Refrigeration of eggs influences the virulence of Salmonella Typhimurium.

Salmonella Typhimurium is a human pathogen associated with eggs and egg-derived products. In Australia, it is recommended that eggs should be refrigerated to prevent condensation that can enhance bacterial penetration across the eggshell. Except for the United States, the guidelines on egg refrigeration are not prescriptive. In the current study, in-vitro and in-vivo experiments were conducted to understand the role of egg storage temperatures (refrigerated vs ambient) on bacterial load and the virulence genes expression of Salmonella Typhimurium. The in-vitro egg study showed that the load of Salmonella Typhimurium significantly increased in yolk and albumen stored at 25 °C. The gene expression study showed that ompR, misL, pefA, spvA, shdA, bapA, and csgB were significantly up-regulated in the egg yolk stored at 5 °C and 25 °C for 96 h; however, an in-vivo study revealed that mice infected with egg yolk stored at 25 °C, developed salmonellosis from day 3 post-infection (p.i.). Mice fed with inoculated egg yolk, albumen, or eggshell wash stored at refrigerated temperature did not show signs of salmonellosis during the period of the experiment. Data obtained in this study highlighted the importance of egg refrigeration in terms of improving product safety.

Salmonella Enteritidis foodborne infection induces altered placental morphometrics in the murine model.

INTRODUCTION: Salmonella foodborne disease during pregnancy causes a significant fetal loss in domestic livestock and preterm birth, chorioamnionitis and miscarriage in humans. These complications could be associated with alterations in placental structure. This study was aimed to determine how a low dose of Salmonella Enteritidis during late gestation affects placental histomorphometric in mice. METHODS: We used a self-limiting enterocolitis murine model. BALB/c pregnant animals received a low dose of Salmonella Enteritidis (3-4 x 102 CFU/mouse) on gestational day (GD) 15. At day 3 post infection bacterial loads, serum cytokines expression and placental histomorphometrics parameters were analyzed. RESULTS: We found that a sub-lethal infection with Salmonella induced a significant drop in fetal weight -to-placental weight-ratio and an increase in the placental coefficient. After bacterial inoculation maternal organs were colonized, inducing placental morphometric alterations, including increased placental thickness, reduced surface area, and diminished major and minor diameters. Also, foci of necrosis accompanied by acute leukocyte infiltration in decidual zone, reduction of vascular spaces and vascular congestion in labyrinth zone, were also evident in placentas from infected females on GD 18. Our data shows that placentas from infected mothers are phenotypically different from control ones. Furthermore, expression of IFN-gamma and IL-6 was up regulated in response to Salmonella in maternal serum. DISCUSSION: Our findings demonstrate that a low dose of Salmonella during late gestation alters the placental morphometry leading to negative consequences on pregnancy outcome such as significant reduction in fetal body weight.

Salmonella versus the Microbiome.

A balanced gut microbiota contributes to health, but the mechanisms maintaining homeostasis remain elusive. Microbiota assembly during infancy is governed by competition between species and by environmental factors, termed habitat filters, that determine the range of successful traits within the microbial community. These habitat filters include the diet, host-derived resources, and microbiota-derived metabolites, such as short-chain fatty acids. Once the microbiota has matured, competition and habitat filtering prevent engraftment of new microbes, thereby providing protection against opportunistic infections. Competition with endogenous Enterobacterales, habitat filtering by short-chain fatty acids, and a host-derived habitat filter, epithelial hypoxia, also contribute to colonization resistance against Salmonella serovars. However, at a high challenge dose, these frank pathogens can overcome colonization resistance by using their virulence factors to trigger intestinal inflammation. In turn, inflammation increases the luminal availability of host-derived resources, such as oxygen, nitrate, tetrathionate, and lactate, thereby creating a state of abnormal habitat filtering that enables the pathogen to overcome growth inhibition by short-chain fatty acids. Thus, studying the process of ecosystem invasion by Salmonella serovars clarifies that colonization resistance can become weakened by disrupting host-mediated habitat filtering. This insight is relevant for understanding how inflammation triggers dysbiosis linked to noncommunicable diseases, conditions in which endogenous Enterobacterales expand in the fecal microbiota using some of the same growth-limiting resources required by Salmonella serovars for ecosystem invasion. In essence, ecosystem invasion by Salmonella serovars suggests that homeostasis and dysbiosis simply represent states where competition and habitat filtering are normal or abnormal, respectively.

