Inactivation of Salmonella enteritidis during boiling of eggs

A series of inactivation curves for Salmonella enteritidis were determined for boiling eggs using different conditions of time and temperature. No significant influence of egg weight could be found on the temperature evolution in the yolk. The inactivation curves consistently showed an initial slow decline in bacterial number at lower temperatures, after which a very rapid inactivation took place. It was not possible to reproduce this behavior using a traditional inactivation model. A pragmatic model existing in two parts was therefore constructed. When the temperature is below a certain threshold, the inactivation follows a second order temperature dependence. Above the temperature threshold, standard Bigelow inactivation kinetics are assumed.This model could describe the data reasonably well, provided that the decimal reduction time in the Bigelow model was assumed to be different for a fast or slow heating process, respectively. The results suggest that the bacteria are more resistant towards a slower heating process, which is confirmed by analyzing the raw data. A fail-safe model can be obtained by using the parameters associated with the slow heating process. The statistical properties of the calibrated model are satisfactory, and a cross-validation shows that it can be used for egg boiling conditions outside its calibration range.     

Factors influencing inactivation of Salmonella enteritidis in hard-cooked eggs

The inside of a hen's egg, once considered sterile, is now known to occasionally harbor Salmonella Enteritidis. At least two recent outbreaks of salmonellosis in which Salmonella Enteritidis PT34 was involved have been associated with hard-cooked eggs. This study was undertaken to compare D56 degrees C values of Salmonella Senftenberg 775W and six strains of Salmonella Enteritidis isolated from outbreaks associated with eggs. D56 degrees C values for Salmonella Enteritidis in liquid egg yolk ranged from 5.14 to 7.39 min; the D56 degrees C value for Salmonella Senftenberg was 19.96 min. The two PT34 strains from outbreaks associated with hard-cooked eggs did not exhibit significantly higher resistance to heat compared with two PT4 strains and one strain each of PT8 and PT13a. A PT4 strain and a PT34 strain of Salmonella Enteritidis were separately inoculated (10(7) to 10(8) CFU) into the yolk of medium and extra large shell eggs at 10 and 21 degrees C, and survival was monitored using two cooking methods: (i) placing eggs in water at 23 degrees C, heating to 100 degrees C, removing from heat, and holding for 15 min (American Egg Board method) and (ii) placing eggs in water at 100 degrees C, then holding for 15 min at this temperature. Within the 15-min holding periods, inactivation was more rapid using the method recommended by the American Egg Board compared with method 2. Within each cooking method, inactivation was most rapid in medium eggs initially at 21 degrees C. The PT4 strain survived in yolk of extra large eggs initially at 10 degrees C when eggs were held in boiling water 9 min using method 2. The final temperature of the yolk in these eggs was 62.3 +/- 2 degrees C. Of the two methods evaluated for hard cooking eggs, the American Egg Board method is clearly most effective in killing Salmonella Enteritidis in the yolk.     

Isolation of antibodies against soy protein from the eggs of immunized laying hens

The authors report on their experiences with immunization of laying hens with alpha-, alpha'- and beta-conglycinin, and on the isolation of specific antibodies from the yolks. Activity and specificity of the immunoglobulins were tested by an enzyme-linked immuno sorbent assay (ELISA). This method of antibody production proved to be very convenient, especially in regard to the high yield of antibodies and the easy handling of the animals. The necessity of using specific antisera for quantitative immunoassays is pointed out.     

Dynamic predictive model for growth of Salmonella enteritidis in egg yolk

ABSTRACT:  Salmonella Enteritidis (SE) contamination of poultry eggs is a major human health concern worldwide. The risk of SE from shell eggs can be significantly reduced through rapid cooling of eggs after they are laid and their storage under safe temperature conditions. Predictive models for the growth of SE in egg yolk under varying ambient temperature conditions (dynamic) were developed. The growth of SE in egg yolk under several isothermal conditions (10, 15, 20, 25, 30, 35, 37, 39, 41, and 43 °C) was determined. The Baranyi model, a primary model, was fitted with growth data for each temperature and corresponding maximum specific growth rates were estimated. Root mean squared error (RMSE) values were less than 0.44 log₁₀ CFU/g and pseudo- R ² values were greater than 0.98 for the primary model fitting. For developing the secondary model, the estimated maximum specific growth rates were then modeled as a function of temperature using the modified Ratkowsky's equation. The RMSE and pseudo- R ² were 0.05/h and 0.99, respectively. A dynamic model was developed by integrating the primary and secondary models and solving it numerically using the 4th-order Runge–Kutta method to predict the growth of SE in egg yolk under varying temperature conditions. The integrated dynamic model was then validated with 4 temperature profiles (varying) such as linear heating, exponential heating, exponential cooling, and sinusoidal temperatures. The predicted values agreed well with the observed growth data with RMSE values less than 0.29 log₁₀ CFU/g. The developed dynamic model can predict the growth SE in egg yolk under varying temperature profiles.     

