Multiplication of Salmonella enteritidis on the yolk membrane and penetration to the yolk contents at 30 degrees C in an in vitro egg contamination model

Refrigeration to limit bacterial multiplication is a critical aspect of efforts to control the transmission of Salmonella enterica serovar Enteritidis (SE) to consumers of contaminated eggs. Although the nutrient-rich yolk interior is an uncommon location for SE contamination in freshly laid, naturally contaminated eggs, migration across the vitelline membrane could lead to rapid bacterial multiplication even when the initial site of deposition is outside the yolk. Multiplication on the yolk membrane (before, or in addition to, multiplication within the yolk contents) could be another source of increased risk to consumers. The present study used an in vitro egg contamination model to compare the abilities of four strains of SE to either multiply in association with the yolk membrane or migrate through that membrane to reach the yolk contents during 36 h of incubation at 30 degrees C. After inoculation onto the exterior surface of intact, whole yolks, all four SE strains penetrated the vitelline membrane to reach the yolk contents (at an overall frequency of 11.5%) after 12 h of incubation. The mean log concentration of SE was significantly higher in whole yolks (including yolk membranes) than in yolk contents at both 12 h (0.818 versus 0.167 CFU/ ml) and 36 h (2.767 versus 1.402 CFU/ml) of incubation. These results demonstrate that SE multiplication on the vitelline membrane may both precede and exceed multiplication resulting from penetration into the yolk contents during the first 36 h of unrefrigerated storage, reinforcing the importance of rapid refrigeration for protecting consumers from egg-transmitted illness.     

Multiplication of Salmonella enteritidis in egg yolks after inoculation outside, on, and inside vitelline membranes and storage at different temperatures

Prompt refrigeration to restrict bacterial growth is important for reducing eggborne transmission of Salmonella enterica serovar Enteritidis (SE). The nutrient-rich yolk interior is a relatively infrequent location for initial SE deposition in eggs, but migration across the vitelline membrane can result in rapid bacterial multiplication during storage at warm temperatures. The objective of the present study was to measure the multiplication of SE in yolks after introduction at three different locations and subsequent storage at a range of temperatures. Using an in vitro egg contamination model, approximately 100 CFU of SE was inoculated either inside yolks, onto the exterior surface of vitelline membranes, or into the adjacent albumen. After storage of samples from each inoculation group at 10, 15, 20, and 25°C for 24 h, SE was enumerated in yolks. For all three inoculation locations, the final SE levels in yolks increased significantly with increasing storage temperatures. At all storage temperatures, significant differences in SE multiplication were observed between inoculation sites (yolk inoculation>vitelline membrane inoculation>albumen inoculation). At 25°C, final log concentrations of 7.759 CFU of SE per ml (yolk inoculation), 2.014 CFU/ml (vitelline membrane inoculation), and 0.757 CFU/ml (albumen inoculation) were attained in yolks after storage. These results demonstrate that, even when the initial site of SE deposition is outside the egg yolk, substantial multiplication supported by yolk nutrients can occur during the first day of storage and the risk of bacterial growth increases at higher ambient storage temperatures.     

Effect of temperature on the growth response of Salmonella enteritidis inoculated onto the vitelline membranes of fresh eggs

The growth response of Salmonella Enteritidis (SE) on the vitelline membrane in vitro was studied with the use of a special tube devised specifically for the inoculation of SE onto the vitelline membrane and for the sampling of the yolk near the inoculation site. This latter ability allowed the detection of the movement of SE into the yolk. The growth of SE on the membrane was compared with that of SE inoculated into yolk and albumen in vitro and in ovo in fresh in-shell eggs. The incubation time was 2 days, and the incubation temperatures were 4, 8, 15, 27, and 37 degrees C. Comparison of the results obtained for in vitro growth showed that at 4, 8, and 15 degrees C, SE behaved as if it were in the albumen, with its numbers decreasing over time. At 27 and 37 degrees C, SE grew as if it were in yolk, with a maximum increase of 4.5 log CFU after 2 days at 37 degrees C. In no experiments involving growth on the vitelline membrane did SE appear in the yolk. Comparisons between in vitro and in ovo growth responses of SE in yolk and albumen indicate that SE growth on the membrane parallels that in the in-shell egg.     

