Metabolic insights and background from naturally affected pigs during Streptococcus suis outbreaks.

Streptococcus suis (S. suis) is an endemic zoonotic pathogen still lacking adequate prevention in pigs. The present case study looked back to the occurrence and consequences of S. suis outbreaks in our swine research facilities in search of new metabolic and physiological insight. From a series of outbreaks, a dataset was created including 56 pigs sampled during disease detection based on clinical signs. Pigs suspected with S. suis infection were defined as diseased (n = 28) and included pigs defined as neurologically diseased (n = 20) when severe neurological signs (central nervous system dysfunctions, i.e., opisthotonos, ataxia, and generalized tremor) were observed. Another set of 28 pigs included respective pen mates from each case and were defined as control. Representative deaths were confirmed to be caused by S. suis. Tonsillar swabs were collected and analyzed by quantitative polymerase chain reaction (qPCR) for total bacteria, total S. suis, and S. suis serotypes (SS) 2 (and/or 1/2) and 9. Blood and sera were analyzed to quantify blood gases, minerals, and S. suis reactive immunoglobulins against current isolates. Data collected included litter sibling associations, birth and weaning body weight (BW), and average daily gain (ADG) 7 d after the disease detection. In general, the disease increased pH, sO2 and the incidence of alkalosis, but reduced pCO2, glucose, Ca, P, Mg, K, and Na in blood/serum compared to control. The SS2 (and/or SS1/2) prevalence was significantly (P < 0.05) increased in neurologically diseased pigs and its relative abundance tended (P < 0.10) to increase in tonsils. In contrast, the relative abundance of total S. suis was lower (P > 0.05) in diseased pigs than control pigs. Levels of S. suis reactive IgG2 were lower, but IgM were higher (P < 0.03) in neurologically affected pigs compared to control. Furthermore, there was an increased proportion of sibling pigs that were diseased compared to control. In conclusion, our results evidence that naturally affected pigs were associated to average performing pigs without any predisease trait to highlight but a sow/litter effect. Besides, neurologically affected pigs had increased S. suis (SS2 and/or 1/2) prevalence and relative abundance, a respiratory alkalosis profile, and mineral loss.

Feed additive blends fed to nursery pigs challenged with Salmonella.

Salmonella in pigs is a concern for human foodborne salmonellosis. Dietary fungal fermented products, coated butyrate, and organic acids (OAs) may be promising control strategies. The objectives of this study were (i) to evaluate in vitro binding affinity of Salmonella enterica serovar Typhimurium (S. Typh) and Enteritidis (S. Ent), and enterotoxigenic Escherichia coli (ETEC) F4 or F18 to mannan-rich hydrolyzed copra meal (MCM) and fermented rye (FR) with Agaricus subrufescens; and (ii) to assess MCM and FR efficacy to control in vivo S. Typh shedding when combined with OAs and compared with coated butyrate strategy. A 31-d study included 32 pigs [6.29 ±â€…0.76 kg BW] individually housed and distributed into four dietary treatments: control diet; OA.BU, 4 kg/t OA plus 6 kg/t coated butyrate; OA.MCM, 4 kg/t OA plus 1 kg/t MCM; and OA.FR, 4 kg/t OA plus 2 kg/t FR. All pigs were challenged for 7 d with 1 mL S. Typh (109 colony forming units daily) at 10 d postweaning. Temperature and fecal samples were collected before and after challenge, and fecal Salmonella shedding quantified. Diarrhea scores were monitored daily and growth performance was evaluated weekly. In vitro, culture with MCM and FR showed significant (P < 0.01) binding affinity for both S. Typh and S. Ent, but not for ETEC F4 and F18. In vivo, pigs fed OA.MCM and OA.FR had lower (P < 0.05) shedding and day 3 peak shedding of S. Typh after infections than pigs fed control and OA.BU diets. Pigs fed OA.FR diet tended to have an 18% increase (P = 0.068) in BW on day 14 post first inoculation compared with control and OA.BU, and 19% increased (P = 0.093) final BW at day 21 compared with control. Diarrhea frequency post infection was overall lower (P = 0.006) for OA.FR (18.9%) than OA.BU (44.8%) and OA.MCM (41.7%) while control (28.7%) was not different. In conclusion, FR and MCM show in vitro-binding affinity to Salmonella enterica serovars Typh and Ent. Feeding FR or MCM combined with OA to nursery pigs reduces the peak and averages S. Typh shedding compared with control. Fermented rye with OA tends to improve pig performance after S. Typh challenge.