Simple and rapid methods for detecting Salmonella enteritidis in raw eggs

The Centers for Disease Control and Prevention estimates there were 300,000 cases of Salmonella enteritidis (SE) in 1997. Egg products were associated with many of the cases. To address this problem, many producers implemented flock surveillance of the SE situation at their facilities. A rapid and simple method for detecting SE from poultry samples is critical for the effective implementation of such testing strategies. A lateral flow device for the detection of SE utilized in this study was manufactured by Neogen, Lansing, MI. The test panel is a presumptive qualitative test system that detects only members of Group D1 Salmonella species. A series of studies were conducted to optimize the test procedure for raw eggs with different sample preparations. A novel antigen extraction method was developed for use with the test panel kit. The detection limit of the test panel kit was increased approximately tenfold when the extraction method was used. Detection of SE was 100% in raw egg pools inoculated with 10 SE cells per ml of egg and incubated at a 1:10 ratio in buffered peptone water (BPW) or tetrathionate brilliant green broth (TBG) for 24 h at 37 degrees C. The developed lateral flow test kit could provide a simple, rapid, and inexpensive method for egg producers and processors to test specifically for Salmonella group D1 serovars, such as SE, in egg samples.     

Growth of Salmonella enteritidis in artificially contaminated eggs: the effects of inoculum size and suspending media.

Growth profiles of two isolates of Salmonella enteritidis phage type (PT) 4 inoculated into either the albumen of whole shell eggs or into separated albumen were found to be markedly affected by the size of the inoculum and the composition of the medium used to suspend the cells prior to inoculation. Using our model with an inoculum of two cells, multiplication of the Salmonella was not seen in 93% of eggs held at 20 degrees C for 8 days. In approximately 7% of eggs, however, growth occurred during the 8 days of storage. If the inoculum equaled or exceeded 25 cells per egg when eggs were subsequently stored at 20 degrees C, or 250 cells per egg when eggs were stored at 30 degrees C, high levels of growth of Salmonella in the egg occurred significantly more frequently than when the inoculum was two cells. High levels of growth were also seen more frequently if the inoculum was suspended in buffered peptone water or maximal recovery diluent rather than in phosphate buffered saline. Growth of Salmonella in separated albumen occurred very infrequently (1.1% of samples) at low inoculum levels and did not become significant until the inoculum was 250 cells or greater. Growth in the albumen was unaffected by the composition of the suspending medium. Provided that the inoculum was approximately 2 cells per egg and the bacteria were suspended in PBS, observed growth profiles of S. enteritidis inoculated into the albumen of whole eggs resembled those in naturally contaminated eggs.     

Individual-based modelling of growth and migration of Salmonella enteritidis in hens' eggs

An individual-based model (IbM) was developed to describe the growth and migration of Salmonella enteritidis in hens' eggs. The Bacteria Simulator (BacSim) environment was used to implement the model; the bacteria are represented by spheres that grow by nutrient uptake and divide in two daughter cells upon exceeding a certain threshold volume. Motility of the Salmonella bacteria was described by a run and tumble mechanism. For the sake of simplicity, the bacteria were assumed to grow exponentially, an appropriate assumption for the initial phase of growth relevant for shelf-life predictions. Both albumen and yolk were assumed to be homogeneous. The impact of several model parameters (chemotaxis, growth rate, initial contamination numbers and bacterial swimming speed) was assessed by a sensitivity analysis. The results show that chemotaxis towards the yolk would have a strong effect on the time needed to reach the vitelline membrane, an aspect that future research should focus on. The contamination position had less impact on the time to reach the vitelline membrane. The simulation results illustrate the need for more detailed knowledge on the subject of bacterial migration in hens' eggs. Our model can easily incorporate this knowledge when it becomes available.     