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.     

Multiplication and motility of Salmonella enterica serovars enteritidis, infantis, and montevideo in in vitro contamination models of eggs

The invasive ability of Salmonella enterica serovars Enteritidis, Infantis, and Montevideo in eggs was examined. Strains of these serovars originating from egg contents, laying chicken houses, and human patients were experimentally inoculated (0.1-ml dose containing 78 to 178 cells) onto the vitelline membrane of eggs collected from specific-pathogen-free chickens and incubated at 25 degrees C. The test strains were detected in 25 of 138 yolk contents by day 6, indicating the penetration of Salmonella organisms through the vitelline membrane. There were no significant differences in overall rates of penetration between serovars. The organisms were also detected in the albumen from 125 of 138 eggs tested by day 6. Growth to more than 10(6) CFU/ml was observed in 48 of the 125 albumen samples. An inoculum of 1000 Salmonella cells was added to 15 ml of albumen at the edge of a petri plate. A 10-mm-diameter cylindrical well, the bottom of which was sealed with a polycarbonate membrane with 3.0-microm pores, was filled with egg yolk and placed into the albumen at the center of the dish, which was maintained at 25 degrees C. Experiments were performed in triplicate with each strain. Salmonella organisms in all the albumen samples were detected by day 11. However, motility of the organisms toward the yolk was observed in only two dishes inoculated with the Salmonella Enteritidis strain from a human patient and in one dish inoculated with the Salmonella Infantis strain from liquid egg. The albumen samples obtained from the dishes inoculated with the Salmonella Enteritidis strain had high numbers of bacteria (>10(8) CFU/ml). The present study suggests that Salmonella organisms in egg albumen are unlikely to actively move toward the yolk, although depositionon or near the vitelline membrane can be advantageous for proliferation.     

Outgrowth of Salmonellae and the physical property of albumen and vitelline membrane as influenced by egg storage conditions

This study was undertaken to determine the influence of storage time and temperature on the volume, weight, and pH of egg albumen, the physical strength of vitelline membrane, and the fate of Salmonella Enteritidis artificially inoculated into egg albumen. A fiber-optic probe was used for inoculation with Salmonella Enteritidis at 10(2), 10(4), or 10(6) cells per egg. Both fresh and inoculated eggs were stored at 4, 10, and 22 degrees C for 6 weeks. Five fresh uninoculated eggs from each storage group were collected each week, and the weight, volume, and pH of the egg albumen were measured. The forces, energies, and degrees of membrane deformation required to rupture the vitelline membranes also were determined from either albumen-free yolks or yolks surrounded by albumen. In separate experiments, five inoculated eggs were evaluated each week for populations of Salmonella Enteritidis. When the eggs were stored at 4 degrees C, the albumen retained significantly more volume and weight and had a relatively lower pH. The vitelline membranes from eggs stored at 4 and 10 degrees C required more force and energy for rupture. Salmonellae flourished at 22 degrees C, even in the albumen with the lowest initial population, 10(2) cells per egg. Storage at 4 and 10 degrees C inhibited the growth of salmonellae in the albumen of eggs with initial populations of 10(2), 10(4), or 10(6) cells per egg. In eggs with initial Salmonella populations of 10(6) cells per egg that were stored at 22 degrees C, the populations of reached as high as 10(10) cells per egg after 4 weeks of storage. Storage at 4 and perhaps 10 degrees C postponed the aging process of chicken eggs, preserved the antimicrobial agents of the albumen, and maintained the integrity of vitelline membrane. Low-temperature storage therefore had a significant impact on the safety and overall quality of the eggs.     

The vitelline membrane: Dynamics of cholesterol metabolism in hens' eggs

Changes in cholesterol levels in egg contents were studied to indicate trends and dynamics of processes of penetration through the vitelline membrane from yolk to egg white. Investigations were carried out on eggs stored for a period of 35 days at room temperature (22°C). Levels of free and esterified cholesterol were determined by the spectrophotometric method at 7-day intervals. The selective character of the vitelline membrane depends on conditions and period of storage. The study demonstrated that the vitelline membrane loses its selective nature between the twentieth and thirtieth days under particular conditions of storage. Following this period, a spontaneous penetration of cholesterol into the egg white is observed—i.e. the barrier equilibrium of the membrane becomes unstable. The presence of free cholesterol in egg white may be indicative of loss of egg freshness.     

