, 46% (11/24; 29% and 17% for strains A and B, respectively), for the first 16 days of observation postinoculation. The proportion of fecal shedding for the IV group of birds was significantly (P = 0.009) lower, 29% (7/24; 33% and 25% respectively for strain A and strain B birds, respectively), than the PO group, 67% (16/24; 75% and 58% for strain A and strain B birds, respectively). Three (2.6%) of 234 egg pools were culture-positive for group D Salmonella from strain A chickens (1 of 119 pools from the IV group and 2 of 115 pools from the PO group of birds). Chickens infected with the field strain of S. enteritidis phage type 4 harbored the organism in tissues only for a brief time, most clearing within 8 DPI and nearly all within 16 DPI. Overall the percentage of culture-positive birds did not differ significantly (P > 0.05) between birds with and without lesions, but isolation of S. enteritidis tended to be more frequent when lesions were evident. This experiment also demonstrated that brown-egg-laying-type chickens were more susceptible than white-egg-laying-type chickens to S. enteritidis phage type 4 isolated from California based on gross and microscopic lesions and bacteriologic findings.     

Detection of airborne Salmonella enteritidis in the environment of experimentally infected laying hens by an electrostatic sampling device

Bacteriologic culturing of environmental samples taken from sources such as manure pits and egg belts has been the principal screening tool in programs for identifying commercial laying flocks that have been exposed to Salmonella enteritidis and are thus at risk to produce contaminated eggs. Because airborne dust and aerosols can carry bacteria, air sampling offers a potentially efficient and inexpensive alternative for detecting S. enteritidis in poultry house environments. In the present study, an electrostatic air sampling device was applied to detect S. enteritidis in a room containing experimentally infected, caged laying hens. After oral inoculation of hens with a phage type 13a S. enteritidis strain, air samples were collected onto agar plates with the electrostatic sampling device, an impaction air sampler, and by passive exposure to the settling of aerosols and dust. Even though the floor of the room was cleaned once per week (removing most manure, dust, and feathers), air samples were positive for S. enteritidis for up to 4 wk postinoculation. On the basis of both the number of S. enteritidis colonies observed on incubated agar plates and the frequency of positive results, the efficiency of the electrostatic device was significantly greater than that of the passive exposure plates (especially at short collection intervals) and was similar to that of the far more expensive impaction sampler. The electrostatic device, used for a 3-hr sampling interval, detected airborne S. enteritidis on 75% of agar plates over the 4 wk of the study.     

Colonization of reproductive organs and internal contamination of eggs after experimental infection of laying hens with Salmonella heidelberg and Salmonella enteritidis

Internal contamination of eggs laid by hens infected with Salmonella enteritidis has been a prominent international public health issue since the mid-1980s. Considerable resources have been committed to detecting and controlling S. enteritidis infections in commercial laying flocks. Recently, the Centers for Disease Control and Prevention also reported a significant association between eggs or egg-containing foods and S. heidelberg infections in humans. The present study sought to determine whether several S. heidelberg isolates obtained from egg-associated human disease outbreaks were able to colonize reproductive tissues and be deposited inside eggs laid by experimentally infected hens in a manner similar to the previously documented behavior of S. enteritidis. In two trials, groups of laying hens were orally inoculated with large doses of four S. heidelberg strains and an S. enteritidis strain that consistently caused egg contamination in previous studies. All five Salmonella strains (of both serotypes) colonized the intestinal tracts and invaded the livers, spleens, ovaries, and oviducts of inoculated hens, with no significant differences observed between the strains for any of these parameters. All four S. heidelberg strains were recovered from the interior liquid contents of eggs laid by infected hens, although at lower frequencies (between 1.1% and 4.5%) than the S. enteritidis strain (7.0%).     

Spiramycin, oxytetracycline and sulphamonomethoxine contents of eggs and egg-forming tissues of laying hens

Spiramycin (SP), oxytetracycline (OTC) or sulphamonomethoxine (SMM) was fed to laying hens at a dietary level of 400 p.p.m. for 7 successive days. After 7 days of medicated feed, the concentrations of SP, OTC and SMM were determined in the blood, liver, ovary, oviducts (magnum and isthmus plus shell gland) and eggs (albumen and yolk) by high-performance liquid chromatography. Of the three drugs, OTC showed the lowest content in the above tissues and eggs, while the reverse was true for SMM. Low concentrations of SP were measured in the blood, whereas contents in the liver and the oviducts were relatively much higher.     