Salmonella secretion systems: Differential roles in pathogen-host interactions.

The bacterial genus Salmonella includes a large group of food-borne pathogens that cause a variety of gastrointestinal or systemic diseases in hosts. Salmonella use several secretion devices to inject various effectors targeting eukaryotic hosts, or bacteria. In the past few years, considerable progress has been made towards understanding the structural features and molecular mechanisms of the secretion systems of Salmonella, particularly regarding their roles in host-pathogen interactions. In this review, we summarize the current advances about the main characteristics of the Salmonella secretion systems. Clarifying the roles of the secretion systems in the process of infecting various hosts will broaden our understanding of the importance of microbial interactions in maintaining human health and will provide information for developing novel therapeutic approaches.

Genetic Ablation of Butyrate Utilization Attenuates Gastrointestinal Salmonella Disease.

Salmonella enterica serovar (S.) Typhi is an extraintestinal pathogen that evolved from Salmonella serovars causing gastrointestinal disease. Compared with non-typhoidal Salmonella serovars, the genomes of typhoidal serovars contain various loss-of-function mutations. However, the contribution of these genetic differences to this shift in pathogen ecology remains unknown. We show that the ydiQRSTD operon, which is deleted in S. Typhi, enables S. Typhimurium to utilize microbiota-derived butyrate during gastrointestinal disease. Unexpectedly, genetic ablation of butyrate utilization reduces S. Typhimurium epithelial invasion and attenuates intestinal inflammation. Deletion of ydiD renders S. Typhimurium sensitive to butyrate-mediated repression of invasion gene expression. Combined with the gain of virulence-associated (Vi) capsular polysaccharide and loss of very-long O-antigen chains, two features characteristic of S. Typhi, genetic ablation of butyrate utilization abrogates S. Typhimurium-induced intestinal inflammation. Thus, the transition from a gastrointestinal to an extraintestinal pathogen involved discrete genetic changes, providing insights into pathogen evolution and emergence.

Outbreak investigation and case-control study: penta-resistant Salmonella Typhimurium DT104 associated with biltong in London in 2008.

In August 2008 an outbreak of Salmonella Typhimurium DT104 occurred in South West London. Sixteen cases were identified with a particular multilocus variable number tandem repeat analysis (MLVA) pattern. In a matched case-control study 14 primary cases were included. These were defined as individuals with gastrointestinal symptoms and Salmonella Typhimurium DT104 isolated from a stool specimen, with a characteristic antibiotic resistance profile and MLVA pattern, and diagnosed in a local laboratory. Four controls per case were matched on age, gender and area of residence. Cases were 26 times more likely than controls to have eaten beef biltong, a South African speciality meat product (odds ratio 25·83, 95% confidence interval 4·92­135·59, P < 0·01). Although environmental investigation failed to identify Salmonella in the food product we conclude that beef biltong consumption led to this outbreak. This conclusion has importance in informing the ongoing risk assessment relating to uncontrolled foodstuffs.

Utility of clinical-epidemiological profiles in outbreaks of foodborne disease, Catalonia, 2002 through 2006.

