Egg-associated Outbreaks Research Articles

Prevalence and Genetic Analysis of Salmonella enterica from a Cross-sectional Survey of the New Zealand Egg Production Environment

Epidemiological evidence suggests that Salmonella on New Zealand eggs is not an important pathway for human salmonellosis. However, robust nationally representative data for Salmonella contamination of eggs is not available to support this. To better understand the exposure of New Zealand commercial eggs to Salmonella, a cross-sectional survey collected data on prevalence and serotypes of Salmonella in the feed, laying sheds (feces, dust, and boot or manure belt swabs), and packhouses (egg contact surfaces) of New Zealand commercial egg layer farms. Salmonella was not detected on 16 of 28 surveyed farms, and 4 farms had only one positive sample. Of the 43 (13.3%) of 323 Salmonella-positive samples, dust samples had the highest prevalence (19 of 67, 28.4%), followed by boot or manure belt swabs (11 of 67, 16.4%), feces (7 of 67, 10.4%), packhouse egg contact surfaces (5 of 87, 5.7%), and feed (1 of 33, 3.0%). A significantly higher prevalence was from caged (33 of 75, 44.0%; P < 0.001) compared with cage-free (4 of 126, 3.2%) systems, yet multiple practices differ between laying systems, which could influence prevalence. Salmonella-positive packhouse samples were only identified on the three farms with the highest laying shed prevalence, and isolates were genetically related (as determined by single nucleotide polymorphism analyses) suggesting cross-contamination between the laying shed and packhouse surfaces. Serotypes isolated included Salmonella Infantis, Salmonella Thompson, Salmonella Typhimurium, Salmonella Anatum, and Salmonella Mbandaka. Importantly, Salmonella Enteritidis, which causes egg-associated outbreaks internationally, was not isolated. Genomic comparisons of isolates supported the presence of a common contamination source in the shed and farm environments rather than multiple sporadic contamination events. This survey establishes a benchmark of Salmonella prevalence and types in the New Zealand egg production environment and provides a reference point for assessing the impact of changes to practices on Salmonella prevalence.

High Intensity Ultrasound for Salmonella Enteritidis Inactivation in Culture and Liquid Whole Eggs.

High intensity ultrasound (HIU) continues to be researched as a nonthermal inactivation technology of appeal to food manufacturers. The advantages of HIU include maintenance of product quality, freshness, product homogenization, along with simultaneous inactivation of pathogens. Besides, it is simple, relatively inexpensive, and easily adaptable to most processing environments. This study investigated the effect of HIU for Salmonella Enteritidis inactivation in culture and liquid whole eggs (LWEs) to decrease egg-associated outbreaks. Overnight S. Enteritidis cultures and spiked LWE (both at 8 log CFU/mL) were treated with 20-kHz HIU for 0, 1, 5, 10, and 30min (n=6) in a temperature-controlled system, not to exceed 20 °C, and replicated thrice. At each time point, samples were enumerated on XLT4 agar and morphologically analyzed using scanning electron microscopy, with measurements of color and rheological properties. Our results revealed significant reduction of healthy S. Enteritidis cells up to 3.6log CFU/mL and 2.3log CFU/25mL after HIU treatment of merely 10min of overnight culture and 30min in LWE, respectively (P < 0.05). After 5 and 10-min HIU treatment, significant reduction of 1.4-log CFU/25mL healthy S. Enteritidis in LWE was obtained (P<0.05). Even at 1-min exposure time, HIU showed significant 1.9 log CFU/mL reduction of cultures (P<0.05); however, no log-reduction was observed in LWE after 1 min. Scanning electron micrographs showed increased cell structural damage using longer HIU exposure. For product color changes, lower redness and yellowness of LWE were observed visually and instrumentally after 5-min HIU treatment (P<0.05). The rheological properties of LWE measured at 0 to 200s-1 shear rate, showed that shear stress of HIU-treated LWEs decreased after 5-min HIU exposure, but increased after 30-min treatment. This study demonstrated that HIU shows promise for rapid Salmonella control in LWE and other liquid foods, as an alternative inactivation method for use in hurdle approaches.

Salmonella Typhimurium and Outbreaks of Egg-Associated Disease in Australia, 2001 to 2011.

Salmonellosis is a significant public health problem, with eggs frequently identified as a food vehicle during outbreak investigations. Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Enteritidis are the two most frequently identified causes of egg-associated disease in industrialized countries. In Australia, a comprehensive review of egg-associated outbreaks has not been previously undertaken. Using a national register of foodborne outbreaks, we undertook a descriptive review of egg-associated outbreaks between 2001 and 2011. Included in our review was additional detail from the findings of trace back investigations conducted to the farm level. Evidence classifications were developed and applied to each outbreak based on descriptive and analytical epidemiology, food safety investigations, and microbiological testing of clinical, food, and trace back-derived samples. Over the study period, the proportion of foodborne Salmonella outbreaks linked to eggs increased significantly (p < 0.001). In total, 166 outbreaks were identified, with 90% caused by Salmonella Typhimurium. The majority of outbreaks were linked to commercial food providers, with raw egg use the major contributing factor. These events resulted in more than 3200 cases, more than 650 hospitalizations, and at least 4 deaths. Fifty-four percent of investigations used analytical epidemiology, food microbiology, and trace back microbiology to demonstrate links between human illness and eggs. Trace back investigations identified S. enterica indistinguishable from outbreak-associated clinical or food samples on 50% of sampled egg farms. Effective control of egg-associated salmonellosis remains a challenge in Australia, with Salmonella Typhimurium dominating as the causative serotype in outbreak events. Although outbreaks predominantly occur in the settings of restaurants, the high recovery rate of indistinguishable Salmonella on epidemiologically implicated egg farms suggests that further efforts to minimize infection pressure at the primary production level are needed in Australia.