Differences among six Salmonella serovars in abilities to colonize reproductive organs and to contaminate eggs in laying hens

The abilities of Salmonella serovars to colonize the reproductive organs of chickens and to contaminate eggs were compared. Mature laying hens were inoculated intravenously with 10(5) colony-forming units of Salmonella enteritidis, Salmonella typhimurium, Salmonella infantis, Salmonella hadar, Salmonella heidelberg, or Salmonella montevideo to cause the systemic infection. Salmonella enteritidis was recovered from three yolks of the laid eggs (7.0%), suggesting egg contamination from the transovarian transmission of S. enteritidis. The liver, spleen, and cecum were colonized by each serovar similarly at 4 or 7 days postinoculation (PI), whereas the ovary and preovulatory follicles were colonized by S. enteritidis with significantly (P < 0.05) higher levels than by the other serovars at 4 and 7 days PI. Salmonella enteritidis was recovered from the cloaca and vagina at 2, 4, and 7 days PI and from the other portions of the oviduct at 4 and 7 days PI. In addition, S. enteritidis had been persistent in the peripheral blood for 7 days PI. These results suggest that S. enteritidis is the predominant serovar to colonize the reproductive organs of mature laying hens among six serovars used in this study, reflecting the field situatibn in which the predominant outbreaks of human salmonellosis were caused by S. enteritidis-contaminated eggs recently. The ability of S. enteritidis to colonize the reproductive organs may be one of the reasons that egg contamination with S. enteritidis has increased.     

Evaluation of a French ELISA for the detection of Salmonella Enteritidis and Salmonella Typhimurium in flocks of laying and breeding hens

In France, the regular and compulsory detection of Salmonella Enteritidis (SE) and Salmonella Typhimurium (ST) in flocks of breeding and laying hens is based on bacteriological examination of environmental swabs and faeces samples. The aim of this study was to compare this bacteriological examination with a serological method (ELISA) developed in our laboratory. This ELISA was first evaluated by use of artificially infected hens. During these experimental infection studies, several groups of hens were inoculated with SE, ST, different vaccines and different Salmonella serovars to calculate the experimental parameters of our ELISA. Then, in a field study, 43 flocks were followed monthly using two bacteriological samples (environmental swab and pool of faeces) and 20 serological samples (sera or yolks). Twenty-seven flocks without SE or ST gave a negative serological response throughout their surveillance. Among the 10 various serovars different from SE and ST isolated in this study, S. Heidelberg, S. Agona and S. Hadar gave seropositive results in seven flocks. Consequently, this ELISA was not specific of SE and ST as it detected serovars sharing or not common antigens with SE and ST. Seropositive results were also obtained each month for two flocks where no Salmonella could be isolated. Finally, in seven flocks found infected with SE or ST, the positive ELISA results appeared later than the bacteriological detection. Therefore, for the detection of chicken flocks recently infected with SE or ST, bacteriological examination currently used in France seems to be more appropriate than this ELISA.     

Characteristics of Salmonella enteritidis contamination in eggs after oral,aerosol, and intravenous inoculation of laying hens

