Influence of light exposure on horizontal transmission of Salmonella typhimurium in weaned pigs

The objective of the following experiment was to examine the effect of light exposure on horizontal transmission of Salmonella typhimurium in weaned pigs. Twenty crossbred pigs (average BW = 15 kg) were housed in isolation rooms (10 pigs/room) and randomly assigned to one of two lighting regimes. Low (8 h light, 16 h dark) or High (16 h light, 8 h dark). Pigs were adjusted to their respective lighting treatments for six days and on the seventh day, two randomly selected pigs/room orally inoculated with 5 ml of tryptic soy broth containing 18 x 108 cfu Salmonella typhimurium/ml. Rectal swabs were collected from each pig daily over the next eight days for direct plating and plating following 24-h enrichment. On day nine, following inoculation of the seeder pigs, all pigs were euthanized and necropsied. Luminal contents were collected from the ileum, colon, cecum and rectum (quantification and qualification of inoculated strain) and tissue samples collected from the above gut segments as well as the tonsils, ileo-cecal lymph nodes, spleen and liver (qualification only)

Calcium-Dependent Association of Calmodulin with the C-Terminal Domain of the Tetraspanin Protein Peripherin/rds

Peripherin/rds (p/rds), an integral membrane protein from the transmembrane 4 (TMF4) superfamily, possesses a multi-functional C-terminal domain that plays crucial roles in rod outer segment (ROS) disk renewal and structure. Here, we report that the calcium binding protein calmodulin (CaM) binds to the C-terminal domain of p/rds. Fluorescence spectroscopy reveals Ca2+-dependent association of CaM with a polypeptide corresponding to the C-terminal domain of p/rds. The fluorescence anisotropy of the polypeptide upon CaM titration yields a dissociation constant (KD) of 320 ± 150 nM. The results of the fluorescence experiments were confirmed by GST-pull down analyses in which a GST-p/rds C-terminal domain fusion protein was shown to pull down CaM in a calcium-dependent manner. Moreover, molecular modeling and sequence predictions suggest that the CaM binding domain resides in a p/rds functional hot spot, between residues E314 and G329. Predictions were confirmed by peptide competition studies and a GST-p/rds C-terminal domain construct in which the putative Ca2+/CaM binding site was scrambled. This GST-polypeptide did not associate with Ca2+/CaM. This putative calmodulin domain is highly conserved between human, mouse, rat, and bovine p/rds. Finally, the binding of Ca2+/CaM inhibited fusion between ROS disk and ROS plasma membranes as well as p/rds C-terminal-domain-induced fusion in model membrane studies. These results offer a new mechanism for the modulation of p/rds function. © 2007 American Chemical Society

Membrane Association and Destabilization by Aggregatibacter Actinomycetemcomitans Leukotoxin Requires Changes in Secondary Structures

Summary: Aggregatibacter actinomycetemcomitans is a common inhabitant of the upper aerodigestive tract of humans and non-human primates and is associated with disseminated infections, including lung and brain abscesses, pediatric infective endocarditis, and localized aggressive periodontitis. Aggregatibacter actinomycetemcomitans secretes a repeats-in-toxin protein, leukotoxin, which exclusively kills lymphocyte function-associated antigen-1-bearing cells. The toxin\u27s pathological mechanism is not fully understood; however, experimental evidence indicates that it involves the association with and subsequent destabilization of the target cell\u27s plasma membrane. We have long hypothesized that leukotoxin secondary structure is strongly correlated with membrane association and destabilization. In this study, we tested this hypothesis by analysing lipid-induced changes in leukotoxin conformation. Upon incubation of leukotoxin with lipids that favor leukotoxin-membrane association, we observed an increase in leukotoxin α-helical content that was not observed with lipids that favor membrane destabilization. The change in leukotoxin conformation after incubation with these lipids suggests that membrane binding and membrane destabilization have distinct secondary structural requirements, suggesting that they are independent events. These studies provide insight into the mechanism of cell damage that leads to disease progression by A. actinomycetemcomitans. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd

Membrane Association and Destabilization by Aggregatibacter Actinomycetemcomitans Leukotoxin Requires Changes in Secondary Structures