The objective of this study was to evaluate the use of clinical-epidemiological profiles for classifying non-laboratory-confirmed outbreaks of foodborne disease (FBD) in Catalonia between 2002 and 2006 and for elucidating associations among factors contributing to these outbreaks. A total of 275 nonfamily outbreaks were studied, of which 190 (69.1%) were laboratory confirmed and 85 (30.9%) were not. In 176 (92.6%) of laboratory-confirmed outbreaks and 69 (81.2%) of non-laboratory-confirmed outbreaks, information was obtained on contributing factors (P = 0.009). In 72% of non-laboratory-confirmed outbreaks, the etiology was assigned by using clinical-epidemiological profiles; thus, 93% of outbreaks eventually were associated with an etiology. In laboratory-confirmed outbreaks, poor personal hygiene was positively associated with norovirus (odds ratio [OR], 2.69; 95% confidence interval [CI], 1.47 to 4.89; P = 0.0007) and negatively associated with Salmonella and Campylobacter (OR, 0.54; 95% CI, 0.33 to 0.89; P = 0.01), and an unsafe source was positively associated with Salmonella and Campylobacter (OR, 4.07; 95% CI, 1.72 to 10.09; P = 0.001) and negatively associated with norovirus (OR, 0.14; 95% CI, 0.04 to 0.58; P = 0.001). No differences were found among contributing factors associated with outbreaks with a laboratory-confirmed etiology and those associated with outbreaks with an etiology assigned according to the clinical-epidemiological profiles. Clinical-epidemiological profiles are useful for determining what prevention and control strategies are appropriate to the agents involved in each community and for designing outbreak investigations.

Outbreaks where food workers have been implicated in the spread of foodborne disease. Part 4. Infective doses and pathogen carriage.

In this article, the fourth in a series reviewing the role of food workers in foodborne outbreaks, background information on the presence of enteric pathogens in the community, the numbers of organisms required to initiate an infection, and the length of carriage are presented. Although workers have been implicated in outbreaks, they were not always aware of their infections, either because they were in the prodromic phase before symptoms began or because they were asymptomatic carriers. Pathogens of fecal, nose or throat, and skin origin are most likely to be transmitted by the hands, highlighting the need for effective hand hygiene and other barriers to pathogen contamination, such as no bare hand contact with ready-to-eat food. The pathogens most likely to be transmitted by food workers are norovirus, hepatitis A virus, Salmonella, Shigella, and Staphylococcus aureus. However, other pathogens have been implicated in worker-associated outbreaks or have the potential to be implicated. In this study, the likelihood of pathogen involvement in foodborne outbreaks where infected workers have been implicated was examined, based on infectious dose, carriage rate in the community, duration of illness, and length of pathogen excretion. Infectious dose estimates are based on volunteer studies (mostly early experiments) or data from outbreaks. Although there is considerable uncertainty associated with these data, some pathogens appear to be able to infect at doses as low as 1 to 100 units, including viruses, parasites, and some bacteria. Lengthy postsymptomatic shedding periods and excretion by asymptomatic individuals of many enteric pathogens is an important issue for the hygienic management of food workers.

The use of clinical profiles in the investigation of foodborne outbreaks in restaurants: United States, 1982-1997.

Improving the efficiency of outbreak investigation in restaurants is critical to reducing outbreak-associated illness and improving prevention strategies. Because clinical characteristics of outbreaks are usually available before results of laboratory testing, we examined their use for determining contributing factors in outbreaks caused by restaurants. All confirmed foodborne outbreaks reported to the Centers for Disease Control and Prevention (CDC) from 1982 to 1997 were reviewed. Clinical profiles were developed based on outbreak characteristics. We compared the percentage of contributing factors by known agent and clinical profile to their occurrence in outbreaks of unclassified aetiology. In total, 2246 foodborne outbreaks were included: 697 (31%) with known aetiology and 1549 (69%) with aetiology undetermined. Salmonella accounted for 65% of outbreaks with a known aetiology. Norovirus-like clinical profiles were noted in 54% of outbreaks with undetermined aetiology. Improper holding times and temperatures were associated with outbreaks caused by Clostridium perfringens, Bacillus cereus, Staphylococcus aureus, and Salmonella, and also with outbreaks of undetermined aetiology that fitted diarrhoea-toxin and vomiting-toxin clinical profiles. Poor personal hygiene was associated with norovirus, Shigella, and Salmonella, and also with outbreaks that fitted norovirus-like and vomiting-toxin clinical profiles. Contributing factors were similar for outbreaks with known aetiology and for those where aetiology was assigned by corresponding clinical profile. Rapidly categorizing outbreaks by clinical profile, before results of laboratory testing are available, can help identification of factors which contributed to the occurrence of the outbreak and will promote timely and efficient outbreak investigations.