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.     

Incubation of egg contents pools at an elevated temperature (42 degrees C) does not improve the rapid detection of Salmonella enteritidis phage type 14b

Detecting internal Salmonella Enteritidis (SE) contamination in eggs is essential for protecting public health. Pooling together > or = 10 eggs for sampling allows many eggs to be screened for contamination, but such pools must be incubated (usually at 25 to 37 degrees C) to permit small numbers of SE to multiply before further testing. The present study determined whether incubating egg contents pools at an elevated temperature (42 degrees C) could increase the rate of multiplication of a phage type 14b strain of SE sufficiently to support the detection of contamination by a rapid lateral flow immunodiffusion method within a single day. Pools of 10 eggs were contaminated with approximately 10 CFU of SE, supplemented with concentrated broth enrichment medium, and incubated at either 37 or 42 degrees C. Incubation of contaminated egg pools at 42 degrees C resulted in significantly higher SE levels after 6, 8, 10, and 12 h. However, incubation at 42 degrees C could only generate a mean log SE concentration of 4.21 CFU/ml within a single working day (8 h), inadequate to support efficient detection by most rapid assays. Detection of SE contamination in egg pools by a rapid lateral flow immunodiffusion test was not achieved at a high frequency until 12 h of incubation at 42 degrees C.     

Laying season and egg shell cracks on the growth of Salmonella enteritidis in the egg albumen during storage

We studied the effects of laying seasons and egg shell cracks on the ability of egg albumen to support the growth of Salmonella Enteritidis (SE) in eggs. Hens eggs used were those laid in February, June, and October in a farm in Japan and stored at 10, 20, and 30 degrees C, and at 30 degrees C after storage at 10 degrees C, immediately after receipt or after cracking the shell. At several-day intervals during storage, the egg contents were poured into a dish, SE was inoculated into albumen, and then the growth of SE during 3 days incubation at 18 degrees C was measured. The results demonstrated that storage temperature and laying season affected the growth of SE in the egg albumen. The proportion of eggs upon which albumen allowed the growth of SE was higher in the eggs stored at 30 degrees C than those stored at 10 degrees C. The growth of SE in eggs was lowest in the following order of laying: February, October, and June. SE grew preferably in albumen of cracked eggs than intact eggs.     

In vitro Survival and Growth of Salmonella Typhimurium inoculated on Yolk Membrane after long Term refrigerated Storage of Shell Eggs

Salmonella enterica serovar Typhimurium has been isolated from commercial egg production facilities in the United States. Given its importance as a causative organism for food-borne salmonellosis, identifying approximate timelines for bacterial invasion of the egg is needed. The objective of this study was to examine net growth of S. Typhimurium in egg components over time. In trial 1 eggs were collected over a 24 hour period from a flock of single comb white leghorn hens while in trial 2 eggs were picked up from a commercial laying source once a week over the course of eight weeks and stored. Eggs were held at refrigeration temperature and each week, subsets of eggs were cracked, separated into yolk and albumen components, and inoculated with 10⁸ CFU/ml of novobiocin and nalidixic acid (NO/NA) resistant S. Typhimurium onto the vitelline membrane of the egg. Yolks were then covered with albumen. Eggs were incubated for twenty-four hours at 25°C. After incubation eggs were again separated into albumen, yolk, and vitelline membrane samples. In trial 1, S. Typhimurium net growth occurred in albumen by the second week and continued from 4 to 8 weeks while in trial 2 net growth only occurred at week 5 and 7. S. Typhimurium net growth on vitelline membranes occurred by 2 weeks and continued from 4 to 8 weeks in trial 1 while no net growth occurred in trial 2 over the 8 week period. Yolk samples showed no net increases in S. Typhimurium populations over the 8 week period. An erratum to this article is available at .     

