Sainfoin – New Data on Anthelmintic Effects and Production in Sheep and Goats

Gastrointestinal nematodes (GIN) are one of the most important problems affecting health and therefore performance and welfare in small ruminant husbandry. The control of these parasites in the past strongly relied on the repeated use of anthelmintic drugs. This has led to nematode populations which are resistant to most of the currently available anthelmintics. Furthermore customer’s demands for organic and residue free animal products are increasing. The aforementioned problems have given a strong impetus for the development of new non-chemical strategies to control GIN. Previous research has pointed out the anthelmintic potential of sainfoin (Onobrychis viciifolia) and other tanniferous (CT) feed sources in goats and lambs infected with GIN. A recent Swiss experiment focussed on the use of sainfoin and field bean (Vicia faba, cv. Scirocco) as single CT sources as well as in combination for additional synergic effects, to reduce periparturient GIN egg rise of ewes in late gestation and early lactation. Another experiment with Alpine goats concentrated on the influence of sainfoin on milk performance and cheese quality. The results of these experiments will be presented and discussed in connection with previous knowledge on (i) anthelmintic effects of sainfoin and (ii) the influence of sainfoin administration on performance

Pumpkin seeds, lemongrass essential oil and ripleaf leaves as feed additives for Ascaridia galli infected laying hens.

The present study was performed to evaluate the in vivo efficiency of Curcurbita pepo (pumpkin) seeds, Cymbopogon citratus (lemongrass) essential oil and Plantago lanceolata (ripleaf) leaves against helminth infections in laying hens. In the first experiment, 75 Lohmann LSL Classic hens naturally infected with Ascaridia galli were assigned to groups of five; groups were randomly assigned to one of three treatments with five replicates each (untreated control; lemongrass oil: 1 g/bird/day; pumpkin seeds: 10 g/bird/day). Feed consumption and egg production were continuously recorded, individual faecal egg counts were determined weekly, and E. coli and Lactobacillus spp. three times during the experimental period of 29 days. After slaughter, intestinal worms were counted and sexed. Pumpkin improved feed conversion as compared to the control (p = 0.008) and to lemongrass (p = 0.021); no treatment effect on any other parameter was found. In the second experiment, 75 LSL pullets were artificially infected with 3 × 200 A. galli eggs, randomly divided into groups of five and assigned to one of three treatments (untreated control, lemongrass oil: 1 g/bird/day; ripleaf: 5% of ration). After 109 days of sampling as described above, hens were slaughtered and worm burdens determined. Performance of the animals did not change regardless of the treatment and none of the treatments resulted in changes of the microbiological and parasitological parameters. In conclusion, with the exception of improved feed conversion in the pumpkin group, no positive nor negative effects of the additives on performance, parasitological and microbiological parameters of naturally and artificially A. galli infected laying hens were observed

A Predictive Model of Intein Insertion Site for Use in the Engineering of Molecular Switches

Inteins are intervening protein domains with self-splicing ability that can be used as molecular switches to control activity of their host protein. Successfully engineering an intein into a host protein requires identifying an insertion site that permits intein insertion and splicing while allowing for proper folding of the mature protein post-splicing. By analyzing sequence and structure based properties of native intein insertion sites we have identified four features that showed significant correlation with the location of the intein insertion sites, and therefore may be useful in predicting insertion sites in other proteins that provide native-like intein function. Three of these properties, the distance to the active site and dimer interface site, the SVM score of the splice site cassette, and the sequence conservation of the site showed statistically significant correlation and strong predictive power, with area under the curve (AUC) values of 0.79, 0.76, and 0.73 respectively, while the distance to secondary structure/loop junction showed significance but with less predictive power (AUC of 0.54). In a case study of 20 insertion sites in the XynB xylanase, two features of native insertion sites showed correlation with the splice sites and demonstrated predictive value in selecting non-native splice sites. Structural modeling of intein insertions at two sites highlighted the role that the insertion site location could play on the ability of the intein to modulate activity of the host protein. These findings can be used to enrich the selection of insertion sites capable of supporting intein splicing and hosting an intein switch

3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors reduce the risk of perioperative stroke and mortality after carotid endarterectomy