Salmonella Typhimurium and Salmonella Sofia: Growth in and Persistence on Eggs under Production and Retail Conditions.

Salmonellosis in Australia has been linked to eggs and egg products with specific serotypes associated with outbreaks. We compared attachment to and survival on egg shells and growth in eggs of two Salmonella serotypes, an egg outbreak associated Salmonella Typhimurium and a non-egg-associated Salmonella enterica ssp. II 1,4,12,27:b:[e,n,x] (S. Sofia). Experiments were conducted at combinations of 4, 15, 22, 37 and 42°C. No significant differences occurred between the serotypes in maximum growth rates, which were significantly greater (P < 0.001) in egg yolk (0.427 log10 CFU/mL/h) compared to whole egg (0.312 log10 CFU/mL/h) and egg white (0.029 log10 CFU/mL/h). Attachment to egg shells varied by time (1 or 20 min) and temperature (4, 22 and 42°C), with S. Typhimurium isolates attaching at higher levels (P < 0.05) than S. Sofia after 1 min at 4°C and S. Typhimurium ATCC 14028 attaching at higher (P < 0.05) levels at 22°C. Survival on egg shells was not significantly different across isolates. Salmonella serotypes behaved similarly regarding growth in egg contents, attachment to egg shells and survival on eggs, indicating that other factors more likely contributed to reasons for S. Typhimurium being implicated in multiple egg-associated outbreaks.

Evidence of microevolution of Salmonella Typhimurium during a series of egg-associated outbreaks linked to a single chicken farm

BackgroundThe bacterium Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the most frequent causes of foodborne outbreaks of gastroenteritis. Between 2005–2008 a series of S. Typhimurium outbreaks occurred in Tasmania, Australia, that were all traced to eggs originating from a single chicken farm. We sequenced the genomes of 12 isolates linked to these outbreaks, in order to investigate the microevolution of a pathogenic S. Typhimurium clone in a natural, spatiotemporally restricted population.ResultsThe isolates, which shared a phage type similar to DT135 known locally as 135@ or 135a, formed a clade within the S. Typhimurium population with close similarity to the reference genome SL1334 (160 single nucleotide polymorphisms, or SNPs). Ten of the isolates belonged to a single clone (<23 SNPs between isolate pairs) which likely represents the population of S. Typhimurium circulating at the chicken farm; the other two were from sporadic cases and were genetically distinct from this clone. Divergence dating indicated that all 12 isolates diverged from a common ancestor in the mid 1990s, and the clone began to diversify in 2003–2004. This clone spilled out into the human population several times between 2005–2008, during which time it continued to accumulate SNPs at a constant rate of 3–5 SNPs per year or 1x10-6 substitutions site-1 year-1, faster than the longer-term (~50 year) rates estimated previously for S. Typhimurium. Our data suggest that roughly half of non-synonymous substitutions are rapidly removed from the S. Typhimurium population, after which purifying selection is no longer important and the remaining substitutions become fixed in the population. The S. Typhimurium 135@ isolates were nearly identical to SL1344 in terms of gene content and virulence plasmids. Their phage contents were close to SL1344, except that they carried a different variant of Gifsy-1, lacked the P2 remnant found in SL1344 and carried a novel P2 phage, P2-Hawk, in place SL1344’s P2 phage SopEϕ. DT135 lacks P2 prophage. Two additional plasmids were identified in the S. Typhimurium 135@ isolates, pSTM2 and pSTM7. Both plasmids were IncI1, but phylogenetic analysis of the plasmids and their bacterial hosts shows these plasmids are genetically distinct and result from independent plasmid acquisition events.ConclusionsThis study provides a high-resolution insight into short-term microevolution of the important human pathogen S. Typhimurium. It indicates that purifying selection occurs rapidly in this population (≤6 years) and then declines, and provides an estimate for the short-term substitution rate. The latter is likely to be more relevant for foodborne outbreak investigation than previous estimates based on longer time scales.

Draft Genome Sequences for Ten Salmonella enterica Serovar Typhimurium Phage Type 135 Variants

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a common cause of gastroenteritis in humans. Here, we report the draft genome sequences of 10 isolates of an S. Typhimurium phage type 135 variant that is linked to egg-associated outbreaks in Tasmania, Australia.

Operational practices associated with foodborne disease outbreaks in the catering industry in England and Wales.