Experimental infection models are useful tools for understanding how Salmonella enteritidis is deposited in eggs and for testing potential strategies to control eggborne transmission of disease to humans. Oral inoculation of laying hens is presumed to provide the closest simulation of naturally occurring infections, but alternatives such as intravenous or aerosol inoculation have sometimes been recommended as options to induce higher incidences of egg contamination. The present study compared the frequency, level, and location of S. enteritidis deposition in egg contents after experimental inoculation by three different routes. In two replicate trials, specific-pathogen-free laying hens were infected with an S. enteritidis culture mixture prepared to optimize invasive behavior. Groups of hens received either an oral dose of 10(9) S. enteritidis, an aerosol dose of 10(9) S. enteritidis, or an intravenous dose of 10(5)-10(7) S. enteritidis. Oral inoculation led to the highest incidence of fecal shedding of S. enteritidis, whereas intravenous inoculation produced the highest specific antibody titers. Eggs laid during the first 21 days postinoculation were cultured to detect and enumerate S. enteritidis in the yolk and albumen. No significant differences were observed among the three inoculation routes in the frequencies of isolation of S. enteritidis from either yolk or albumen. For all three routes of administration, S. enteritidis was recovered more often from yolk (at frequencies ranging from 4% to 7%) than from albumen (0 to 2%). Over 73% of contaminated eggs harbored fewer than 1 colony-forming unit (CFU) of S. enteritidis per milliliter, and only 3% of such eggs contained more than 100 CFUs/ml. Significantly higher levels of S. enteritidis contaminants were associated with intravenous inoculation than with the other routes. No advantage of using aerosol or intravenous administration of S. enteritidis as an alternative to oral inoculation for inducing the production of contaminated eggs was evident in this study.     

Deposition of phage type 4 and 13a Salmonella enteritidis strains in the yolk and albumen of eggs laid by experimentally infected hens

Because egg yolk and albumen differ substantially in their abilities to support bacterial growth, the initial level and location of Salmonella enteritidis deposition are critical for determining whether proposed standards for refrigerating eggs are likely to protect public health by preventing extensive microbial multiplication. In the present study, three groups of laying hens were infected with oral doses of approximately 10(9) cells of different S. enteritidis strains (two were phage type 4 and one was phage type 13a) in two replicate trials. For all three S. enteritidis strains, the incidence of yolk contamination (approximately 2.5% overall) was significantly greater than the incidence of albumen contamination (approximately 0.5% overall). The phage type 13a strain was less often isolated from fecal samples at 2 wk post-inoculation than were the phage type 4 strains, but no significant differences between strains were observed in the incidence of egg contamination. Most freshly laid contaminated eggs contained fewer than 1 S. enteritidis cell/ml of egg yolk or albumen, and no sample contained more than 67 S. enteritidis cells/ml.     

Secretion of Salmonella-specific antibodies in the oviducts of hens experimentally infected with Salmonella enteritidis

The production and secretion of Salmonella enteritidis whole cell antigen-specific antibodies in the oviducts and in the serum of laying hens experimentally infected with Salmonella enteritidis, was analyzed by ELISA. The dynamics of the antibody levels in the oviducts were identical to that in the serum. Subclasses of antibodies (IgA, IgG, and IgM) in the infected hens were found to increase significantly (p < 0.01) compared to those in the control uninfected hens throughout the experiment. IgG and IgM levels in both oviducts and in sera reached to a peak by 14 days post-inoculation, and remained elevated throughout. The secretion of IgA seemed to be transient since the IgA levels increased to a peak 7 days after both primary and secondary inoculations, and declined rapidly. The elevated levels of antibodies were followed by partial clearance of Salmonella organisms from the oviducts. The present results indicate a significant local immune reaction against the Salmonella infection and suggest an association of the local antibodies with the clearance of Salmonella from the oviducts at least partially.     

Impacts of dried whole eggs on productive performance, quality of fresh and stored eggs, reproductive organs and lipid metabolism of laying hens

1. Rejected eggs were processed by freezing, freezing and boiling, or autoclaving, dried at 55°C, and the chemical and amino-acid analysis and microbiological properties of the dried whole eggs (DWE) determined. The processed DWE was included in a laying-hen mash diet at 50 or 100 g/kg, mainly to replace soybean meal and maize, in a factorial design of 3×2 and fed to hens between 25 and 41 weeks of age. In addition, a control diet without DWE was used to create a total of 7 treatments; each having 5 replicates of 5 hens. 2. Hens fed the diet containing 50-g/kg frozen and boiled DWE and 100-g/kg frozen DWE laid the largest egg size, while hens fed the 50-g/kg frozen and boiled or autoclaved DWE diets yielded the highest egg mass and best feed conversion efficiency. 3. Hens fed the 50-g/kg frozen DWE laid eggs with a higher yolk indices, Haugh unit scores, and eggshell thickness and maintained higher Haugh unit scores during egg storage. 4. Yolk cholesterol significantly decreased in eggs from groups fed either the 50-g/kg frozen or autoclaved DWE diet, with freezing having the stronger effect. 6. In conclusion, 50-g/kg autoclaved DWE diets fed to laying hens between 25 and 41 weeks of age had no adverse effects on production, egg quality or reproductive organs, but decreased yolk cholesterol, and thus could be beneficially included in laying-hen diets.     