ObjectiveThere is increasing evidence that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) reduce cardiovascular and cerebrovascular events through anti-inflammatory, plaque stabilization, and neuroprotective effects independent of lipid lowering. This study was designed to investigate whether statin use reduces the incidence of perioperative stroke and mortality among patients undergoing carotid endarterectomy (CEA).MethodsAll patients undergoing CEA from 1994 to 2004 at a large academic medical center were retrospectively reviewed. The independent association of statin use and perioperative morbidity was assessed via multivariate logistic regression analysis.ResultsCEA was performed by 13 surgeons on 1566 patients (987 men and 579 women; mean age, 72 ± 10 years), including 1440 (92%) isolated and 126 (8%) combined CEA/coronary artery bypass grafting procedures. The indication for CEA was symptomatic disease in 660 (42%) cases. Six hundred fifty-seven (42%) patients received a statin medication for at least 1 week before surgery. Statin use was associated with a reduction in perioperative strokes (1.2% vs 4.5%; P < .01), transient ischemic attacks (1.5% vs 3.6%; P < .01), all-cause mortality (0.3% vs 2.1%; P < .01), and median (interquartile range) length of hospitalization (2 days [2-5 days] vs 3 days [2-7 days]; P < .05). Adjusting for all demographics and comorbidities in multivariate analysis, statin use independently reduced the odds of stroke threefold (odds ratio [95% confidence interval], 0.35 [0.15-0.85]; P < .05) and death fivefold (odds ratio [95% confidence interval], 0.20 [0.04-0.99]; P < .05).ConclusionsThese data suggest that perioperative statin use may reduce the incidence of cerebrovascular events and mortality among patients undergoing CEA

Pumpkin seeds, lemongrass essential oil and ripleaf leaves as feed additives for Ascaridia galli infected laying hens

The present study was performed to evaluate the in vivo efficiency of Curcurbita pepo (pumpkin) seeds, Cymbopogon citratus (lemongrass) essential oil and Plantago lanceolata (ripleaf) leaves against helminth infections in laying hens. In the first experiment, 75 Lohmann LSL Classic hens naturally infected with Ascaridia galli were assigned to groups of five; groups were randomly assigned to one of three treatments with five replicates each (untreated control; lemongrass oil: 1 g/bird/day; pumpkin seeds: 10 g/bird/day). Feed consumption and egg production were continuously recorded, individual faecal egg counts were determined weekly, and E. coli and Lactobacillus spp. three times during the experimental period of 29 days. After slaughter, intestinal worms were counted and sexed. Pumpkin improved feed conversion as compared to the control (p = 0.008) and to lemongrass (p = 0.021); no treatment effect on any other parameter was found. In the second experiment, 75 LSL pullets were artificially infected with 3 × 200 A. galli eggs, randomly divided into groups of five and assigned to one of three treatments (untreated control, lemongrass oil: 1 g/bird/day; ripleaf: 5% of ration). After 109 days of sampling as described above, hens were slaughtered and worm burdens determined. Performance of the animals did not change regardless of the treatment and none of the treatments resulted in changes of the microbiological and parasitological parameters. In conclusion, with the exception of improved feed conversion in the pumpkin group, no positive nor negative effects of the additives on performance, parasitological and microbiological parameters of naturally and artificially A. galli infected laying hens were observed

Conservation of intron and intein insertion sites: implications for life histories of parasitic genetic elements

<p>Abstract</p> <p>Background</p> <p>Inteins and introns are genetic elements that are removed from proteins and RNA after translation or transcription, respectively. Previous studies have suggested that these genetic elements are found in conserved parts of the host protein. To our knowledge this type of analysis has not been done for group II introns residing within a gene. Here we provide quantitative statistical support from an analyses of proteins that host inteins, group I introns, group II introns and spliceosomal introns across all three domains of life.</p> <p>Results</p> <p>To determine whether or not inteins, group I, group II, and spliceosomal introns are found preferentially in conserved regions of their respective host protein, conservation profiles were generated and intein and intron positions were mapped to the profiles. Fisher's combined probability test was used to determine the significance of the distribution of insertion sites across the conservation profile for each protein. For a subset of studied proteins, the conservation profile and insertion positions were mapped to protein structures to determine if the insertion sites correlate to regions of functional activity. All inteins and most group I introns were found to be preferentially located within conserved regions; in contrast, a bacterial intein-like protein, group II and spliceosomal introns did not show a preference for conserved sites.</p> <p>Conclusions</p> <p>These findings demonstrate that inteins and group I introns are found preferentially in conserved regions of their respective host proteins. Homing endonucleases are often located within inteins and group I introns and these may facilitate mobility to conserved regions. Insertion at these conserved positions decreases the chance of elimination, and slows deletion of the elements, since removal of the elements has to be precise as not to disrupt the function of the protein. Furthermore, functional constrains on the targeted site make it more difficult for hosts to evolve immunity to the homing endonuclease. Therefore, these elements will better survive and propagate as molecular parasites in conserved sites. In contrast, spliceosomal introns and group II introns do not show significant preference for conserved sites and appear to have adopted a different strategy to evade loss.</p