Mechanism of infection of a human isolate Salmonella (3,10:r:-) in chicken ileum: ultrastructural study.

BACKGROUND & OBJECTIVE: Originally isolated from severe human food-poisoning cases, Salmonella (3,10:r:-), a monophasic variety of otherwise diphasic serotypes such as S. weltevreden and S. simi, causes serious infections in man, animals and poultry. Mechanism of infection of this versatile and deadly organism is important to understand for its control. The objective of this study was to enhance our understanding of infection of Salmonella (3,10:r:-) in vivo at cellular level. METHODS: Aliquots of 10(9) cfu of Salmonella (3,10:r:-) organisms were injected intra-ileally in 24 h pre-fasted 3 month old broiler chickens by standard ligated ileal loop method. After 18 h, the fluid accumulated in the ileum was drained and small tissue pieces were fixed in 2.5 per cent buffered (pH 7) glutaraldehyde and subsequently in 1 per cent aqueous osmium tetraoxide. Ultra-thin sections of araldite-embedded tissue pieces were examined under transmission electron microscope operated at 100 KV after staining with uranyl acetate and lead citrate. RESULTS: Over 70 per cent of salmonellae interacting within 300 nm with ileal epithelial cells developed numerous surface blebs of periplasmic extensions designated "periplasmic organelles" (POs). Large sized POs were apparently pinched off as outer membrane vesicles (OMVs), 50-90 nm in diameter. Type III secretion needle complex-like "rivet complexes" (RCs) were viewed to rivet the bacterial outer and inner membranes together, allowing only pockets of periplasm to expand/inflate in order to liberate OMVs. Many OMVs were found visibly docked on the plasma membrane of host epithelial cells. The invading organisms appeared to leave the epithelial cells so as to find entry into the lymphatic vessels, where, they again appeared to be closely interacting with ileal macrophages, by forming numerous POs and concomitantly liberating OMVs. Inside the cytoplasm of macrophages, numerous tight phagosomes were seen, each containing two organisms. The final stage appeared to contain replicated salmonellae, four in each loose phagosome and, at the same time, macrophages also showed signs of apoptotic disintegration, culminating in the release of replicated salmonellae. INTERPRETATION & CONCLUSION: Outer membrane vesicles released from a fiercely virulent human isolate, Salmonella 3,10:r:- pathogens have been implicated in translocating biochemical signals from the host-interactive organisms to the eukaryotic cells at both stages of invasion leading to epithelial cell and macrophage infection in vivo, in the chicken ileal model. A comprehensive cellular mechanism at ultrastructural level is outlined for typhoid-like Salmonella infections caused by this humans-infecting organism.

Reactive joint symptoms following an outbreak of Salmonella typhimurium phage type 135a.

OBJECTIVE: To quantify the incidence and clinical features of reactive arthritis (ReA) developing in a cohort exposed to an outbreak of Salmonella typhimurium phage type 135a, and factors affecting host susceptibility to ReA. METHODS: A screening questionnaire was mailed to 493 patients with confirmed Salmonella infection. Musculoskeletal symptoms and extraarticular manifestations of ReA were quantified. Positive responders with joint pain were invited to participate further, with a detailed history, examination, and investigations including HLA-B27 status. RESULTS: A total of 261/461 (57%) subjects responded to the questionnaire, with 23/54 adults (43%) and 41/207 children (20%) reporting joint symptoms. Although joint pains were less common in children compared with adults, those children affected usually had eye (34%) or mucocutaneous (37%) symptoms. The incidence of ReA was 14.6%, with adults more frequently affected (24%) than children (12%). This may be an underestimate given the large proportion of children involved. Associated clinical features were similar to previous studies, with the distribution of arthritis affecting the lower limbs predominantly in an oligoarticular pattern, as were the extraarticular manifestations and enthesopathy. We found 17% of subjects were HLA-B27 positive, and 55% were still symptomatic after 6 months. CONCLUSION: In an Australian cohort study of a S. typhimurium phage type 135a outbreak, joint symptoms were common, affecting 25% of subjects. The incidence of ReA of 14.6% and the clinical features were comparable to previous studies. There was a small effect of HLA-B27 status on the development of ReA.

