Research Note: Internal organ colonization by Salmonella Enteritidis in experimentally infected layer pullets reared at different stocking densities in indoor cage-free housing.

Contamination of eggs by Salmonella has often been identified as a source of food-borne human illness. S. Enteritidis is deposited inside developing eggs when invasive infections of laying hens reach the reproductive organs. The susceptibility of hens in cage-based housing systems to S. Enteritidis has been associated with their stocking density, but the applicability of this information to extensive (cage-free) systems is uncertain. The present study assessed internal organ colonization by S. Enteritidis in egg-type pullets reared at 2 different stocking densities in cage-free housing. Pullets were reared at either 374 cm2 or 929 cm2 of floor space per bird. At 16 wk of age, 4 groups of 72 pullets were moved into isolation rooms simulating commercial cage-free barns; 1/3 of the pullets in 2 rooms were orally inoculated with S. Enteritidis immediately after transfer and pullets in 2 rooms were similarly infected at 19 wk. At 6 and 12 d postinoculation, the pullets were euthanized and samples of liver, spleen, and intestinal tract were removed for bacteriologic culturing. No significant differences (P > 0.05) in S. Enteritidis isolation frequencies from any tissue were observed between high and low density rearing groups following infection at either age. However, S. Enteritidis was found significantly (P < 0.05) more frequently among pullets infected orally at 19 wk than at 16 wk in spleens and intestines. Likewise, the frequency of S. Enteritidis isolation from all birds (inoculated plus contact-exposed) at 19 wk was significantly higher than at 16 wk in livers and spleens. This increased susceptibility to invasive S. Enteritidis infection at reproductive maturity emphasizes the importance of risk reduction at a critical stage in the egg production cycle.

Serotype Screening of Salmonella enterica Subspecies I by Intergenic Sequence Ribotyping (ISR): Critical Updates.

(1) Background: Foodborne illness from Salmonella enterica subspecies I is most associated with approximately 32 out of 1600 serotypes. While whole genome sequencing and other nucleic acid-based methods are preferred for serotyping, they require expertise in bioinformatics and often submission to an external agency. Intergenic Sequence Ribotyping (ISR) assigns serotype to Salmonella in coordination with information freely available at the National Center for Biotechnology Information. ISR requires updating because it was developed from 26 genomes while there are now currently 1804 genomes and 1685 plasmids. (2) Methods: Serotypes available for sequencing were analyzed by ISR to confirm primer efficacy and to identify any issues in application. Differences between the 2012 and 2022 ISR database were tabulated, nomenclature edited, and instances of multiple serotypes aligning to a single ISR were examined. (3) Results: The 2022 ISR database has 268 sequences and 40 of these were assigned new NCBI accession numbers that were not previously available. Extending boundaries of sequences resolved hdfR cross-alignment and reduced multiplicity of alignment for 37 ISRs. Comparison of gene cyaA sequences and some cell surface epitopes provided evidence that homologous recombination was potentially impacting results for this subset. There were 99 sequences that still had no match with an NCBI submission. (4) The 2022 ISR database is available for use as a serotype screening method for Salmonella enterica subspecies I. Finding that 36.9% of the sequences in the ISR database still have no match within the NCBI Salmonella enterica database suggests that there is more genomic heterogeneity yet to characterize.

Research Note: Contamination of eggs by Salmonella Enteritidis and Salmonella Typhimurium in experimentally infected laying hens in indoor cage-free housing.

Contaminated eggs are a leading source of human Salmonella infections and this problem continues to challenge public health authorities and egg industries around the world. Salmonella invasion of the ovaries and oviducts of infected laying hens can result in bacterial deposition inside the edible portions of developing eggs. The introduction, persistence, and transmission of salmonellae in commercial egg-laying flocks are influenced by flock management practices, but the food safety ramifications of different types of laying hen housing remain unresolved. The present study assessed the frequency of internal contamination of eggs after experimental Salmonella Enteritidis and S. Typhimurium infection of laying hens in indoor cage-free housing. Groups of 72 hens were housed on wood shavings in isolation rooms simulating commercial cage-free barns with community kick-out nest boxes and perches and 1/3 of the hens in each room were orally inoculated with 8.0 × 107 cfu of 2-strain mixtures of either S. Enteritidis (2 rooms) or S. Typhimurium (2 rooms), and the entire internal contents of all eggs laid 5 to 30 d postinoculation in nest boxes or on the flooring substrate were cultured to detect Salmonella. Contaminated eggs were laid between 8 and 28 d postinoculation. The overall incidence of S. Enteritidis isolation from eggs (3.41%) was significantly (P = 0.0005) greater than S. Typhimurium (1.19%). The contamination frequencies associated with the 2 egg collection locations were not significantly different (P > 0.05). These results demonstrate that oral infection of a relatively small proportion of laying hens in indoor cage-free housing with invasive Salmonella serovars can result in the production of internally contaminated eggs at low frequencies over a period of nearly a month postinoculation.