Effects of different molting procedures on incidence of Salmonella infection in flocks of naturally contaminated laying hens in a commercial egg-producing farm by detection of yolk antibodies to Salmonella in eggs

Indirect enzyme-linked immunosorbent assays (ELISAs) have been applied to detect immunoglobulin Y antibodies to different serotypes of Salmonella in the yolks of chicken eggs with heat-extracted antigens of Salmonella enterica serotypes Agona (SA), Cerro (SC), Enteritidis (SE), Montevideo (SM), and Putten (SP). The egg yolk samples examined were classified as positive if their ELISA absorbance values exceeded the value for eggs from specific-pathogen-free flocks by more than two standard deviations. Of 30 egg yolk samples from three flocks vaccinated with a killed SE vaccine, 29 were antibody positive by the ELISA assay for the SE antigen. Four to 29 of the 29 yolk samples showed positive results for the other serovars, although the absorbance values for SE were higher than those obtained for the other serotypes in each of the yolk samples. All 30 yolks from three flocks that were not administered any SE vaccines were found to be antibody negative for SE, and two samples were determined to be positive for SC. Thirty-nine or 40 eggs were obtained from each of four layer flocks in a commercial egg production farm where the laying houses were naturally contaminated with SA, SC, SM, SP, Salmonella serovar Infantis (SI), and untypeable strains. The ELISA absorbance values for SM in the egg yolks obtained from the two flocks molted through feed withdrawal when the birds restarted laying were significantly (P < 0.05) higher than those observed in the yolks obtained before the molt. In egg yolks from the two other flocks that were molted through a wheat bran diet, there was no significant difference between the absorbance values before and after the molt. The observations in the present study provide further evidence to suggest that a molt initiated through the administration of a wheat bran diet can reduce the risk for Salmonella problems in a commercial egg-producing setting.     

Provision of lactose to molting hens enhances resistance to Salmonella enteritidis colonization.

Older leghorn hens, more than 50 weeks of age, were divided into three groups designated 1, unmolted controls; 2, molted; or 3, molted treated with lactose. Forced molt was induced by 14 days of feed removal. Lactose was provided to the hens in group 3 as 2.5% (wt/vol) of the daily drinking water. Each hen in all groups was challenged orally with 10(5) Salmonella enteritidis (SE) cells on day 7 of feed removal. The study was repeated in three replicated trials. The concentrations of acetic, propionic, and total volatile fatty acids (VFA) in the cecal contents of the molted hens in groups 2 and 3 decreased significantly (P < 0.05) on days 6 and 14 of molt compared with the unmolted controls. Forced molt had no apparent effect on pH or on the oxidation-reduction potential of the ceca. Compared to the unmolted controls, SE cecal and spleen and liver colonization was significantly increased (P < 0.05) in the molted hens in group 2. Compared to the molted hens in group 2, SE cecal and spleen and liver colonization was significantly decreased (P < 0.05) in two of three trials in the hens in group 3 provided with lactose. The results suggested that the increased susceptibility of molting hens to SE colonization may be associated with decreased fermentation and production of VFA by cecal bacteria or by a depletion of the number of VFA-producing bacteria present in the ceca. The results further suggest that providing lactose in the drinking water during molting may significantly enhance resistance to SE colonization.     

不同温度下肠炎沙门氏菌在鸡蛋清和蛋黄中的生存特点

目的比较不同温度下肠炎沙门氏菌(SE)在蛋黄、蛋清中的生存能力。方法无菌分离蛋黄蛋清,将SE稀释液分别接种于蛋黄、蛋清中,使SE-蛋清/蛋黄混合液中SE的浓度约为103CFU/mL。不同温度下培养,每4h/8h用平板计数法测定SE-蛋清/蛋黄混合液中菌液浓度,据此作出SE浓度变化曲线。结果 4℃时SE缓慢增殖,20℃和37℃时SE快速增殖,39℃时蛋清成分开始发挥有效的抑菌作用,42℃时蛋清中的SE能够被快速清除。结论严格控制污染源,并低温保存鸡蛋以限制鸡蛋中SE含量的快速增加。     

Detection of Salmonella enteritidis in shell and liquid eggs using enrichment and plating

Detection methods using various enrichment and plating media and immunoconcentration for Salmonella enteritidis in shell and liquid eggs were evaluated. For liquid egg samples naturally contaminated with S. enteritidis, pre-enrichment in 225 ml of buffered peptone water with cysteine followed by selective enrichment in 10 ml of tetrathionate broth was the superior, resulting in the detection of S. enteritidis in all samples on six of the seven types of selective agar substrate investigated. This enrichment procedure also enabled detection of S. enteritidis in most of artificially inoculated shell egg and pasteurized liquid egg samples.     