Aggregatibacter actinomycetemcomitans is a common inhabitant of the upper aerodigestive tract of humans and non-human primates and is associated with disseminated infections, including lung and brain abscesses, pediatric infective endocarditis in children, and localized aggressive periodontitis. A. actinomycetemcomitans secretes a repeats-in-toxin protein, leukotoxin, which exclusively kills lymphocyte function-associated antigen-1-bearing cells. The toxin\u27s pathological mechanism is not fully understood; however, experimental evidence indicates that it involves the association with and subsequent destabilization of the target cell\u27s plasma membrane. We have long hypothesized that leukotoxin secondary structure is strongly correlated with membrane association and/or destabilization. In this study, we tested this hypothesis by analyzing lipid-induced changes in leukotoxin conformation. Upon incubation of leukotoxin with lipids that favor leukotoxin-membrane association, we observed an increase in leukotoxin α-helical content that was not observed with lipids that favor membrane destabilization. The change in leukotoxin conformation after incubation with these lipids suggests that membrane binding and membrane destabilization have distinct secondary structural requirements, suggesting that they are independent events. These studies thus provide insight into the mechanism of cell damage that leads to disease progression by A. actinomycetemcomitans

Isolation of Salmonella spp. and bacteriophage active against Salmonella spp. from commercial swine

Bacteriophage are viruses that prey on bacteria and may be a potential strategy to reduce foodborne pathogemc bactena in the gastromtestlnal tract of food animals Phages are fairly common in the gastrointestinal microbial ecosystem of mammals, but the incidence is unknown. If phage are to be an intervention strategy, we must understand their role in the microbial ecology of the gut. From a regulatory perspective, knowing incidence of phage is crucial. Therefore the current study was designed to determine the incidence of phage active against Salmonella spp in the feces of commercial finishing swine in the United States. Fecal samples (n=60) were collected from each of six commercial swine finishing operations. Samples were collected from 10 randomly selected pens throughout each operation. Total number of fecal samples collected in this study was n=360 Salmonella spp were found in 66% of the fecal samples Salmonella spp. were isolated from only 2 farms and the serotypes represented were Schwarzengrund, Anatum, Ohio and Heidelberg Bacteriophages were isolated from fecal sample through 2 parallel methods, 1) initlal enrichment in Salmonella Typhimunum, or 2) initial ennchment in E. colt B (a strain very sensitive to phages), followed by direct spot-testing against Salmonella Typhimurium Bacteriophages active against Salmonella Typhimunum were isolated from 1.1% 4/360) of the individual fecal samples when initially enriched in Salmonella Typhimurium, but E coli S-killing phages were 1solated from 43.8% (158/360) of the fecal samples but only 2 of these Isolates were capable of k1ll1ng Salmonella Typhimunum. Our results mdicate that bactenophage capable of killing Salmonella Typh1murium are fairly w1despread across commercial swine production facilities but may be present at relatively low populat1ons These results md1cate that phage (predator) populations may vary along w1th Salmonella (prey) populations and that phage could potentially be used as a food safety pathogen reduction strategy

Reduction of Campylobacter and Salmonella in pigs treated with A select nitrocompound

The aim of this study was to test the effectiveness of administering a select nitrocompound (S-NO) on reducing naturally colonized Campylobacter and experimentally infected Salmonella in the weaned pig gut. Pigs were divided into four groups; control (0 g S-NO/pig), 1X (0.2 g S-NO/pig), 5X (1 g S-NO/pig), and 10X (2 g S-NO/pig). Treatments were administered via oral gavage 24 h before sacrifice. Mean ± SD populations (log10 cfu/g) of Campylobacter in the cecum were reduced (P \u3c 0.05) in pigs receiving the 10X dose when compared with untreated controls (1.64 ± 1.30 vs 5.31 ± 0.58, respectively). Campylobacter concentrations in rectal contents from pigs administered the 5X dose were reduced (P \u3c 0.05) compared to control (2.65 ± 2.86 vs 5.90 ± 0.94, respectively). Rectal Salmonella concentrations were reduced (P \u3c 0.05) in all of the S-NO-treated groups. Adverse effects of S-NO on pig health were not observed. These results demonstrate that S-NO may have potential as an intervention to reduce pig colonization by Campylobacter and Salmonella

Effect of thymol or diphenyliodonium chloride on feed intake, average daily gain and gut Campylobacter concentrations in growing swine

Food producing animals can be reservoirs of Campylobacter, a leading bacterial cause of human foodborne illness. Campylobacter differ from most other gut bacteria in that they do not ferment carbohydrates but can utilize amino acids as major energy substrates, a process that can be inhibited by thymol and diphenyliodonium chloride (DIC). To evaluate palatability issues pertaining to feeding thymol or DIC, growing pigs were provided ad libitum access to standard growing diets supplemented with or without 0.0067 or 0.0201% thymol or 0.0001 4 or 0.00042% DIC in a replicated study design (n=4 pens per treatment per replicate, 2 pigs/pen). The diets were offered 2X/day for 7 days during which time average daily feed intake (2.39 ± 0.06 kg d-1; mean ± SEM) and average daily gain (0.62 ± 0.04 kg d-1) were not affected (P \u3e 0.05) by treatment