Equivalency of peroxyacetic acid to chlorine as a shell egg sanitizing rinse.

In the United States, all shell eggs processed under the USDA Agricultural Marketing Service voluntary grading standards must receive a shell sanitizing rinse of 100-200 ppm chlorine or its equivalent after leaving the washing process. A study was conducted to determine the concentration of peroxyacetic acid (PAA) which would be equivalent to 100-200 ppm chlorine (Cl) in reducing target organisms under the required washing conditions for shell eggs. Three isolates of Salmonella spp. (Enteritidis, Braenderup, and Typhimurium), as well as Enterobacter cloacae were used as inocula. Sanitizing treatments were negative control; deionized water; 100 and 200 ppm Cl; and 50-500 ppm PAA (7 concentrations). Considering all isolates tested, 100 and 200 ppm chlorine had 2.6 and 2.3 log cfu/mL cultural organisms remaining on shell surface; 50 and 100 ppm peracetic acid had 1.9 and 1.0 log cfu/mL cultural organisms remaining, respectively, compared with untreated control average of 3.8 log cfu/mL (P < 0.001). Salmonella Typhimurium was least resistant to shell sanitizer treatments. Peroxyacetic acid concentrations >250 ppm did not produce significant reductions in microbial populations as PAA concentration increased. Culturing for the prevalence of viable and injured organisms, 400-500 ppm PAA resulted in fewer eggs (P < 0.0001) being positive for Salmonella spp. E. cloacae was culturable via enrichment from 99.4% of inoculated eggs, regardless of sanitizer treatment. The results of this study indicate that 50-100 ppm PAA is equivalent to 100-200 ppm chlorine in reducing egg surface microorganisms. The use of 400-500 ppm PAA resulted in a lower incidence of viable, but not culturable, Salmonella spp. on the shell surface. E. cloacae resulted in almost 100% viable, but not culturable, organism recovery for all sanitizing treatments and should be considered as an indicator organism when studying processing facility sanitation procedures.

Research Note: Horizontal transmission and internal organ colonization by Salmonella Enteritidis and Salmonella Kentucky in experimentally infected laying hens in indoor cage-free housing.

The transmission of Salmonella to humans via contaminated eggs is an international public health concern. S. Enteritidis is deposited inside eggs after colonizing reproductive tissues of infected hens. Diverse housing facility characteristics and flock management practices influence Salmonella persistence and transmission in poultry, but the food safety consequences of different housing systems for laying hens remain unresolved. The present study compared the horizontal transmission of infection and invasion of internal organs during the first 2 wk after experimental S. Enteritidis and S. Kentucky infection of laying hens in indoor cage-free housing. Groups of 72 hens were housed in isolation rooms simulating commercial cage-free barns, and 1/3 of the hens in each room were orally inoculated with either S. Enteritidis (2 rooms) or S. Kentucky (2 rooms). At 6 d and 12 d postinoculation, 12 inoculated and 24 contact-exposed hens in each room were euthanized, and samples of liver, spleen, ovary, oviduct, and intestinal tract were removed for bacteriologic culturing. All orally inoculated hens were positive for intestinal colonization by S. Enteritidis at 6 d postinfection, and 70.8% of contact-exposed hens had become colonized by 12 d. S. Enteritidis was isolated from 100% of livers and 50.0% of ovaries from inoculated birds at 6 d and from 41.7% of livers and 10.4% of ovaries from contact-exposed birds at 12 d. The majority of both orally inoculated and contact-exposed hens were positive for intestinal colonization by S. Kentucky at 6 d, but S. Kentucky was found in other internal organs of both inoculated and contact-exposed hens significantly (P < 0.05) less often than S. Enteritidis at both sampling intervals. These results indicate that Salmonella infection can spread rapidly and extensively among hens in cage-free indoor housing, including a high frequency of internal organ involvement for invasive S. Enteritidis.