Correlation of eggshell strength and Salmonella enteritidis contamination of commercial shell eggs

Shell quality has been identified as a heritable trait that can be manipulated by genetic selection. Previous research has concluded that many methods of determining shell quality produce variable results. With the development of newer, more precise measuring technologies, shell strength can now be assessed in a consistent, objective fashion. A research project was conducted to determine what role shell strength might play in affecting external Salmonella Enteritidis contamination of egg contents. Visibly clean eggs were collected from an in-line shell egg-processing facility at the accumulator. Eggs were inoculated by dipping in a concentrated suspension of nalidixic acid-resistant Salmonella Enteritidis. After storage, eggs were assessed for shell strength and both external and internal Salmonella Enteritidis contamination. In the first study, there was a significant difference (P < 0.05) in shell strength among the three replicates. No differences between treatments were found for shell strength or Salmonella Enteritidis contamination of contents. In the second study, there were no replicate differences for any of the monitored factors. When rinsate and content samples were enriched, 100% of the rinsates were positive for Salmonella Enteritidis. No content samples were shown to be contaminated with Salmonella Enteritidis during direct plating, but 3 to 5% of the samples from each replicate were positive after enrichment. Correlation analysis of the results from each study found only weak correlations between shell strength and Salmonella Enteritidis contamination on eggshell surface or contents. Within the range of shell strengths recorded in this study, the correlation analysis suggests that shell strength does not play a major role in Salmonella Enteritidis contamination. Further work with eggs that represent a greater range of shell strengths could provide a clearer indication of the interaction of shell strength and Salmonella Enteritidis contamination.     

High voltage atmospheric cold plasma decontamination of Salmonella enteritidis on chicken eggs

Salmonella enteritidis (SE) accounts for more than 70% of Salmonella spp. infections in humans with a primary source being chicken eggs, that can result from post-lay SE cross-contamination of the shell from contaminated equipment or the environment. The objective of this study was to apply a HVACP treatment that can achieve a minimum 5-log reduction in SE on the surface of artificially inoculated shell eggs with an initial bacterial load of 10⁸ CFU/egg, after a previous disinfection. Optimized HVACP treatment conditions were an indirect treatment with air at 60% humidity at 100 kV for one minute treatment and six hours post-treatment or alternatively, five minutes of treatment and four hours post-treatment. Egg quality parameters of Haugh unit (HU), pH, color, and vitelline membrane and shell strength were tested under the optimized conditions and showed no significant difference (p > 0.05) between treated and untreated eggs.Industrial relevance: Missing information for a possible scale up of a cold plasma system for egg surface decontamination has been addressed by an optimization of HVACP treatment focused on treatment and post-treatment time, essential parameters to have into account in the food industry. These results demonstrate that HVACP is an effective decontamination method for SE on chicken shell eggs and provides a baseline for a future scale up of the process, showing that different combinations of treatment variables can achieve the desired decontamination without affecting to key quality parameters of the egg such as Haugh Unit or vitelline membrane strength.     

Effect of refrigeration on in vitro penetration of Salmonella enteritidis through the egg yolk membrane

Internally contaminated eggs have been implicated as leading sources of transmission of Salmonella Enteritidis (SE) to humans. Although SE is not often deposited inside the nutrient-rich yolks of naturally contaminated eggs, penetration through the vitelline membrane to reach the yolk contents could result in rapid bacterial multiplication. In previous studies, such penetration has been observed occasionally at warm temperatures during experiments with in vitro egg contamination models. The present study was conducted to determine whether refrigeration affects the frequency of in vitro SE penetration of the egg yolk membrane. After inoculation of small numbers of SE onto the outside of the vitelline membranes of intact yolks, immediate refrigeration of contaminated samples prevented the penetration of SE into the egg yolk contents during 24 h of storage. However, SE penetrated inside the yolk contents in 4% of contaminated egg samples refrigerated after 2 h of storage at 30 degrees C, 15% of samples refrigerated after 6 h of storage at 30 degrees C, and 40% of samples stored at 30 degrees C for 24 h (48 samples per treatment group). These results highlight the value of prompt refrigeration for restricting the opportunities for SE to multiply to high numbers inside the yolks of contaminated eggs.     

