The prevalence of Escherichia coli O157:H7 fecal shedding in feedlot pens is affected by the water-to-cattle ratio: A randomized controlled trial.

Escherichia coli O157:H7 fecal shedding in feedlot cattle is common and is a public health concern due to the risk of foodborne transmission that can result in severe, or even fatal, disease in people. Despite a large body of research, few practical and cost-effective farm-level interventions have been identified. In this study, a randomized controlled trial was conducted to assess the effect of reducing the level of water in automatically refilling water-troughs on fecal shedding of E. coli O157:H7 in feedlot cattle. Pens in a feedlot in the Texas Panhandle were randomly allocated as control (total number: 17) or intervention (total number: 18) pens. Fecal samples (2,759 in total) were collected both at baseline and three weeks after the intervention, and tested for the presence of E. coli O157:H7 using immunomagnetic bead separation and selective culture. There was a strong statistical association between sampling date and the likelihood of a fecal sample testing positive for E. coli O157:H7. Pen was also a strong predictor of fecal prevalence. Despite accounting for this high level of clustering, a statistically significant association between reduced water levels in the trough and increased prevalence of E. coli O157:H7 in the feces was observed (Odds Ratio = 1.6; 95% Confidence Interval: 1.2-2.0; Likelihood Ratio Test: p = 0.02). This is the first time that such an association has been reported, and suggests that increasing water-trough levels may be effective in reducing shedding of E. coli O157:H7 in cattle feces, although further work would be needed to test this hypothesis. Controlling E. coli O157:H7 fecal shedding at the pre-harvest level may lead to a reduced burden of human foodborne illness attributed to this pathogen in beef.

Gold Nanocluster Containing Polymeric Microcapsules for Intracellular Ratiometric Fluorescence Biosensing.

A new approach to sensing and imaging hydrogen peroxide (H2O2) was developed using microcapsule-based dual-emission ratiometric luminescent biosensors. Bovine serum albumin-capped gold nanoclusters (BSA-AuNCs) sensitive to H2O2 were coencapsulated with insensitive FluoSpheres (FSs) within polymeric capsules fabricated via the layer-by-layer method. Under single-wavelength excitation, the microcapsule-based biosensors exhibited emission bands at ∼516 and ∼682 nm resulting from the FSs and BSA-AuNCs, respectively. The polyelectrolyte multilayers lining the microcapsules were effective in protecting BSA-AuNCs from the degradation catalyzed by proteases (chymotrypsin, trypsin, papain, and proteinase K) and subsequent luminescent quenching, overcoming a key limitation of prior BSA-AuNC-based sensing systems. The luminescent response of the sensors was also found to be independent of local changes in pH (5-9). Quenching of the AuNCs in the presence of H2O2 enabled the spectroscopic quantification and imaging of changes in H2O2 concentration from 0 to 1 mM. The microcapsule sensors were easily phagocytized by murine macrophage cells (RAW 264.7), were effective as intracellular H2O2 imaging probes, and were successfully used to detect local release of H2O2 in response to an external chemical stimulus.

Weaned pigs experimentally infected with Salmonella display sexually dimorphic innate immune responses without affecting pathogen colonization patterns.

Sexually dimorphic innate immune responses have been observed in several species, but have not been studied in response to a live pathogen challenge in pigs. This study aimed to elucidate sexually dimorphic innate immune responses along with Salmonella translocation patterns in newly weaned pigs orally inoculated with Salmonella. Newly weaned pigs (n = 8 gilts and 12 barrows; 6.2 ± 0.2 kg BW) were obtained from a commercial swine facility and were maintained in an environmentally-controlled facility in individual pens equipped with feeders and nipple waterers. Pigs were allowed ad libitum access to a commercial non-medicated starter ration and water throughout the study. On d 12 post-weaning, pigs were anesthetized to allow placement of a temperature measuring device in the abdominal cavity for measurement of intraperitoneal temperature (TEMP). On d 17, pigs were anesthetized and fitted with indwelling jugular vein catheters. On the following day (d 18), pigs were orally inoculated with 4.7×109 Salmonella typhimurium. Blood samples were collected at 0.5-h intervals from -2 to 8 h, and at 8-h intervals from 8 to 72 h post-challenge. Whole blood was analyzed for complete blood cell counts. Serum was isolated for measurement of cortisol. Following collection of the 72 h sample, pigs were humanely euthanized and tissues were collected for Salmonella isolation. There was a sex × time interaction (P < 0.001) for TEMP such that gilts had a greater TEMP response to the Salmonella challenge compared to barrows. There was also a sex × time interaction (P = 0.03) for serum cortisol with gilts having decreased cortisol at 16 h yet greater cortisol at 32 h than barrows. Barrows had greater total white blood cells (17.8 vs. 16.2 ± 0.4 103 cells/µL; P < 0.01; respectively) and neutrophils (7.8 vs. 6.1 ± 0.4 103 cells/µL; P < 0.01; respectively) than gilts. However, gilts had greater lymphocytes (9.6 vs. 9.0 ± 0.2 103 cells/µL; P = 0.05; respectively) than barrows. While immune parameters were influenced by sex, there was no effect of sex (P > 0.05) on Salmonella concentrations from fecal shedding 3 d post-inoculation in the cecum, mesenteric and subiliac lymph nodes, liver, spleen, gallbladder, or kidney tissues. These data demonstrate that weaned gilts appear to produce a stronger acute phase response to a Salmonella challenge compared to barrows, without affecting the tissue translocation or shedding of Salmonella.