Pooling of Laying Hen Environmental Swabs and Efficacy of Salmonella Detection.

ABSTRACT: Environmental testing for Salmonella Enteritidis is required for U.S. shell egg producers with ≥3,000 hens on a farm. The egg producer assumes all costs for the mandatory testing. According to the U.S. Food and Drug Administration (FDA) Egg Rule, either manure scraper or drag swabs can be collected according to published guidelines and requirements. The present study was undertaken to determine the efficacy of Salmonella detection with one-, two-, and four-swab pools of either manure scraper or drag swabs. Resistant isolates of Salmonella serovars Enteritidis (1,000 ppm of streptomycin), Heidelberg (200 ppm of nalidixic acid [NA]), Typhimurium (200 ppm of NA), and Kentucky (200 ppm of NA) were utilized. Low (approximately 8.4 CFU) and high (approximately 84 CFU) levels of inocula were introduced onto a single swab within a pool. Single flocks from each conventional cage (manure scraper swabs) and cage-free barn (drag swabs) were monitored throughout the study at the ages required under the FDA Egg Rule. The highest and most consistent recovery of inoculum was found in single swab samples. For low dose inocula, recovery of isolates was low from single manure scraper swabs (57.9 to 29.2%) and decreased as more swabs were added to the pool. Recovery of isolates from manure scraper swabs was higher for high dose inocula, although Salmonella Heidelberg was outcompeted by the naturally occurring flora and had the lowest rate of recovery among the isolates tested. One- and two-swab pools of drag swabs had similar rates of recovery at both low and high doses for Salmonella Enteritidis, Salmonella Heidelberg, and Salmonella Typhimurium. When Salmonella Enteritidis and Salmonella Kentucky were combined in an inoculum, Salmonella Enteritidis was recovered at a much higher rate than was Salmonella Kentucky for all types of swabs and doses of inocula. Pooling of two drag swabs allowed for similar detection of low and high dose Salmonella, but the pooling of manure scraper swabs decreased detection of low dose Salmonella.

Thermal Inactivation Kinetics of Three Heat-Resistant Salmonella Strains in Whole Liquid Egg.

The heat resistance of three heat-resistant strains of Salmonella was determined in whole liquid egg (WLE). Inoculated samples in glass capillary tubes were completely immersed in a circulating water bath and held at 56, 58, 60, 62, and 64°C for predetermined lengths of time. The recovery medium was tryptic soy agar with 0.1% sodium pyruvate and 50 ppm of nalidixic acid. Survival data were fitted using log-linear, log-linear with shoulder, and Weibull models using GInaFiT version 1.7. Based on the R2 and mean square error, the log-linear with shoulder and Weibull models consistently produced a better fit to Salmonella survival curves obtained at these temperatures. Contaminated WLE must be heated at 56, 60, and 64°C for at least 33.2, 2.7, and 0.31 min, respectively, to achieve a 4-log reduction of Salmonella; 39.0, 3.1, and 0.34 min, respectively, for a 5-log reduction; and 45.0, 3.5, and 0.39 min, respectively, for a 6-log reduction. The z-values calculated from the D-values were 3.67 and 4.18°C for the log-linear with shoulder and Weibull models, respectively. Thermal death times presented in this study will be beneficial for WLE distributors and regulatory agencies when designing pasteurization processes to effectively eliminate Salmonella in WLE, thereby ensuring the microbiological safety of the product.

Contamination of eggs by Salmonella Enteritidis in experimentally infected laying hens of four commercial genetic lines in conventional cages and enriched colony housing.