Estimating the annual fraction of eggs contaminated with Salmonella enteritidis in the United States

Using available data on the occurrence of Salmonella enteritidis (SE) in US layer flocks and eggs, and a probabilistic scenario tree method, an estimate of the fraction of SE-contaminated eggs produced annually is derived with attendant uncertainty. In lieu of a definitive prevalence survey, the approach presented here provides insight to the relative contribution of various pathways leading to contaminated eggs. A Monte Carlo model with four branches is developed. The first branch predicts the proportion of all US flocks that are SE-affected. The second branch apportions SE-affected flocks into three categories (high, moderate, and low level affected flocks) based on population-adjusted epidemiologic data. The third branch predicts the proportion of affected flocks that are molted and producing eggs during a high risk period subsequent to molt. The fourth branch predicts the fraction of contaminated eggs produced by flocks of the type described by the pathway (e.g. high level affected flocks that are not molted) based on egg sampling evidence from naturally infected flocks. The model is simulated to account for uncertainty in the data used to estimate the branch probabilities. Correlation analysis is used to estimate the sensitivity of model output to various model inputs. The output of this model is an uncertainty distribution for the fraction of all eggs that are SE-contaminated during 1 year of production in the US. The expected value of this distribution is approximately one SE-affected egg in every 20,000 eggs annually produced, and the 90% certainty interval is between one SE-contaminated egg in 30,000 eggs, and one SE-contaminated egg in 12,000 eggs. The model estimates that an average of 14% of all eggs (i.e. contaminated and not contaminated) from affected flocks are produced by high level, non-molted affected flocks, but these flocks are estimated to account for more than two-thirds of the total fraction of contaminated eggs produced annually. Sensitivity analysis also suggests that the proportion of affected flocks that are high level flocks - and the egg contamination frequency for these types of flocks - are the most sensitive model inputs. The model's pathways provide a framework for evaluating interventions to reduce the number of contaminated eggs produced in the US. Furthermore, sensitivity analysis of the model identifies those inputs whose uncertainty is most influential on the model's output. Future farm-level research priorities can be established on the basis of this analysis, but public policy decisions require a fuller exposure assessment and dose-response analysis to account for microbial growth dynamics, meal preparation, and consumption demographics among US egg consumers.     

Efficacy of electrolyzed water in inactivating Salmonella enteritidis and Listeria monocytogenes on shell eggs

The efficacy of acidic electrolyzed (EO) water produced at three levels of total available chlorine (16, 41, and 77 mg/ liter) and chlorinated water with 45 and 200 mg/liter of residual chlorine was investigated for inactivating Salmonella Enteritidis and Listeria monocytogenes on shell eggs. An increasing reduction in Listeria population was observed with increasing chlorine concentration from 16 to 77 mg/liter and treatment time from 1 to 5 min, resulting in a maximal reduction of 3.70 log CFU per shell egg compared with a deionized water wash for 5 min. There was no significant difference in antibacterial activities against Salmonella and Listeria at the same treatment time between 45 mg/liter of chlorinated water and 14-A acidic EO water treatment (P > or = 0.05). Chlorinated water (200 mg/liter) wash for 3 and 5 min was the most effective treatment; it reduced mean populations of Listeria and Salmonella on inoculated eggs by 4.89 and 3.83 log CFU/shell egg, respectively. However, reductions (log CFU/shell egg) of Listeria (4.39) and Salmonella (3.66) by 1-min alkaline EO water treatment followed by another 1 min of 14-A acidic EO water (41 mg/liter chlorine) treatment had a similar reduction to the 1-min 200 mg/liter chlorinated water treatment for Listeria (4.01) and Salmonella (3.81). This study demonstrated that a combination of alkaline and acidic EO water wash is equivalent to 200 mg/liter of chlorinated water wash for reducing populations of Salmonella Enteritidis and L. monocytogenes on shell eggs.     

Sensory evaluation of egg products and eggs laid from hens fed diets with different fatty acid composition and supplemented with antioxidants

Is the sensory quality of eggs influenced by adding vegetable lipids, animal and vegetable sources of n − 3 polyunsaturated fatty acids (n − 3 PUFA), and/or natural antioxidants to the hens diet? To answer this question three feeding experiments were conducted adding either palm butter, grape seed oil, flax seed oil, n − 3 PUFA such as flax seed and marine algae and the natural antioxidant rosemary to the hens diet. For each experiment a standard diet was used as control. The results suggested that vegetable lipids (palm butter, grape seed, flax seed), n − 3 PUFA (flax seed and marine algae) and rosemary may be used to hens fed diet without affecting the sensory properties of eggs.The sensory quality of eggs was evaluated on hard boiled, scrambled eggs and Madeira cake.In this work, we report the first sensory characterization of eggs and products containing eggs obtained from hens diet based on grape and algae plus vitamin E and rosemary extract.     

