Multiplication of Salmonella Enteritidis on the Yolk Membrane and Penetration to the Yolk Contents at 30°C in an In Vitro Egg Contamination Model

2008 ◽  
Vol 71 (9) ◽  
pp. 1905-1909 ◽  
Author(s):  
RICHARD K. GAST ◽  
RUPA GURAYA ◽  
JEAN GUARD-BOULDIN ◽  
PETER S. HOLT
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Vitelline Membrane ◽  
Critical Aspect ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Exterior Surface ◽  
Increased Risk ◽  
The Mean ◽  
Uncommon Location

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°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

2010 ◽  
Vol 73 (10) ◽  
pp. 1902-1906 ◽  
Author(s):  
RICHARD K. GAST ◽  
RUPA GURAYA ◽  
JEAN GUARD ◽  
PETER S. HOLT
Keyword(s):  
Bacterial Growth ◽  
Salmonella Enteritidis ◽  
Egg Yolk ◽  
Vitelline Membrane ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Exterior Surface ◽  
Different Temperatures ◽  
Subsequent Storage ◽  
Storage 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.

Multiplication and Motility of Salmonella enterica Serovars Enteritidis, Infantis, and Montevideo in In Vitro Contamination Models of Eggs

2006 ◽  
Vol 69 (5) ◽  
pp. 1012-1016 ◽  
Author(s):  
TOSHIYUKI MURASE ◽  
KAZUHIKO FUJIMOTO ◽  
RUI NAKAYAMA ◽  
KOICHI OTSUKI
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Egg Yolk ◽  
Vitelline Membrane ◽  
Human Patient ◽  
Polycarbonate Membrane ◽  
Specific Pathogen ◽  
Egg Albumen ◽  
Petri Plate

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°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 106 CFU/ml was observed in 48 of the 125 albumen samples. An inoculum of 1,000 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-μm pores, was filled with egg yolk and placed into the albumen at the center of the dish, which was maintained at 25°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 (>108 CFU/ml). The present study suggests that Salmonella organisms in egg albumen are unlikely to actively move toward the yolk, although deposition on or near the vitelline membrane can be advantageous for proliferation.

Inhibitory Effects of Enterococcus Strains Obtained from a Probiotic Product on In Vitro Growth of Salmonella enterica Serovar Enteritidis Strain IFO3313

2006 ◽  
Vol 69 (9) ◽  
pp. 2258-2262 ◽  
Author(s):  
WATTHANA THEPPANGNA ◽  
KOICHI OTSUKI ◽  
TOSHIYUKI MURASE
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Viable Cell ◽  
Mixed Cultures ◽  
Inhibitory Effects ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Cell Counts ◽  
Pure Cultures ◽  
Probiotic Product

Enterococcus faecium and Enterococcus gallinarum strains were isolated from a commercial probiotic product and the effects of these strains on the growth of Salmonella enterica serovar Enteritidis strain IFO3313 were investigated. Viable cell counts of Salmonella Enteritidis in mixed cultures with the probiotic product isolate of E. faecium were significantly (P < 0.05) lower than those in pure cultures after 6, 8, and 24 h when the cultures were incubated in heart infusion broth at 37 and 41°C. Significant differences in viable cell counts of Salmonella Enteritidis in mixed cultures with the probiotic product isolate of E. gallinarum and those in pure cultures were also observed after 8 and 24 h at 37 and 41°C. Similar observations were shown in mixed cultures of Salmonella Enteritidis with the reference strains of E. faecium GIFU8355 and E. gallinarum ATCC 49573. Significant differences in viable cell counts of these enterococcal strains were not shown among pure and mixed cultures with Salmonella Enteritidis. The pH values in pure and mixed cultures were 7.0 or 7.5 throughout the experiments. E. faecium strains were found to harbor the genes encoding enterocins A and B and showed inhibitory zones with a diameter of 4 to 6 mm against growth of Salmonella Enteritidis in the enterocin production assays. However, the E. gallinarum strains possessed neither of the enterocin genes tested and exhibited no inhibition zone in the enterocin production assays. These results indicated that enterococcal strains exhibit inhibitory effects on the growth of Salmonella Enteritidis and these effects were due to both enterocin and nonenterocin factors.