In Vivo Characterization of the Homing Endonuclease within the polB Gene in the Halophilic Archaeon Haloferax volcanii

Inteins are parasitic genetic elements, analogous to introns that excise themselves at the protein level by self-splicing, allowing the formation of functional non-disrupted proteins. Many inteins contain a homing endonuclease (HEN) gene, and rely on its activity for horizontal propagation. In the halophilic archaeon, Haloferax volcanii, the gene encoding DNA polymerase B (polB) contains an intein with an annotated but uncharacterized HEN. Here we examine the activity of the polB HEN in vivo, within its natural archaeal host. We show that this HEN is highly active, and able to insert the intein into both a chromosomal target and an extra-chromosomal plasmid target, by gene conversion. We also demonstrate that the frequency of its incorporation depends on the length of the flanking homologous sequences around the target site, reflecting its dependence on the homologous recombination machinery. Although several evolutionary models predict that the presence of an intein involves a change in the fitness of the host organism, our results show that a strain deleted for the intein sequence shows no significant changes in growth rate compared to the wild type

Application of Ni(II)-Assisted Peptide Bond Hydrolysis to Non-Enzymatic Affinity Tag Removal

In this study, we demonstrate a non-enzymatic method for hydrolytic peptide bond cleavage, applied to the removal of an affinity tag from a recombinant fusion protein, SPI2-SRHWAP-His6. This method is based on a highly specific Ni(II) reaction with (S/T)XHZ peptide sequences. It can be applied for the protein attached to an affinity column or to the unbound protein in solution. We studied the effect of pH, temperature and Ni(II) concentration on the efficacy of cleavage and developed an analytical protocol, which provides active protein with a 90% yield and ∼100% purity. The method works well in the presence of non-ionic detergents, DTT and GuHCl, therefore providing a viable alternative for currently used techniques

A Cost-Effective ELP-Intein Coupling System for Recombinant Protein Purification from Plant Production Platform

BACKGROUND: Plant bioreactor offers an efficient and economical system for large-scale production of recombinant proteins. However, high cost and difficulty in scaling-up of downstream purification of the target protein, particularly the common involvement of affinity chromatography and protease in the purification process, has hampered its industrial scale application, therefore a cost-effective and easily scale-up purification method is highly desirable for further development of plant bioreactor. METHODOLOGY/PRINCIPAL FINDINGS: To tackle this problem, we investigated the ELP-intein coupling system for purification of recombinant proteins expressed in transgenic plants using a plant lectin (PAL) with anti-tumor bioactivity as example target protein and rice seeds as production platform. Results showed that ELP-intein-PAL (EiP) fusion protein formed novel irregular ER-derived protein bodies in endosperm cells by retention of endogenous prolamins. The fusion protein was partially self-cleaved in vivo, but only self-cleaved PAL protein was detected in total seed protein sample and deposited in protein storage vacuoles (PSV). The in vivo uncleaved EiP protein was accumulated up to 2-4.2% of the total seed protein. The target PAL protein could be purified by the ELP-intein system efficiently without using complicated instruments and expensive chemicals, and the yield of pure PAL protein by the current method was up to 1.1 mg/g total seed protein. CONCLUSION/SIGNIFICANCE: This study successfully demonstrated the purification of an example recombinant protein from rice seeds by the ELP-intein system. The whole purification procedure can be easily scaled up for industrial production, providing the first evidence on applying the ELP-intein coupling system to achieve cost-effective purification of recombinant proteins expressed in plant bioreactors and its possible application in industry