Human illness caused by the consumption of eggs contaminated with Salmonella Enteritidis is a continuing international public health concern. This pathogen is deposited inside the edible contents of eggs as a consequence of its ability to colonize reproductive tissues in infected hens. Conditions in the housing environment can influence the persistence and transmission of avian Salmonella infections, but the food safety ramifications of different poultry management systems are not entirely clear. The present study assessed the deposition of S. Enteritidis inside eggs laid by groups of experimentally infected laying hens of 4 commercial genetic lines (designated as white egg lines W1 and W2 and brown egg lines B1 and B2). Groups of hens from each line were housed at 555 cm2 of floor space per bird in both conventional cages and colony units enriched with access to perches and nesting areas. All hens were orally inoculated with 5.75 × 107 cfu of a 2-strain S. Enteritidis mixture, and the internal contents of eggs laid 5 to 24 D post-inoculation were cultured to detect the pathogen. No significant differences in egg contamination frequencies were found between the 2 housing systems for any of the hen lines. Contaminated eggs were laid between 7 and 21 D post-inoculation at an overall frequency of 2.47%, ranging from 0.25 to 4.38% for the 4 hen lines. The frequency of S. Enteritidis recovery from egg samples was significantly (P < 0.05) lower for line B2 than for any of the other lines, and the egg contamination frequency for line W1 was significantly greater than for line W2. The overall incidence of contamination among white eggs (3.38%) was significantly higher than among brown eggs (1.56%). These results demonstrate that S. Enteritidis deposition inside eggs can vary between genetic lines of infected laying hens, but housing these hens in 2 different systems did not affect the production of contaminated eggs.

Can acceptable quality angel food cakes be made using pasteurized shell eggs? The effects of mixing factors on functional properties of angel food cakes.

Due to recent Salmonella outbreaks, the pasteurized shell egg market is rapidly growing. One objection to using pasteurized eggs is the belief that they will produce unacceptable angel food cakes. Eggs were pasteurized using a hot water immersion process (56.7°C for 60 min) similar to that used by industry. Angel food cakes were made from the pasteurized egg white (PEW) as well as from raw egg white (REW) for comparison. Meringues were made using three mixer speed settings (low, medium, and high) and three durations for each speed. Functional qualities such as egg foaming were evaluated. Angel food cakes were compared in terms of cake volume, texture profile, and color values. When the optimal processing factors used for REW were applied to PEW, an inferior meringue was formed. However, by increasing the mixing time for PEW by 200% at the highest speed, an acceptable meringue was formed. The best angel food cake prepared from PEW had a volume only 6.8% less than that of the best cake prepared from REW. Texture profile analyses showed that the best angel food cake made from PEW was 13% firmer, 7.4% less springy, and 62% chewier than that from REW. Color analyses showed that PEW made a slightly darker colored cake crust than REW, although there were no significant differences in the crumb color. Modifying the mixing conditions for PEW resulted in angel food cakes with quality similar to that of cakes made with REW, thus overcoming an objection to using safer pasteurized shell eggs.

Colonization of internal organs by Salmonella Enteritidis in experimentally infected laying hens of four commercial genetic lines in conventional cages and enriched colony housing.

The prevalence of Salmonella Enteritidis in commercial egg-laying flocks is a prominent public health concern because contaminated eggs cause human illness. Deposition of this pathogen inside eggs results from bacterial colonization of reproductive tissues in infected hens. Environmental conditions can influence avian Salmonella infections, but the food safety consequences of different poultry housing systems remain uncertain. The present study assessed the invasion of internal organs by Salmonella Enteritidis in groups of experimentally infected laying hens of four commercial genetic lines (designated as white egg lines W1 and W2 and brown egg lines B1 and B2). Groups of hens from each line were housed at 555 cm2 of floor space per bird in both conventional cages and colony units enriched with access to perches and nesting areas. All hens were orally inoculated with 5.75 × 107 colony-forming units of a two-strain Salmonella Enteritidis mixture. At 6 to 7 d post-inoculation, hens were euthanized, and samples of liver, spleen, ovary, oviduct, and intestinal tract were removed for bacteriologic culturing. The frequency of Salmonella Enteritidis recovery from intestinal samples was significantly (P < 0.05) greater for the two white egg lines combined than for the two brown egg lines combined in both conventional cage (72.2% vs. 50.0%) and enriched colony housing systems (66.7% vs. 37.5%). The frequency of intestinal Salmonella Enteritidis isolation from line B1 was significantly higher from hens in conventional cages (47.2%) than in enriched colonies (22.2%), but no differences were observed for other hen lines. Line W1 yielded more positive intestinal samples than either brown egg line in conventional cages, and line B2 had fewer positive intestinal samples than all other lines in enriched colonies. There were no significant differences between hen lines or housing systems in Salmonella Enteritidis isolation from other internal organs. These results demonstrate that Salmonella Enteritidis colonization of the intestinal tract can vary between genetic lines of egg-laying hens and that some lines are subject to housing system influences on Salmonella susceptibility.