Heat transfer models for predicting Salmonella enteritidis in shell eggs through supply chain distribution

ABSTRACT:  Egg and egg preparations are important vehicles for Salmonella enteritidis infections. The influence of time–temperature becomes important when the presence of this organism is found in commercial shell eggs. A computer-aided mathematical model was validated to estimate surface and interior temperature of shell eggs under variable ambient and refrigerated storage temperature. A risk assessment of S. enteritidis based on the use of this model, coupled with S. enteritidis kinetics, has already been reported in a companion paper published earlier in JFS . The model considered the actual geometry and composition of shell eggs and was solved by numerical techniques (finite differences and finite elements). Parameters of interest such as local ( h ) and global ( U ) heat transfer coefficient, thermal conductivity, and apparent volumetric specific heat were estimated by an inverse procedure from experimental temperature measurement. In order to assess the error in predicting microbial population growth, theoretical and experimental temperatures were applied to a S. enteritidis growth model taken from the literature. Errors between values of microbial population growth calculated from model predicted compared with experimentally measured temperatures were satisfactorily low: 1.1% and 0.8% for the finite difference and finite element model, respectively.     

Sensitivity analysis of Salmonella enteritidis levels in contaminated shell eggs using a biphasic growth model

Salmonella enteritidis (SE) is a common foodbome pathogen, the transmission of which is primarily associated with the consumption of contaminated Grade A shell eggs. In order to estimate the level of SE present in raw shell eggs, it is necessary to consider the protective effects of the egg albumin, which effectively inhibits SE growth in a time- and temperature-dependent manner. In this study, a SE growth model was produced by combining two mathematical equations that described both the extended lag phase of SE growth (food component) and a SE growth model (pathogen component). This biphasic growth model was then applied to various egg handling scenarios based on the farm-to-table continuum, including in-line and off-line processing facilities with consideration of key events in production, processing, transportation, and storage. Seasonal effects were also studied. Monte Carlo simulation was used to characterize variability in temperature and time parameter values influencing the level of SE to which individuals are exposed. The total level of SE consumed was estimated under best, most likely, and time-temperature abusive handling scenarios. The model estimated that, in most cases, there was no SE growth in contaminated eggs handled under most likely practices, because 10-70% of the yolk membrane remained intact. Under abusive handling scenarios, complete loss of yolk membrane integrity frequently occurred by the time eggs reach the distribution phase, followed by subsequent SE growth, which was often quite rapid. In general, the effect of season and processing method (in-line vs. off-line) was minimal. Further sensitivity analysis demonstrated that the initial SE contamination level significantly influenced the final exposure levels only under no-abuse or mildly abusive conditions. The results of our study suggest that, for maximum reduction of SE exposure level, cooling strategies should not only focus on the on-farm or processing phases, but should emphasize the importance of cooling strategies at the distribution and consumer phases of the farm-to-fork continuum.     

Growth of Salmonella enterica serovar Enteritidis in albumen and yolk contents of eggs inoculated with this organism onto the vitelline membrane

By using an in vitro model simulating the potential opportunities for Salmonella enterica serovar Enteritidis (SE) to proliferate within eggs contaminated with this organism following oviposition, we investigated growth of SE in eggs. Seventy to 140 CFU of one of three SE strains originating either from egg contents, chicken meat, or a human infection were experimentally inoculated onto the vitelline membrane of eggs collected from specific-pathogen-free flocks of chickens and incubated at 25 degrees C. SE organisms were detected in 6 of 71 yolk contents of the eggs inoculated with any of the test strains attaining levels ranging from 2.0 x 10(2) to 4.2 x 10(8) CFU/ml by day 6. The organisms were also detected in the albumen from 38 of 55 eggs tested, growing to levels ranging from 1.0 x 10(2) to 4.3 x 10(8) CFU/ml by day 6 after inoculation. An additional three yolk contents and 15 albumen samples were culture positive for SE following enrichment. There was no correlation between the number of the organisms in the yolk contents and that in the albumen from each of the eggs. When 73 to 91 CFU of the egg strain were inoculated into samples of separated albumen obtained from eggs that were stored at 4 degrees C for 1 to 4 weeks or at 25 degrees C for 1 week, slight growth (3.0 x 10(2) to 7.4 x 10(3) CFU/ml) was found in only 3 of the 60 albumen samples by day 6 after inoculation, but the organisms were recovered from 52 samples following enrichment. The results suggest that the environment on or near the vitelline membrane can be conducive to SE proliferation over time.