In vitro Survival and Growth of Salmonella Typhimurium inoculated on Yolk Membrane after long Term refrigerated Storage of Shell Eggs

Salmonella enterica serovar Typhimurium has been isolated from commercial egg production facilities in the United States. Given its importance as a causative organism for food-borne salmonellosis, identifying approximate timelines for bacterial invasion of the egg is needed. The objective of this study was to examine net growth of S. Typhimurium in egg components over time. In trial 1 eggs were collected over a 24 hour period from a flock of single comb white leghorn hens while in trial 2 eggs were picked up from a commercial laying source once a week over the course of eight weeks and stored. Eggs were held at refrigeration temperature and each week, subsets of eggs were cracked, separated into yolk and albumen components, and inoculated with 10⁸ CFU/ml of novobiocin and nalidixic acid (NO/NA) resistant S. Typhimurium onto the vitelline membrane of the egg. Yolks were then covered with albumen. Eggs were incubated for twenty-four hours at 25°C. After incubation eggs were again separated into albumen, yolk, and vitelline membrane samples. In trial 1, S. Typhimurium net growth occurred in albumen by the second week and continued from 4 to 8 weeks while in trial 2 net growth only occurred at week 5 and 7. S. Typhimurium net growth on vitelline membranes occurred by 2 weeks and continued from 4 to 8 weeks in trial 1 while no net growth occurred in trial 2 over the 8 week period. Yolk samples showed no net increases in S. Typhimurium populations over the 8 week period.     

Comparison of homogenization methods for recovering Salmonella Enteritidis from eggs

For Salmonella Enteritidis (SE) detection, shell eggs have been homogenized with stomachers, with electric blenders, and by hand massaging. However, to date, there have been no published reports addressing whether the method of homogenization affects the recovery of SE from raw eggs. Three inoculum levels (10, 126, and 256 SE cells per pool of 10 eggs) were used to conduct three experiments. The 10-egg pools were homogenized by one of four homogenization methods--mechanical stomaching, electric blending, hand massaging, and hand stirring-for 30 s. The homogenized eggs were then incubated at 37 degrees C, and SE colonies were enumerated after 24 and 48 h of incubation. After 24 h of incubation, no SE was recovered from egg samples from stomached or electrically blended pools inoculated with <10 cells, while levels of 106 CFU/ml were found for samples from whipped or hand-massaged pools inoculated with <10 cells. Similarly, after 24 h of incubation, the numbers of SE cells recovered from hand-massaged or hand-stirred egg pools inoculated with 126 cells were significantly larger than the numbers recovered from stomached or electrically blended egg pools inoculated with 126 cells. The number of SE cells recovered from samples homogenized with a blender was still significantly smaller than the numbers recovered from samples homogenized by the other three methods when the inoculum level was increased to 256 CFU per pool. However, the SE count for all samples approached 9 log10 CFU/ml after 48 h of incubation. It is concluded that the detection of small SE populations in shell egg samples could be improved with the use hand massaging and hand stirring for homogenization.     

Influence of eggshell condensation on eggshell penetration and whole egg contamination with Salmonella enterica serovar enteritidis

Shells of agar-filled and whole eggs were inoculated with 10(3) to 10(4) CFU of Salmonella enterica serovar Enteritidis per eggshell. The agar-filled eggs were used to study bacterial eggshell penetration, and the whole egg results were used to characterize contamination of the egg contents. In each group, half of the eggs were stored for 21 days at 20 degrees C and 60% relative humidity (RH), and the other half was stored for 24 h at 6 degrees C and then for 20 days at 20 degrees C. The latter conditions resulted in condensation on the eggshell for 30 min from the moment the eggs were placed in the 20 degrees C chamber. Taking into account the ages at which hens were studied (39, 53, and 67 weeks), an average of 62% of the eggshells with condensate were penetrated compared with 43% for the control group; this difference was significant (P < 0.01). No significant difference in whole egg contamination was found; 18% of the control eggs were contaminated compared with 22% of the condensate eggs. Whole egg contamination was significantly higher for eggs from the hens at an older age (67 weeks). This difference probably was not due to a higher penetration potential because differences were not observed for the corresponding agar-filled eggs. Condensation on the eggshell seemed to encourage bacterial penetration of the eggshell but had a smaller impact on whole egg contamination.     