Prevalence, Persistence, and Antimicrobial Resistance of Campylobacter spp. from Eggs and Laying Hens Housed in Five Commercial Housing Systems.

Husbandry practices for laying hens in commercial egg production is a topic of interest from a social, economic, and regulatory standpoint. Animal welfare concerns regarding the use of conventional cages have arisen and consumer perceptions of hen welfare have led to a higher demand for cage-free eggs. The aim of this study was to assess the impact of housing systems on prevalence, persistence, and antimicrobial resistance (AMR) of Campylobacter from laying hens and shell eggs. A total of 425 samples were collected over a 10-month period from the North Carolina Layer Performance and Management Test and Campylobacter isolates were identified by serological, biochemical, and molecular tests. Genetic variability was evaluated using pulsed-field gel electrophoresis (PFGE) and AMR testing was performed. Prevalence of Campylobacter spp. ranged from 11.1% in the enrichable cages to 19.7% in the conventional systems. A greater prevalence of Campylobacter was found in the fecal swab samples from free-range birds compared with those of birds housed in the more intensive housing systems (p > 0.05). Overall, 72 isolates were confirmed as Campylobacter spp. by PCR. More than 90% of the isolates (n = 66) were identified as Campylobacter jejuni, followed by Campylobacter coli (n = 6). C. jejuni isolates displayed high levels of resistance to tetracycline (67%). Genetic variability of Campylobacter was high, with more than 20 PFGE patterns identified. Pattern "a" comprised 42% of isolates from all housing systems and was also the most persistent. This study suggests that housing systems of laying hens used for commercial shell egg production may impact the rate of Campylobacter shedding by layers. Isolation rates and tetracycline resistance levels of this pathogen are still of concern, emphasizing the need for well-implemented biosecurity measures on the farm.

Serotyping of Salmonella Enterica Isolated from Mice Caught on US Poultry Farms 1995 through 1998.

In this study, a collection of Salmonella enterica subspecies obtained from live mice caught on 32 poultry farms in the Northeast US between 1995 to 1998 was evaluated to provide a historical reference for serotype distribution during a time when egg contamination by serotype Enteritidis was at its peak. Of 821 mice cultured, 157 were positive (19.1%). Seven mice harbored two serotypes of Salmonella. Nine serotypes were detected, eight of which are often associated with foodborne illness. The three most prevalent serotypes were Enteritidis, Heidelberg, and Typhimurium. Enteritidis and Typhimurium were obtained from both spleens and intestines without preference according to type of sample. In contrast, Heidelberg was isolated most often from intestines and Schwarzengrund was most often obtained from spleens. These results support that the house mouse Mus musculus was a risk factor for introduction of multiple pathogenic Salmonella serotypes in poultry raised in the Northeast US during the mid-1990s. Isolates were submitted to the Food and Drug Administration and draft genomes for 64 isolates of Salmonella enterica serovar Enteritidis data have been released through the National Center for Biotechnology Information via the GenomeTrakr network.

Frequency and Duration of Fecal Shedding of Salmonella Serovars Heidelberg and Typhimurium by Experimentally Infected Laying Hens Housed in Enriched Colony Cages at Different Stocking Densities.

Eggs contaminated with Salmonella Enteritidis are leading sources of human salmonellosis, but Salmonella Heidelberg and Salmonella Typhimurium are also egg-associated pathogens. The management practices and housing facilities characterizing different systems for housing commercial egg flocks can influence Salmonella persistence and transmission. Animal welfare aspects of poultry housing have been widely debated, but their food safety ramifications are not thoroughly understood. The present study assessed the effects of two different bird stocking densities on the frequency and duration of fecal shedding of strains of Salmonella Heidelberg and Salmonella Typhimurium in groups of experimentally infected laying hens housed in colony cages enriched with perching and nesting areas. In separate trials, laying hens were distributed into two groups housed in enriched colony cages at stocking densities of 648 and 973 cm2/bird, and a third group was housed in conventional cages at 648 cm2/bird. All hens were orally inoculated with doses of 108 colony-forming units (CFU) of either Salmonella Heidelberg or Salmonella Typhimurium. At eight weekly postinoculation intervals, samples of voided feces were collected from beneath each cage and cultured to detect Salmonella. Fecal shedding of Salmonella Heidelberg continued for 8 wk in all housing groups, but Salmonella Typhimurium shedding ceased after as little as 5 wk in enriched colony cages at low stocking density. After Salmonella Heidelberg infection, the overall frequency of positive fecal cultures for all sampling dates combined was significantly (P < 0.05) greater from either conventional cages (51.0%) or enriched colony cages (46.5%) at high stocking density than from enriched colony cages at low stocking density (33.3%). No significant differences in Salmonella Typhimurium fecal isolation were identified between housing groups. These results demonstrate that stocking density can affect intestinal colonization and fecal shedding in laying hens for some (but not necessarily all) Salmonella serovars or strains.