Protection of epithelial cells from Salmonella enterica serovar Enteritidis invasion by antibodies against the SPI-1 type III secretion system

2010 ◽  
Vol 56 (6) ◽  
pp. 522-526 ◽  
Author(s):  
Taseen S. Desin ◽  
Claudia S. Mickael ◽  
Po-King S. Lam ◽  
Andrew A. Potter ◽  
Wolfgang Köster
Keyword(s):  
Epithelial Cells ◽  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Secretion Systems ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Specific Antibodies ◽  
Type Iii ◽  
Food Borne Illness

Salmonella enterica serovar Enteritidis (Salmonella Enteritidis) is one of the major causes of bacterial food-borne illness in humans. During the course of infection, Salmonella Enteritidis uses 2 type III secretion systems (T3SS), one of which is encoded on Salmonella pathogenicity island 1 (SPI-1). SPI-1 plays a major role in the invasion process. In the present study, we evaluated the effect of sera against the SPI-1 T3SS components on invasion in vitro using polarized human intestinal epithelial cells (Caco-2). Antisera to SipD protected Caco-2 cells against entry of wild-type Salmonella Enteritidis. On the other hand, sera against InvG, PrgI, SipA, SipC, SopB, SopE, and SopE2 did not affect Salmonella Enteritidis entry. To illustrate the specificity of anti-SipD mediated inhibition, SipD-specific antibodies were depleted from the serum. Antiserum depleted of SipD-specific antibodies lost its capacity to inhibit Salmonella Enteritidis entry. Thus, we demonstrate for the first time that antibodies against the SPI-1 needle tip protein (SipD) inhibit Salmonella Enteritidis invasion and that the SipD protein may be an important target in blocking SPI-1 mediated virulence of Salmonella Enteritidis.

Growth of Salmonella enterica Serovar Enteritidis in Albumen and Yolk Contents of Eggs Inoculated with This Organism onto the Vitelline Membrane

2005 ◽  
Vol 68 (4) ◽  
pp. 718-721 ◽  
Author(s):  
TOSHIYUKI MURASE ◽  
PETER S. HOLT ◽  
RICHARD K. GAST
Keyword(s):  
Salmonella Enterica ◽  
In Vitro Model ◽  
Human Infection ◽  
Vitelline Membrane ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Specific Pathogen ◽  
Vitro Model ◽  
Slight Growth ◽  
Culture Positive

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°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 × 102 to 4.2 × 108 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 × 102 to 4.3 × 108 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°C for 1 to 4 weeks or at 25°C for 1 week, slight growth (3.0 × 102 to 7.4 × 103 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.

scholarly journals Genotypic characterization of antibiotic-resistant Salmonella Enteritidis isolates in Dakar, Senegal

2007 ◽  
Vol 1 (03) ◽  
pp. 284-288 ◽  
Author(s):  
Amy Gassama Sow ◽  
Abdoul Aziz Wane ◽  
Mamadou Hadi Diallo ◽  
Cheikh Saad-Bouh Boye ◽  
Awa Aïdara-Kane
Keyword(s):  
Antibiotic Resistance ◽  
Antimicrobial Resistance ◽  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Data Bank ◽  
Human Infection ◽  
Molecular Techniques ◽  
Conjugative Plasmid ◽  
Bacterial Strains ◽  
Salmonella Enterica Serovar Enteritidis

Background: It is well established that Salmonella enterica is a major cause of food-borne disease worldwide. In Africa, according to the Who Global Salm-Surv country data bank from 2000 to 2002 Salmonella enterica serovar Enteritidis was the most common serotype involved in human salmonellosis. In Dakar this serotype of Salmonella has been reported as a frequent and an increasing cause of human infection. Methodology: The genetic determinants of the antimicrobial resistance of 25 selected multiresistant strains of Salmonella enterica serovar Enteritidis referred to the National Reference Center for Enterobacteria (NRCE) in Dakar were investigated using molecular techniques. Results: All strains carried blaTEM 1 genes. Five harboured three types of class 1 integrons with gene cassettes dfrA15, dfrA1-aadA1 and dfrA7. Multiresistance was due to a 23 Kb conjugative plasmid. DNA fingerprinting by macrorestriction of genomic DNA revealed a single related group suggesting that strains might be clonal. Conclusions: The spread of resistance genes through plasmid transfer plays an important role in the dissemination of antibiotic resistance in enteric pathogens such as Salmonella Enteritidis; the risk of transmissibility of antibiotic resistance between different bacterial strains highlights the urgent need to develop strategies to limit the spread of antimicrobial resistance among bacterial enteropathogens.

scholarly journals Salmonella enterica Serovar Enteritidis Pathogenicity Island 1 Is Not Essential for but Facilitates Rapid Systemic Spread in Chickens

2009 ◽  
Vol 77 (7) ◽  
pp. 2866-2875 ◽  
Author(s):  
Taseen S. Desin ◽  
Po-King S. Lam ◽  
Birgit Koch ◽  
Claudia Mickael ◽  
Emil Berberov ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Systemic Infection ◽  
Poultry Meat ◽  
Wild Type ◽  
Secretion Systems ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Systemic Spread ◽  
Mutant Strains ◽  
Vitro Analysis