Salmonella Enteritidis Risk Assessment: A Kinetic Analysis

ABSTRACT: Egg and egg preparations are important vehicles for Salmonella enteritidis infections. The influence of time-temperature becomes important when the presence of this microorganism is found in commercial shell eggs, particularly in countries where refrigeration is not mandatory as in Chile. The objective of this research was to develop a mathematical model to analyze the Salmonella enteritidis risk under variable ambient temperatures. Breakdown of vitelline egg membrane was assumed to be required for initiation of bacterial growth. When the critical factor concerning safety is the vitelline membrane breakdown, 15 °C was found to be the storage threshold temperature for a 30-d shelf life. This computer based tool can be used as a contribution in current regulation adjustments or modifications.     

Growth ofSalmonella enteritidis(SE) in egg contents from hens vaccinated with an SE bacterin

Vaccination of hens withSalmonella enteritidis(SE) bacterin has become an important industry management tool to reduce both the incidence of SE in flocks and the production of SE-contaminated eggs. After vaccination, antibodies to SE can be found in both the serum and egg yolks. The current study was undertaken to examine whether the antibodies deposited in eggs after vaccination would have any effect on thein vitrogrowth of 10 SE seeded into pooled egg contents and incubated at 37°C for 24 h. The SE inoculum grew well in egg contents from control-vaccinated hens, and a high percentage of these samples were culture positive for SE. Conversely, significantly fewer egg contents from vaccinated hens were positive for SE at weeks 2, 3, 4, and 5 post-primary vaccination and weeks 1, 2, and 4 post-secondary vaccination. After 24 h incubation, the SE inoculum grew to 10⁸–10⁹organisms ml⁻¹in egg contents from control hens, while growing to less than 10⁴organisms ml⁻¹in egg contents from vaccinated hens. No inhibition of growth was observed for a second organism,Proteus mirabilis, in egg contents from bacterin-vaccinated hens, indicating antigen-specificity of the activity. Inhibition of SE growth was observed in egg contents from vaccinated hens diluted 1:5 in eggs from control hens. Mixed results were observed for egg contents receiving a 10-fold higher SE inoculum with significant inhibition of growth observed in one trial and minimal inhibition observed in a second trial. Supplementing the egg samples with an iron source abrogated the inhibition, suggesting that the iron- restricted environment found in eggs may play a role in the observed inhibition.     

Effectiveness of chitosan‐based coating in improving shelf‐life of eggs

Despite eggs having a natural packaging—shell—they are perishable and can lose their quality during storage. Chitosan‐based coatings were applied to shell eggs to examine potential effects on egg quality properties (weight loss, Haugh unit, yolk index) during 4 weeks of storage. Mineral amounts in yolks were also evaluated after 4 weeks of storage. Three chitosan‐based coatings produced with organic acids (acetic‐(C‐AA), lactic‐(C‐LA), and propionic (C‐PA)) were evaluated on shelf‐life enhancements of fresh egg quality. All chitosan‐coated eggs showed greater interior quality than the non‐coated eggs. The coatings significantly maintained weight loss compared to the control specimen (4.96%). Lower weight loss (3.45% for C‐PA, 3.53% for C‐LA) was observed in the coated eggs. Eggshell chitosan coat containing lactic and propionic acids maintained higher Haugh unit and yolk index than eggs coated with acetic acid. Uncoated (UC) eggs changed from grade ‘A’ to ‘B’ after 1 week of storage. Chitosan‐based coating containing lactic and propionic acids maintained eggs in grade ‘A’ for 4 weeks. Haugh unit showed that C‐LA and C‐PA effectively maintained eggs at grade ‘A’ quality for at least 3 weeks more than control and 1 week more than C‐AA. Results also indicated that the chitosan coating maintained mineral amounts at nutritional values (especially calcium, iron and magnesium concentration) in yolks after 4 weeks storage. Copyright © 2006 Society of Chemical Industry     

Super Chilling Enhances Preservation of the Freshness of Salted Egg Yolk During Long-Term Storage