Frequency and Duration of Fecal Shedding of Salmonella Enteritidis by Experimentally Infected Laying Hens Housed in Enriched Colony Cages at Different Stocking Densities.

Human infections with Salmonella Enteritidis are often attributed to the consumption of contaminated eggs, so the prevalence of this pathogen in egg-laying poultry is an important public health risk factor. Numerous and complex environmental influences on Salmonella persistence and transmission are exerted by management practices and housing facilities used in commercial egg production. In recent years, the animal welfare implications of poultry housing systems have guided the development of alternatives to traditional cage-based housing, but their food safety consequences are not yet fully understood. The present study assessed the effects of different bird stocking densities on the frequency and duration of fecal shedding of S. Enteritidis in groups of experimentally infected laying hens housed in colony cages enriched with perching and nesting areas. In two trials, groups of laying hens were distributed at two stocking densities (648 and 973 cm2/bird) into enriched colony cages and (along with a group housed in conventional cages at 648 cm2/bird) orally inoculated with doses of 1.0 × 108 cfu of S. Enteritidis. At 10 weekly postinoculation intervals, samples of voided feces were collected from beneath each cage and cultured to detect S. Enteritidis. Fecal shedding of S. Enteritidis was detected for up to 10 weeks postinoculation by hens in all three housing treatment groups. The overall frequency of positive fecal cultures was significantly (P < 0.05) greater from conventional cages than from enriched colony cages (at the lower stocking density) for the total of all sampling dates (45.0 vs. 33.3%) and also for samples collected at 4-9 weeks postinfection. Likewise, the frequency of S. Enteritidis isolation from feces from conventional cages was significantly greater than from enriched colony cages (at the higher hen stocking density) for the sum of all samples (45.0 vs. 36.7%) and at 6 weeks postinoculation. Moreover, the frequency of S. Enteritidis fecal recovery from enriched colony cages at the higher hen stocking was significantly greater than from similar cages at the lower stocking density for all 10 sampling dates combined (39.4 vs. 33.3%). These results suggest that stocking density can affect S. Enteritidis intestinal colonization and fecal shedding in laying hens, but some other difference between conventional and enriched colony cage systems appears to exert an additional influence.

Assessing the impact of egg sweating on Salmonella Enteritidis penetration into shell eggs.

Salmonella Enteritidis (SE) prevalence in eggs is a major concern to the egg industry. Some research has shown that egg sweating can increase Salmonella penetration into egg contents when refrigerated eggs are moved to a warmer temperature. This occurs when eggs are tempered before wash, to minimize thermal cracks. The effect of egg sweating on SE penetration into shell eggs over a 6 week storage period at 4°C was assessed. A 2 × 2 factorial of SE inoculation and egg sweating was utilized. Treatments included (SES) nalidixic acid (NA)-resistant SE inoculated and sweated, (SENS) NA-resistant SE inoculated and not sweated, (NSES) buffered peptone water (BPW) inoculated and sweated, and (NSENS) BPW inoculated and not sweated. Eggs were inoculated with 108 SE. Eggs formed condensation for approximately 17 min in a 32°C incubator. Shell rinse, shell emulsion, and egg contents were sampled then enumerated and assessed for prevalence of SE over a 6 wk storage period at 4°C. After wk 1, the SENS shell rinse had higher SE counts (0.32 log10 CFU/mL) than the other 3 treatments, where no SE was enumerated. A significant week by treatment interaction was found for the shell rinse SE detection (P < 0.05). In subsequent weeks, no SE counts were obtained from the egg shell rinse, shell emulsion, or egg contents. The SENS shell rinses had significantly higher SE prevalence than the SES rinses in weeks 1 (100% vs. 34.3%), 2 (57.6% vs. 22.2%), and 3 (38.2% vs. 11.1%) (P < 0.05). In samples from weeks 4, 5, and 6, there was no difference in SE prevalence between SES and SENS. Egg sweating did not increase SE penetration into the shell emulsion across treatment or week (P < 0.05). The decreasing trend of SE prevalence obtained over the study period indicate that refrigeration is effective at inhibiting SE growth. These results indicate that egg sweating occurring under common US egg handling practices is not harmful to egg safety.