Life-threatening infantile diarrhea from fluoroquinolone-resistant Salmonella enterica typhimurium with mutations in both gyrA and parC.

Salmonella Typhimurium DT12, isolated from a 35-day-old infant with diarrhea, was highly resistant to ampicillin, tetracycline, chloramphenicol, streptomycin, gentamycin, sulfamethoxazole/trimethoprim, nalidixic acid, and fluoroquinolones. The patient responded to antibiotic therapy with fosfomycin. Multidrug-resistance may become prevalent in Salmonella infections in Japan, as shown in this first case of a patient infected with fluoroquinolone-resistant Salmonella.

Septicemic salmonellosis in two cats fed a raw-meat diet.

Salmonella gastroenteritis and septicemia were diagnosed in two cats presented for necropsy. Both cats resided in the same household and were fed a home-prepared, raw meat-based diet. Salmonella was isolated from multiple organs in both cats and from samples of raw beef incorporated into the diet fed to one of the cats. Subtyping of the bacterial isolates yielded Salmonella newport from one cat and from the diet it had been fed. This report provides evidence that the practice of feeding raw meat-based diets to domestic cats may result in clinical salmonellosis.

Interferon-gamma plays a critical role in intestinal immunity against Salmonella typhimurium infection.

Salmonella bacteria are a major cause of food-borne infectious diarrhoea and there is great interest in understanding the pathogenesis of Salmonella infection and in vaccine development. Potential vaccines include the aromatic mutants of S. typhimurium. Such non-lethal Aro mutants have also been useful for studying Salmonella infections in mouse models. Studies of systemic infection, using these Aro mutants, in both normal and cytokine gene knockout mice, indicate that interferon-gamma (IFN-gamma) plays a key role in the resolution of Salmonella infection. The present studies have investigated the outcome of oral infection in mice with attenuated Salmonella because this infection route mimics natural infection in humans. In IFN-gamma gene knockout (IFN-gamma-/-) mice, intestinal immunity was impaired and oral challenge resulted in disseminated septicaemia 2 weeks later. No dissemination of infection was seen in wild-type mice. In wild-type mice, both CD4 and CD8 cell numbers increased in the gut following Salmonella challenge, together with increased expression of major histocompatibility complex (MHC) II and vascular cell adhesion molecule-1 (VCAM-1). No such changes were seen in IFNgamma-/- mice. Following oral challenge, antilipopolysaccharide (LPS) and antiphosphoryl choline antibodies increased by more than 100-fold in both serum and faecal pellet extracts of IFNgamma-/- mice compared with wild-type mice. Our data show that IFN-gamma production is essential for resolution of enteric Salmonella infection and that antibody has little effect on this process.

Pathology of the alimentary tract in Salmonella typhimurium food poisoning.

The pathology of the alimentary tracts of nine patients dying of Salmonella typhimurium infection is reviewed. Two patients had previous gastric operations, supporting previous reports that such patients are more susceptible to food poisoning. Four had no parietal (oxyntic) cells in the gastric mucosa, suggesting hypo- or anacidity. Only one had acute gastritis. None had acute enteritis, but in half of the patients, subtle histological changes suggested an 'enteropathy'. Acute diffuse colitis with abundant crypt abscesses, without stromal abscesses in the lamina propria, was the most constant finding and reparative features started very early, and occurred in later deaths. Under ideal circumstances this crypt abscess is readily distinguished from that of idiopathic ulcerative colitis, but can be confused with the crypt abscess of acute bacillary (sonne) dysentery. While the florid colonic changes may have settled in the late deaths, active inflammation is commonly present in the appendix mucosa on histology. The pathology of the alimentary tract in S typhimurium infection differs from that of S typhi and S paratyphi infections. There is little evidence of gastroenteritis, although subtle changes occur in the stomach and small intestine. The features are those of acute diffuse colitis with histological appendicitis, distinguishable from idiopathic ulcerative colitis.