Lead concentrations in the blood and eggs of backyard laying hens

AIMS: To investigate the prevalence of lead exposure in hens and eggs from backyard poultry in a sample of Auckland households, the relationship between concentrations of lead in the blood of the hens and in the shells and yolks of eggs from the same household, and to examine associations with measures of hen health, environment and husbandry factors.

METHODS: Thirty households participated in the study from August to November 2016, each providing one adult hen for sampling, an egg from the household if available, and completing a questionnaire on hen husbandry. Concentrations of lead in blood were determined using a portable lead analyser. Eggs were analysed for concentrations of lead in the yolk and shell using inductively coupled plasma mass spectrometry after biological digestion with a mixture of nitric and hydrochloric acid.

RESULTS: Twenty three of 30 hens (77%) showed evidence of lead exposure, with median concentrations of lead in blood of 0.77 (min?<0.16, max 8.02) μmol/L. All eggs showed evidence of lead exposure, with concentrations of lead in the yolk ranging from 0.003–1.07?mg/kg, and concentrations of lead in the eggshell ranging from <0.1–0.82?mg/kg. A positive correlation existed between concentrations of lead in the blood of a hen and concentrations of lead in egg yolk from the same hen (R²=0.97), and both the yolk (R²=0.58) and shell (R²=0.30) of an egg from her flock. No association was found between concentrations of lead in blood and hen health indices measured in this study. Concentrations of lead in blood were higher in hens from properties with homes built before 1941 than between 1941–1960 (p=0.03), and in hens from properties with weatherboard homes than brick homes (p=0.049).

CONCLUSIONS AND CLINICAL RELEVANCE: There was a high prevalence of lead exposure in this sample of Auckland backyard chickens, with the majority of hens being sub-clinically affected. Associations were found between concentrations of lead in the blood of the hens, and properties with homes built before 1941 and clad in weatherboard. Concentrations of lead in over half the egg yolks sampled were at levels sufficient to warrant human health concern. The assessment of concentrations of lead in backyard poultry and eggs intended for human consumption is recommended to protect human and bird health.     

The absence of a relationship between egg production and dominance in caged laying hens

1. The individual egg production of two strains of laying hen in colony battery cages was recorded from 20 to 72 weeks of age by feeding a different fat‐soluble dyestuff to each bird and then breaking open the eggs to determine the yolk colour and hence the identity of the hen that laid the egg.

2. Observations of agonistic behaviour and other pecking were made and the combs were examined for pecking damage on two occasions; each group of hens was hierarchically arranged in terms of these three measures, which were then taken as an indication of each hen's dominance.

3. The three measures of dominance correlated fairly well with each other, but none showed any relationship with egg production.     

The relationship between the magnitude of the specific antibody response to experimental salmonella enteritidis infection in laying hens and their production of contaminated eggs.

Detecting infected laying flocks is a vital part of many efforts to control egg-associated transmission of Salmonella enteritidis to humans. The relationship between the development of a specific antibody response in infected hens and the deposition of S. enteritidis in eggs is important for establishing the epidemiologic relevance of serologic testing methods. In two trials, laying hens were infected with large oral doses of phage types 13a and 14b isolates of S. enteritidis. Approximately 38% of all infected hens produced at least one contaminated egg, at an overall incidence of 5.2%, between 3 and 23 days postinoculation. As determined by enzyme-linked immunosorbent assay with an S. enteritidis flagellar antigen, 91.7% of inoculated hens produced specific serum antibodies. Although hens with very high antibody titers were associated with a significantly elevated frequency of egg contamination, a consistently direct relationship was not evident between the magnitude of the antibody responses of individual hens and the frequency at which they laid contaminated eggs. Accordingly, although serologic tests can be valuable screening tools for preliminary detection of S. enteritidis infections in poultry, the magnitude of the antibody responses detected in individual hens may not predict the overall risk of egg contamination associated with particular laying flocks.