Colonization of internal organs by Salmonella serovars Heidelberg and Typhimurium in experimentally infected laying hens housed in enriched colony cages at different stocking densities.

Contaminated eggs produced by infected commercial laying flocks are often implicated as sources of human infections with Salmonella Enteritidis, but Salmonella serovars Heidelberg and Typhimurium have also been associated with egg-transmitted illness. Contamination of the edible contents of eggs is a consequence of the colonization of reproductive tissues in systemically infected hens. In recent years, the animal welfare implications of diverse poultry housing and management systems have been vigorously debated, but the food safety significance of laying hen housing remains uncertain. The present study evaluated the effects of 2 different bird stocking densities on the invasion of internal organs by Salmonella serovars Heidelberg and Typhimurium in groups of experimentally infected laying hens housed in colony cages enriched with perching and nesting areas. Laying hens were distributed at 2 different stocking densities (648 and 973 cm2/bird) into colony cages and (along with a group housed in conventional cages at 648 cm2/bird) orally inoculated with doses of 107 cfu of 2-strain cocktails of either Salmonella Heidelberg or Salmonella Typhimurium. At 5 to 6 d post-inoculation, hens were euthanized and samples of internal organs (cecum, liver, spleen, ovary, and oviduct) were removed for bacteriologic culturing. The overall frequency of Salmonella isolation from ceca after inoculation with strains of serovar Heidelberg (83.3%) was significantly (P < 0.001) greater than the corresponding value for strains of serovar Typhimurium (53.8%), whereas Salmonella was recovered significantly more often from both livers (85.2% vs. 53.7%; P < 0.0001) and spleens (78.7% vs. 56.5%; P = 0.0008) after inoculation with strains of serovar Typhimurium than strains of serovar Heidelberg. However, there were no significant differences (P > 0.05) between stocking densities or cage systems in the frequencies of isolation of either Salmonella serovar from any of the five sampled tissues. These results contrast with prior studies, which reported increased susceptibility to internal organ invasion by Salmonella Enteritidis among hens in conventional cages at higher stocking densities.

Metabolic parameters linked by phenotype microarray to acid resistance profiles of poultry-associated Salmonella enterica.

Phenotype microarrays were analyzed for 51 datasets derived from Salmonella enterica. The top 4 serotypes associated with poultry products and one associated with turkey, respectively Typhimurium, Enteritidis, Heidelberg, Infantis and Senftenberg, were represented. Datasets were partitioned initially into two clusters based on ranking by values at pH 4.5 (PM10 A03). Negative control wells were used to establish 90 respiratory units as the point differentiating acid resistance from sensitive strains. Thus, 24 isolates that appeared most acid-resistant were compared initially to 27 that appeared most acid-sensitive (24 × 27 format). Paired cluster analysis was also done and it included the 7 most acid-resistant and -sensitive datasets (7 × 7 format). Statistical analyses of ranked data were then calculated in order of standard deviation, probability value by the Student's t-test and a measure of the magnitude of difference called effect size. Data were reported as significant if, by order of filtering, the following parameters were calculated: i) a standard deviation of 24 respiratory units or greater from all datasets for each chemical, ii) a probability value of less than or equal to 0.03 between clusters and iii) an effect size of at least 0.50 or greater between clusters. Results suggest that between 7.89% and 23.16% of 950 chemicals differentiated acid-resistant isolates from sensitive ones, depending on the format applied. Differences were more evident at the extremes of phenotype using the subset of data in the paired 7 × 7 format. Results thus provide a strategy for selecting compounds for additional research, which may impede the emergence of acid-resistant Salmonella enterica in food.

Alternative Antimicrobial Commercial Egg Washing Procedures.