Catering businesses continue to be the most common setting for foodborne disease outbreaks. In a study of catering businesses in England and Wales, operational practices relating to the supply, preparation, and service of food in 88 businesses associated with outbreaks were compared with those practices at 88 control businesses. Operational practices did not differ significantly between case and control businesses but larger small medium-size enterprise (SME) businesses were more likely to be associated with foodborne disease outbreaks than were micro-SME businesses. Businesses associated with outbreaks of Salmonella infection were less likely to use local or national suppliers but instead used regional suppliers, especially for eggs. This practice was the only significantly independent operational practice associated with outbreaks of Salmonella infection. Regional egg suppliers also were more likely to be used by businesses associated with outbreaks attributed to food vehicles containing eggs. Businesses associated with egg-associated outbreaks were less likely to use eggs produced under an approved quality assurance scheme, suggesting that the underlying risk associated with using regional suppliers may relate to the use of contaminated eggs.

Determination of Close Genetic Relatedness of the Major Salmonella enteritidis Phage Types by Pulsed-Field Gel Electrophoresis and DNA Sequence Analysis of Several Salmonella Virulence Genes

Salmonella enteritidis (SE) is an important cause of egg-associated outbreaks in both Europe and the United States. Phage typing has become an important epidemiologic tool in identifying the source of outbreaks. Limitations of phage typing have become apparent with wholesale egg distributors that have multiple suppliers in an area where a particular phage type is endemic. Several different molecular typing methods were evaluated for their discriminatory power to identify genetic differences among different SE phage types isolated in Europe and the United States. Pulsed-field gel electrophoresis (PFGE) identified a single DNA pattern among the different SE phage types. Comparison of the nucleotide sequence for several Salmonella virulence genes failed to identify a single nucleotide change in the gene sequences from most SE isolates, regardless of phage type. On the basis of these results, the different SE phage types appear to be genetically related or clonal. However, with primers 1283 and Opa4, it was possible to differentiate not only SE isolates from different geographic locations but those within a specific geographic locale as well by random amplified polymorphic DNA polymerase chain reaction. Any chance for discerning genetic differences among isolates will need to rely on molecular techniques other than PFGE.

A Flow-Injection System for Studying Heat Inactivation of Listeria monocytogenes and Salmonella enteritidis in Liquid Whole Egg

A flow-injection system was devised to mimic continuous flow-through pasteurization systems for laboratory thermal inactivation studies. Air bubbles were introduced into the sample stream to create separate moving segments (plugs) of liquid stream during pasteurization while residence time was adjusted by a combination of pump speed and column length. The method was used to obtain thermal inactivation data for Listeria monocytogenes Scott A and Salmonella enteritidis ATCC 13076 in liquid whole eggs at different temperatures and heating times. Thermal inactivation of L. monocytogenes using the capillary tube method (Zcap = 7.3°C) gave results comparable to those obtained with the flow-injection system (Zflow = 7.2°C). The flow-injection system also was used to examine thermal inactivation of S. enteritidis (SE) grown in either tryptic soy broth (TSB) or egg yolk medium (EYM) before inoculation into liquid whole egg (LWE). D-values were obtained by regression analysis and the data showed that SE grown in EYM gave D-values 15 to 120% higher than those obtained for SE grown in TSB. Thermal inactivation studies performed with S. enteritidis grown in commercial broth media may therefore inaccurately represent thermal resistance of S. enteritidis grown in liquid or shell raw egg as may occur in egg-associated outbreaks. The continuous flow-injection system described herein may be adapted to study continuous flow pasteurization processes not easily examined by the traditional capillary tube method.

Role of Egg Consumption in Sporadic Salmonella enteritidis and Salmonella typhimurium Infections in Minnesota

The incidence of Salmonella enteritidis infections has increased in the eastern United States, and consumption of undercooked eggs has been associated with outbreaks of S. enteritidis. In Minnesota, the incidence of S. enteritidis infections doubled from 1980 to 1990; however, no egg-associated outbreaks were identified. A case-control study was conducted to examine potential exposures for S. enteritidis and Salmonella typhimurium infections in Minnesota adults. Sporadic cases of S. enteritidis (odds ratio [OR], 5.2; 95% confidence interval [CI], 1.9-14.2; P = .003) and S. typhimurium infection (OR, 2.4; CI, 1.1-5.5; P = .03) were more likely to have consumed undercooked eggs or egg-containing foods during the 3 days before onset of illness compared to a similar reference period for controls. In addition, the extent to which eggs were cooked was directly associated with illness (chi 2 test for trend, P < .001). These findings demonstrate that eggs are important vehicles for S. enteritidis and S. typhimurium, even in the absence of recognized outbreaks.

Trends in Foodborne Salmonellosis Outbreaks: 1963–1975

Since 1963 651 outbreaks of human salmonellosis have been reported to the Center for Disease Control. The vehicle of transmission was identified in 463 (71%) of the reported outbreaks. The three most important vehicles have been poultry, meat, and eggs, which accounted for 99 (21%), 69 (15%), and 53 (11%) of the outbreaks respectively. Egg-associated outbreaks decreased markedly during the study period while meat-associated outbreaks (particularly beef) gradually increased and poultry-associated outbreaks showed no constant trend.