ABSTRACT:  Pasteurized egg yolk with 10% (w/w) salt was stored at 5, –5, –15, –20, and –30 °C for 1 to 6 mo, respectively. Changes in generation of volatiles of the stored samples (5 and −5 °C for 6 mo) were analyzed by SPME-GC-MS. Emulsifying properties of egg yolk stored at −5, −15, −20, and −30 °C for 1 mo, respectively, were also evaluated by measurement of emulsion particle diameters in model emulsions prepared with the yolk samples. In addition, structural changes in low-density lipoprotein (LDL) in the egg yolks dependent on storage conditions for 6 mo were evaluated by ³¹P–NMR. Volatile compounds such as hexanal, 2-methylbutanal, and 3-methylbutanal increased in egg yolk during storage at 5 °C; however, volatile compounds hardly increased in any samples stored at −5 °C (super chilling). The mean emulsion particle diameter in super chilled egg yolk was significantly smaller than that in egg yolk stored at the other lower temperatures. In addition, the results of ³¹P–NMR evaluation suggested that prevention of structural changes of LDL resulted in maintenance of emulsifying properties of egg yolk. Thus, these results indicate that super chilling is an effective means of preserving salted egg yolk during long-term storage.     

Combination of Super Chilling and High Carbon Dioxide Concentration Techniques Most Effectively to Preserve Freshness of Shell Eggs during Long-Term Storage

ABSTRACT:  This study was made to examine the combined effects of stored temperature and carbon dioxide atmosphere on shell egg quality. The shell eggs were packed into polyethylene terephthalate/polyethylene (PET/PE) pouches and stored at 0 °C (super chilling), 10 °C, and 20 °C, respectively for 90 d. The atmospheric carbon dioxide concentration was controlled to obtain the 3 concentration levels of high (about 2.0%), medium (about 0.5%), and low (below 0.01%). Changes in Haugh unit (HU) values, weakening of vitelline membranes, and generation of volatiles were analyzed to evaluate the freshness of shell eggs. Results showed that, compared with the other combinations, the technique of super chilling and high carbon dioxide concentration enabled shell eggs to be most effectively stored for 90 d, based on estimations of the statistical significances of differences in HU values, and on maintaining the initial HU values during storage. In addition, the storage of shell eggs using this combination technique was found to significantly prevent the weakening of the vitelline membrane based on the estimations of numbers of eggs without vitelline membrane breakage when eggs broke, and significantly lowered the incidence of hexanal in the yolk from exposure to the gas chromatographic-mass spectrometric analyses of volatiles. Thus, these results confirmed that the combination of super chilling and high carbon dioxide concentration was the most effective technique for preserving shell eggs during a long term of 90 d compared with other combination techniques.     

Multiplication in egg yolk and survival in egg albumen of Salmonella enterica serotype Enteritidis strains of phage types 4, 8, 13a, and 14b

Refrigeration of eggs is vital for restricting the multiplication of Salmonella enterica serotype Enteritidis contaminants, but differences between Salmonella Enteritidis strains or phage types in their survival and multiplication patterns in egg contents might influence the effectiveness of refrigeration standards. The present study compared the abilities of 12 Salmonella Enteritidis isolates of four phage types (4, 8, 13a, and 14b) to multiply rapidly in egg yolk and to survive for several days in egg albumen. The multiplication of very small numbers of Salmonella Enteritidis inoculated into yolk (approximately 10(1) CFU/ml) was monitored during 24 h of incubation at 25 degrees C, and the survival of much larger numbers of Salmonella Enteritidis inoculated into albumen (approximately 10(5) CFU/ml) was similarly evaluated during the first 3 days of incubation at the same temperature. In yolk, the inoculated Salmonella Enteritidis strains multiplied to mean levels of approximately 10(3) CFU/ml after 6 h of incubation and 10(8) CFU/ml after 24 h. In albumen, mean levels of approximately 10(4) CFU/ml or more of Salmonella Enteritidis were maintained through 72 h. Although a few differences in multiplication and survival were observed between individual isolates, the overall range of values was relatively narrow, and no significant differences (P < 0.05) were evident among phage types.