Salmonella Typhimurium and Multidirectional Communication in the Gut.

The mammalian digestive tract is home to trillions of microbes, including bacteria, archaea, protozoa, fungi, and viruses. In monogastric mammals the stomach and small intestine harbor diverse bacterial populations but are typically less populated than the colon. The gut bacterial community (microbiota hereafter) varies widely among different host species and individuals within a species. It is influenced by season of the year, age of the host, stress and disease. Ideally, the host and microbiota benefit each other. The host provides nutrients to the microbiota and the microbiota assists the host with digestion and nutrient metabolism. The resident microbiota competes with pathogens for space and nutrients and, through this competition, protects the host in a phenomenon called colonization resistance. The microbiota participates in development of the host immune system, particularly regulation of autoimmunity and mucosal immune response. The microbiota also shapes gut-brain communication and host responses to stress; and, indeed, the microbiota is a newly recognized endocrine organ within mammalian hosts. Salmonella enterica serovar Typhimurium (S. Typhimurium hereafter) is a food-borne pathogen which adapts to and alters the gastrointestinal (GI) environment. In the GI tract, S. Typhimurium competes with the microbiota for nutrients and overcomes colonization resistance to establish infection. To do this, S. Typhimurium uses multiple defense mechanisms to resist environmental stressors, like the acidic pH of the stomach, and virulence mechanisms which allow it to invade the intestinal epithelium and disseminate throughout the host. To coordinate gene expression and disrupt signaling within the microbiota and between host and microbiota, S. Typhimurium employs its own chemical signaling and may regulate host hormone metabolism. This review will discuss the multidirectional interaction between S. Typhimurium, host and microbiota as well as mechanisms that allow S. Typhimurium to succeed in the gut.

Adhesion of Fusobacterium necrophorum to bovine endothelial cells is mediated by outer membrane proteins.

Fusobacterium necrophorum, a Gram-negative anaerobe, is frequently associated with suppurative and necrotic infections of animals and humans. The organism is a major bovine pathogen, and in cattle, the common fusobacterial infections are hepatic abscesses, foot rot, and necrotic laryngitis. The species comprises two subspecies: F. necrophorum subsp. necrophorum and F. necrophorum subsp. funduliforme. Bacterial adhesion to the host cell surface is a critical initial step in the pathogenesis, and outer membrane proteins (OMP) play an important role in adhesion and establishment of certain Gram-negative bacterial infections. The means by which F. necrophorum attaches to epithelial or endothelial cells has not been determined. We evaluated whether OMP of F. necrophorum, isolated from a liver abscess, mediated adhesion to bovine endothelial cells (adrenal gland capillary endothelial cell line). The extent of binding of subsp. necrophorum to the endothelial cells was higher than that of F. necrophorum subsp. funduliforme. Trypsin treatment of bacterial cells decreased their binding to endothelial cells indicating the protein nature of adhesins. Preincubation of endothelial cells with OMP extracted from F. necrophorum decreased the binding of bacterial cells. In addition, binding of each subspecies to endothelial cells was inhibited by polyclonal antibodies raised against respective OMP and the antibody-mediated inhibition was subspecies specific. The western blot analysis of OMP bound to endothelial cells with anti-OMP antibodies showed four OMP of 17, 24, 40 and 74 kDa. We conclude that OMP of F. necrophorum play a role in adhesion of bacterial cells to the endothelial cells.

Catecholamines increase conjugative gene transfer between enteric bacteria.

The ability of pathogenic bacteria to sense and respond to periods of host stress is critical to their lifestyle. Adrenaline and norepinephrine are catecholamines that mediate acute host stress in vertebrates and invertebrates. Catecholamines are also used as environmental cues to enhance growth, motility and virulence of bacterial pathogens via specific binding receptors. Incidence of multidrug resistant and highly virulent bacterial pathogens is on the rise, and majority of the genes for antimicrobial resistance (AMR) and virulence are carried on horizontally transferable genetic elements. Conjugation machinery offers an efficient method for acquisition of AMR and virulence genes, which may be responsible for propelling the evolution of pathogenic bacteria. Here we show that norepinephrine (NE) at physiological concentrations enhances horizontal gene transfer (HGT) efficiencies of a conjugative plasmid from a clinical strain of Salmonella Typhimurium to an Escherichia coli recipient in vitro. Expressions of plasmid encoded transfer (tra) genes necessary for conjugation were also significantly upregulated in the presence of NE. Phentolamine, an α-adrenergic receptor antagonist, negated the effects of NE on conjugation more strongly than propranolol, a ß-adrenergic receptor antagonist. This study for the first time provides evidence that innate mediators of acute host stress may influence evolution and adaptation of bacterial pathogens.