Commercial table eggs are washed prior to packaging. Standard wash procedures use an alkaline pH and warm water. If a cool water method could be developed that would still provide a microbiologically safe egg, the industry may save energy costs associated with water heating. Four wash procedures were evaluated for Salmonella reduction: pH 11 at 48.9°C (industry standard), pH 11 at ambient temperature (∼20°C), pH 6 at 48.9°C, and pH 6 at ambient temperature. Alkaline washes contained potassium hydroxide-based detergent, while pH 6 washes contained approximately 200 ppm of chlorine and a proprietary chlorine stabilizer (T-128). When eggs were inoculated by immersion in a cell suspension of Salmonella Enteritidis and Salmonella Typhimurium, all treatments resulted in a slight and similar reduction of Salmonella numbers (approximately 0.77 log CFU/ml of shell emulsion reduction). When eggs were inoculated by droplet on the shell surface, Salmonella counts were reduced by approximately 5 log CFU when washed with chlorine plus the chlorine stabilizer at both temperatures and with the alkaline wash at the high temperature. The reductions in Salmonella by these treatments were not significantly (P > 0.05) different from each other but were significantly (P < 0.05) more than the reduction observed for the 20°C alkaline treatment and 20°C control water treatments. Ambient temperature acidic washes reduced Salmonella contamination to the same degree as the standard pH 11 warm water wash and may be a viable option to reduce cost, increase shelf life, and slow pathogen growth in and on shell eggs.

Colonization of internal organs by Salmonella Enteritidis in experimentally infected laying hens housed in enriched colony cages at different stocking densities.

Epidemiologic analyses have linked the frequency of human infections with Salmonella enterica subspecies enterica serovar Enteritidis to the consumption of contaminated eggs and thus to the prevalence of this pathogen in commercial egg-laying flocks. Contamination of the edible contents of eggs by Salmonella Enteritidis is a consequence of the colonization of reproductive tissues in systemically infected hens. The animal welfare implications of laying hen housing systems have been widely debated, but no definitive consensus has yet emerged about the food safety significance of poultry housing options. The present study sought to determine the effects of two different bird stocking densities on the invasion of internal organs by Salmonella Enteritidis in groups of experimentally infected laying hens housed in colony cages enriched with perching and nesting areas. In two trials, groups of laying hens were distributed at two different stocking densities into colony cages and (along with a group housed in conventional cages) orally inoculated with doses of 1.0 × 10(7) cfu of Salmonella Enteritidis. At 5 to 6 d post-inoculation, hens were euthanized and samples of internal organs were removed for bacteriologic culturing. For both trials combined, Salmonella Enteritidis was recovered at a significantly (P < 0.05) greater frequency from hens in enriched colony cages at the higher stocking density than at the lower density from livers (75.0% vs. 51.4%) and ovaries (51.4% vs. 30.6%). However, spleens from hens in enriched colony cages at the higher stocking density were significantly less often positive for Salmonella Enteritidis than from hens in conventional cages at that same density (90.3% vs. 68.1%). These results suggest that stocking density can influence the susceptibility of hens to Salmonella Enteritidis, but other housing systems parameters may also contribute to the outcome of infections.

Inactivation of Salmonella in Shell Eggs by Hot Water Immersion and Its Effect on Quality.

Thermal inactivation kinetics of heat resistant strains of Salmonella Enteritidis in shell eggs processed by hot water immersion were determined and the effects of the processing on egg quality were evaluated. Shell eggs were inoculated with a composite of heat resistant Salmonella Enteritidis (SE) strains PT8 C405, 2 (FSIS #OB030832), and 6 (FSIS #OB040159). Eggs were immersed in a circulating hot water bath for various times and temperatures. Come-up time of the coldest location within the egg was 21 min. SE was reduced by 4.5 log at both hot water immersion treatments of 56.7 C for 60 min and 55.6 °C for 100 min. Decimal reduction times (D-values) at 54.4, 55.6, and 56.7 °C were 51.8, 14.6, and 9.33 min, respectively. The z-value was 3.07 °C. Following treatments that resulted in a 4.5 log reduction (56.7 °C/60 min and 55.6 °C/100 min), the surviving population of SE remained static during 4 wk of refrigerated storage. After processing under conditions resulting in 4.5 log reductions, the Haugh unit and albumen height significantly increased (P < 0.01) and yolk index significantly decreased (P < 0.05). The shell dynamic stiffness significantly increased (P < 0.05), while static compression shell strength showed no significant difference (P < 0.05). Vitelline membrane strength significantly increased (P < 0.05); although, no significant difference (P < 0.05) was observed in vitelline membrane elasticity. In summary, the hot water immersion process inactivated heat resistant SE in shell eggs by 4.5 log, but also significantly affected several egg quality characteristics.