Most Salmonella enterica serovars are believed to have a cyclical lifestyle involving both host-associated and environment-associated, persistent phases. Their ability to persist in the environment increases the probability that they will be transmitted. Our hypothesis is that the genetic factors required for cellular aggregation and biofilm formation are important for host-to-host transmission. A link between biofilm formation, environmental persistence and transmissibility has not been observed, due to the lack of an appropriate model. We developed a murine model of Salmonella transmission allowing us to study the genetic factors involved in the transmission process. To test the role of aggregation and biofilm formation we used the ∆csgD mutant, which is deficient in both processes. We also engineered luciferase reporter strains of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) to track infection within a mouse population before the onset of clinical signs using bioluminescent imaging. We determined that mice shed high levels of Salmonella Typhimurium in their feces when pre-treated with streptomycin. To observe the transmission efficiency of Salmonella, we tracked their spread from infected mice to naive mice, and determined that Salmonella could be transmitted only after pre-treatment with streptomycin. We compared the shedding potential and colonization levels of mice challenged with either wild-type Salmonella Typhimurium or the ∆csgD mutant and determined them to be statistically similar when challenged separately. We found that wild-type Salmonella Typhimurium persisted in fecal pellets at higher levels than the ∆csgD mutant. We compared both the short- and long- transmission potential of the ∆csgD mutant to wild type Salmonella Typhimurium, and found that the mutant did not have a defect in either process. Though not observed in our model, we believe that environmental persistence and biofilm formation are important for the transmission of Salmonella due to its cyclical lifestyle. The model we generated remains useful to test the role of other genes in transmission. It can be further refined to more accurately mimic environmental transmission of Salmonella. Further understanding of the transition of Salmonella from infected hosts to the environment and back into new hosts will aid in reducing its environmental persistence and transmission.

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / abstract / Microbiology / Doctoral / Doctor of Philosophy

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In order to determine the effects that Maillard Reaction Products (MRP) have on Salmonella Typhimurium, growth rates and virulence expression, in the presence of Maillard reaction products, were observed, using the β−galactosidase Miller Assay and Reverse Transcription Polymerase Chain Reaction (RT-PCR). The presence of MRP compounds in liquid media caused no negative effect on the growth rate of Salmonella cells. However, the addition of MRP compounds at a 1% level in the media caused a significant increase in hilA expression in Salmonella Typhimurium, and the highest induction levels were observed in media supplemented with arginine and histidine-MRP compounds. There was no effect on the induction of hilA with the 0.5% addition of the MRP compounds in the amended media as shown by the Miller Assay. However, there was an effect seen when using the Real Time RT-PCR assay that resulted in the same levels of significance seen at 1.0% additions of MRPs being seen at the 0.5% level as well. Since rsmC was shown to be a constitutive gene that had continuous levels of expression in Salmonella based on cell number, Real-Time PCR was then used to assess the hilA expression of Salmonella Typhimurium under different oxygen, pH levels, and osmolarity conditions. The results under low oxygen indicate that the combination of low osmolarity and high pH have the highest inducing effect on hilA expression. The hilA response under the same media conditions and a high oxygen environment showed the same pattern of expression as those bacteria grown under a non-aerobic environment. The media with a pH of 8 and low osmolarity conditions had the greatest effect on the induction of hilA with none of the other media showing any significant effect. The relative expression of hilA did decrease for those bacteria grown under aerobic conditions versus those grown under low oxygen conditions.

The purpose of this study was to describe the herd antibody status to Salmonella Typhimurium as estimated from co-mingled milk samples and to describe the resulting geographical patterns found in Texas dairy herds. Bulk tank milk samples were collected from 852 Grade A dairies throughout Texas during the summer of 2001. An indirect enzyme-linked immunosorbent assay (ELISA) using S. Typhimurium lipopolysaccharide was performed with signal to noise ratios calculated for each sample. The ELISA ratio was used in fitting a theoretical variogram and kriging was used to develop a predicted surface for these ratios in Texas. A spatial process with areas of higher risk located in the panhandle and near Waller County was apparent. Lower risk areas included Atascosa, Cooke, Collin, Titus, Comanche and Cherokee Counties. Subsets representing large dairy sheds in northeast Texas, the Erath County area, and the Hopkins County area were also evaluated individually. Each result illustrated a spatial process with areas of low and high ELISA ratio predictions. Cluster analysis was performed for the entire state with cases defined as herds having milk ELISA ratios greater than or equal to 1.8. Using this cutoff, the prevalence of herds with positive bulk tank milk ELISAs was 4.3%. Significant clustering of cases was demonstrated by the Cuzick and Edward's test. The spatial scan statistic then identified the two most likely clusters located in and near the Texas Panhandle. This study demonstrated that the distribution of S. Typhimurium antibodies in bulk tank milk in Texas is describable by a spatial process. Knowledge of this process will help elucidate geospatial influences on the presence of S. Typhimurium in dairy herds and enhance our understanding of the epidemiology of salmonellosis.

The control of foodborne pathogens especially Salmonella and Campylobacter are of great concern to the commercial poultry industry. The control of these pathogens could be essential in the reduction of foodborne illness and deaths related to eggs and poultry meat. Previous studies have found that the presence or disappearance of Salmonella or Campylobacter is linked to various environmental and management-based factors, of which include vaccines used in the industry. Presently, we evaluated the effect of the infectious bronchitis virus (IBV) vaccine on the incidence of Salmonella or Campylobacter prevalence in broiler chicks. In the current study, a high vaccine dosage of IBV vaccine was associated with an increase the prevalence of Campylobacter during the first two weeks of age. Although in a previous study a high vaccine dose of IBV was linked in to increased prevalence of Salmonella, this was not seen in our study. In a subsequent trial, we also evaluated the potential cross-protection against three Salmonella serotypes of two-previously formulated vaccines when used in various dosage combinations. The combination vaccine was effective in reducing shedding of S. Enteritidis however reduction of S. Typhimurium and S. Hadar were not seen consistently. The vaccines were also shown to not significantly affect the body weights of the birds. Vaccines have been an essential component in the control of diseases within flocks in the commercial poultry industry. Ensuring the uniform application of IBV vaccine could help prevent and/or reduce the prevalence of Campylobacter in broiler flocks. The combination vaccine was effective against one serotype of Salmonella but further trials are needed to complete evaluate its potential as a vaccine that could be used in the poultry industry.

Salmonella Typhimurium continues to be a leading cause of human gastroenteritis worldwide. This organism is a facultative intracellular pathogen, meaning that it is able grow and reproduce within the host cell it inhabits. S. Typhimurium has the ability to invade and replicate within human intestinal epithelial cells, which in turn causes induced cell death or apoptosis. The human intestinal epithelial cells, HCT-8, were challenged with live, heat inactivated, and electron beam inactivated S. Typhimurium for various time points. Infected cell monolayers were collected for RNA extractions, and Real-time PCR was performed on the samples to analyze differential gene expression. Genes of the host cell that were expected to be differentially expressed were shortlisted and Real-Time PCR analysis was performed. Internalized Salmonella within the host cell was unable to be successfully visualized using fluorescent light microscopy. However, differential gene expression for a common transcriptional regulator and inflammatory chemokine were observed to be expressed significantly higher in response to e-beam inactivated Salmonella infection. Genes coding for extracellular and intracellular pattern-recognition receptors of the host cells were shown to be up-regulated in response to e-beam inactivated Salmonella infection at 4 and 24 hours, but were not statistically significant. Additional studies must be conducted to definitively confirm e-beam irradiated Salmonella has the ability to invade human host cells.