ABSTRACT Salmonella enterica subsp. enterica serovar Enteritidis is a leading cause of human food-borne illness that is mainly associated with the consumption of contaminated poultry meat and eggs. To cause infection, S. Enteritidis is known to use two type III secretion systems, which are encoded on two salmonella pathogenicity islands, SPI-1 and SPI-2, the first of which is thought to play a major role in invasion and bacterial uptake. In order to study the role of SPI-1 in the colonization of chicken, we constructed deletion mutants affecting the complete SPI-1 region (40 kb) and the invG gene. Both ΔSPI-1 and ΔinvG mutant strains were impaired in the secretion of SipD, a SPI-1 effector protein. In vitro analysis using polarized human intestinal epithelial cells (Caco-2) revealed that both mutant strains were less invasive than the wild-type strain. A similar observation was made when chicken cecal and small intestinal explants were coinfected with the wild-type and ΔSPI-1 mutant strains. Oral challenge of 1-week-old chicken with the wild-type or ΔSPI-1 strains demonstrated that there was no difference in chicken cecal colonization. However, systemic infection of the liver and spleen was delayed in birds that were challenged with the ΔSPI-1 strain. These data demonstrate that SPI-1 facilitates systemic infection but is not essential for invasion and systemic spread of the organism in chickens.

Effect of Temperature on the Growth Response of Salmonella Enteritidis Inoculated onto the Vitelline Membranes of Fresh Eggs

2003 ◽  
Vol 66 (8) ◽  
pp. 1368-1373 ◽  
Author(s):  
G. J. FLEISCHMAN ◽  
C. L. NAPIER ◽  
D. STEWART ◽  
S. A. PALUMBO
Keyword(s):  
Salmonella Enteritidis ◽  
Growth Response ◽  
Effect Of Temperature ◽  
Vitelline Membrane ◽  
Growth Responses ◽  
In Ovo ◽  
Maximum Increase ◽  
Comparison Of The Results ◽  
As If

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°C. Comparison of the results obtained for in vitro growth showed that at 4, 8, and 15°C, SE behaved as if it were in the albumen, with its numbers decreasing over time. At 27 and 37°C, SE grew as if it were in yolk, with a maximum increase of 4.5 log CFU after 2 days at 37°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 Enteritidis Phage Type 30 in Almond Hull and Shell Slurries and Survival in Drying Almond Hulls

2006 ◽  
Vol 69 (4) ◽  
pp. 712-718 ◽  
Author(s):  
AARON R. UESUGI ◽  
LINDA J. HARRIS
Keyword(s):  
Incubation Time ◽  
Rapid Growth ◽  
Salmonella Enterica ◽  
Salmonella Enteritidis ◽  
Phage Type ◽  
Weather Conditions ◽  
Water Sources ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Outbreak Strain ◽  
High Concentrations

Traceback investigation of a 2000 to 2001 outbreak of salmonellosis associated with consumption of raw almonds led to isolation of the outbreak strain Salmonella enterica serovar Enteritidis phage type (PT) 30 on three geographically linked almond farms. Interviews with these growers revealed that significant rain fell during the 2000 harvest when many almonds were drying on the ground. The objectives of this study were to document weather conditions during the 2000 harvest, determine the potential for growth of Salmonella Enteritidis PT 30 in hull or shell slurries, and evaluate survival of Salmonella Enteritidis PT 30 on wet almond hulls during drying. Dry almond hulls and in-shell kernels wetted for 24 h increased in weight by 250 to 300% and 100%, respectively. Both hull and shell slurries supported rapid growth of Salmonella Enteritidis PT 30 at 24°C; slurries containing hulls also supported growth at 15°C. Maximum Salmonella Enteritidis PT 30 concentrations of 6.2 and 7.8 log CFU/ml were observed at 15 and 24°C, respectively. Salmonella Enteritidis PT 30 grown in wet hulls that were incubated at 24°C survived drying at either 15 or 37°C. Reductions of 1 to 3 log CFU/g of dry hull were observed during drying; reductions generally declined as incubation time increased from 2 to 7 days. Evaluation of shipping records revealed that approximately 60% of outbreak-associated almonds had not been exposed to rain, eliminating this factor as the sole cause of the outbreak. However, the data provide evidence that wet almonds may be a greater risk for high concentrations of Salmonella, and specific guidelines should be established for harvesting and processing almonds that have been exposed to rain or other water sources.

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