New perspectives to the enterotoxigenic E. coli F4 porcine infection model: Susceptibility genotypes in relation to performance, diarrhoea and bacterial shedding.

Enterotoxigenic E. coli (ETEC), causing post-weaning diarrhoea, is a major problem in weaned piglets. Individual animal responses to ETEC infection show high variability in animal experiments. Two studies were designed to optimize the ETEC F4ac infection model in piglets by combining the genotype susceptibility with performance, diarrhoea incidence and bacterial shedding. The studies were performed with respectively 120 and 80 male piglets that were tested for susceptibility or resistance towards ETEC O149:F4ac by a DNA marker based test. Three different genotypes were observed; resistant (RR), susceptible heterozygote (RS) and susceptible homozygote (SS). Piglets, were orally infected with an inoculum suspension (containing 1.5E8 CFU/ml ETEC F4ac) at day 0, 1 and 2 of the study. Performance, diarrhoea incidence and bacterial shedding were followed for 21days. In the first week after challenge a difference in average daily gain was observed between resistant and susceptible piglets in both studies. For the complete study period no significant differences were observed. Diarrhoea incidence was significantly higher in susceptible pigs compared to the resistant pigs in the first week after challenge. Bacterial shedding was much higher in the susceptible pigs and ETEC excretion lasted longer. ETEC was hardly detected in the faecal material of the resistant pigs. In conclusion, susceptible pigs showed higher diarrhoea incidence and higher numbers of faecal ETEC shedding in the first week after challenge compared to resistant pigs. The DNA marker based test can be used to select pigs that are susceptible for ETEC for inclusion in ETEC infection model, resulting in less animals needed to perform infection studies.

Gender-specific increase of bone mass by CART peptide treatment is ovary-dependent.

Cocaine- and amphetamine-regulated transcript (CART) has emerged as a neurotransmitter and hormone that has been implicated in many processes including food intake, maintenance of body weight, and reward, but also in the regulation of bone mass. CART-deficient mice are characterized by an osteoporotic phenotype, whereas female transgenic mice overexpressing CART display an increase in bone mass. Here we describe experiments that show that peripheral subcutaneous sustained release of different CART peptide isoforms for a period up to 60 days increased bone mass by 80% in intact mice. CART peptides increased trabecular bone mass, but not cortical bone mass, and the increase was caused by reduced osteoclast activity in combination with normal osteoblast activity. The observed effect on bone was gender-specific, because male mice did not respond to treatment with CART peptides. In addition, male transgenic CART overexpressing mice did not display increased bone mass. Ovariectomy (OVX) completely abolished the increase of bone mass by CART peptides, both in CART peptide-treated wild-type mice and in CART transgenic mice. The effect of CART peptide treatment on trabecular bone was not mediated by 17ß-estradiol (E(2)) because supplementation of OVX mice with E(2) could not rescue the effect of CART peptides on bone. Together, these results indicate that sustained release of CART peptides increases bone mass in a gender-specific way via a yet unknown mechanism that requires the presence of the ovary.

Molecular cloning and characterization of a broad substrate terpenoid oxidoreductase from Artemisia annua.

From Artemisia annua L., a new oxidoreductase (Red 1) was cloned, sequenced and functionally characterized. Through bioinformatics, heterologous protein expression and enzyme substrate conversion assays, the elucidation of the enzymatic capacities of Red1 was achieved. Red1 acts on monoterpenoids, and in particular functions as a menthone:neomenthol oxidoreductase. The kinetic parameter k(cat)/K(m) was determined to be 939-fold more efficient for the reduction of (-)-menthone to (+)-neomenthol than results previously reported for the menthone:neomenthol reductase from Mentha x piperita. Based on its kinetic properties, the possible use of Red1 in biological crop protection is discussed.

Self-assembling protein arrays on DNA chips by auto-labeling fusion proteins with a single DNA address.

The high-throughput deposition of recombinant proteins on chips, beads or biosensor devices would be greatly facilitated by the implementation of self-assembly concepts. DNA-directed immobilization via conjugation of proteins to an oligonucleotide would be preeminently suited for this purpose. Here, we present a unique method to attach a single DNA address to proteins in one step during the purification from the E. coli lysate by fusion to human O6-alkylguanine-DNA-alkyltransferase (SNAP-tag) and the Avitag. Use of the conjugates in converting a DNA chip into a protein chip by self assembly is demonstrated.