Non-targeted metabolomics of saliva to explore potential biomarkers for gastric ulceration in pigs fed hemp

Gastric ulceration is a common disease in pig production worldwide and is associated with economic losses as well as animal health and welfare issues. The aim of this study was to explore potential salivary biomarkers for gastric ulceration in pigs. In addition, the aim was to study the effect of hemp on the incidence of gastric ulcers. Approximately 440 growing-finishing pigs in the period from 30 to 110 kg BW were allocated to four different diets: meal feed (Meal); pelleted feed (Pellets); pelleted feed added 4% hempseed cake (Hemp Cake); pelleted feed added 4% hempseed hulls (Hemp Hulls). The day before slaughter, saliva samples from each pig were collected. After slaughter, the stomachs were emptied to assess the consistency of the stomach content and examined for gastric ulceration using an index scale (0–10). Noticeable changes of the gastric mucosa (total index score ≥ 6) were observed in 291 pigs. The odds of having index scores 0–5 relative to index scores 6–8 and 9–10, respectively, were higher (P < 0.001) for pigs fed Meal compared to pigs fed Pellets. The odds of suffering from severe gastric ulcers tended (P = 0.08) to be lower in pigs fed Hemp Hulls compared to pigs fed Pellets. A non-targeted liquid chromatography mass spectrometry based metabolomics analysis was performed on saliva samples to determine any separation between pigs with healthy stomachs and those with gastric ulcers and to examine a possible correlation between gastric ulcer index and potential biomarkers. Partial least-squares discriminant analysis showed a separation between pigs with ulcers and those with healthy stomachs/hyperkeratosis (HK). Metabolites contributing to the separation between groups were identified. Levels of oxylipins deriving from linoleic acid were lower (P < 0.001) in pigs with ulcers compared to healthy/HK pigs. This may indicate a shift in the metabolic pathways towards more pro-inflammatory arachidonic acid-derived eicosanoids, which might reflect an increased inflammatory response. Thus, reduced levels of oxylipins derived from linoleic acid seemed to be associated with active gastric ulcers, and thereby they might function as biomarkers for gastric ulceration in pigs. In addition, supplementation of hempseed hulls had a beneficial effect on severe gastric ulcers, as hempseed hulls changed the consistency of the gastric content by conferring more solidness. However, it was not possible to observe any reliable separation between pigs fed pellets supplemented with hemp products and pigs fed non-supplemented pellets according to the identified salivary metabolites.     

The formation of filamentous structures from iodinated neurotubules

The porcine neurotubule and its basic subunit were found to be modified in vitro by iodination of amino acids (principally tyrosine) using lactoperoxidase. Iodide ion, H₂O₂, or lactoperoxidase singly or in any pairwise combination had virtually no effect on neurotubules. However, when all three reagents were present, permitting covalent iodination, it was found that at 0.1 iodotyrosines per tubulin dimer the microtubules unravel to form structures which morphologically resemble strands of protofilaments twisted or wound around each other. These abnormal tubules are stable at room temperature and 4°C. Both monomers of tubulin are labeled to approximately the same extent. Iodinated tubulin (0.1 iodotyrosines/dimer) is unable to assemble in vitro under normal assembly conditions. Heavily iodinated microtubules (8 iodines per tubulin dimer) are similar in morphology to the slightly iodinated structures.     

High throughput detection of antibody self-interaction by bio-layer interferometry

Self-interaction of an antibody may lead to aggregation, low solubility or high viscosity. Rapid identification of highly developable leads remains challenging, even though progress has been made with the introduction of techniques such as self-interaction chromatography (SIC) and cross-interaction chromatography (CIC). Here, we report a high throughput method to detect antibody clone self-interaction (CSI) using bio-layer interferometry (BLI) technology. Antibodies with strong self-interaction responses in the CSI-BLI assay also show delayed retention times in SIC and CIC. This method allows hundreds of candidates to be screened in a matter of hours with minimal material consumption.     

Enzyme-linked enzyme-binding assay for Pin1 WW domain ligands

Peptidyl prolyl cis-trans isomerase (PPIase) interacting with NIMA-1 (Pin1) catalyzes the cis-trans isomerization of pSer/pThr-Pro amide bonds. Pin1 is a two-domain protein that represents a promising target for the treatment of cancer. Both domains of Pin1 bind the pSer/pThr-Pro motif; PPIase enzymatic activity occurs in the catalytic domain, and the WW domain acts as a recognition module for the pSer/pThr-Pro motif. An assay we call an enzyme-linked enzyme-binding assay (ELEBA) was developed to measure the K_d of ligands that bind selectively to the WW domain. A ligand specific for the WW domain of Pin1 was covalently immobilized in a 96-well plate. Commercially available Pin1 conjugated to horseradish peroxidase was used for chemiluminescent detection of ligands that block the association of the WW domain with immobilized ligand. The peptide ligands were derived from the cell cycle regulatory phosphatase, Cdc25c, residues 45-50. The K_d values for Fmoc-VPRpTPVGGGK-NH₂ and Ac-VPRpTPV-NH₂ were determined to be 36 ± 4 and 110 ± 30 μM, respectively. The ELEBA offers a selective approach for detecting ligands that bind to the Pin1 WW domain, even in the presence of the catalytic domain. This method may be applied to any dual specificity, multidomain protein.     

The thermal physiology of Stenopelmus rufinasus and Neohydronomus affinis (Coleoptera: Curculionidae), two biological control agents for the invasive alien aquatic weeds,Azolla filiculoides and Pistia stratiotes in South Africa

Water lettuce, Pistia stratiotes, and red water fern, Azolla filiculoides, are floating aquatic macrophytes that have become problematic in South Africa. Two weevils, Neohydronomus affinis and Stenopelmus rufinasus, are successful biological control agents of these two species in South Africa. The aim of this study was to investigate the thermal requirements of these two species to explain their establishment patterns in the field. Laboratory results showed that both weevils are widely tolerant to cold and warm temperatures. The critical thermal minima (CT_min) of N. affinis was determined to be 5.58?±?0.31°C and the critical thermal maxima (CT_max) was 44.52?±?0.27°C, while the CT_min of S. rufinasus was 5.38?±?0.33°C and the CT_max?44.0?±?0.17°C. In addition, the lower lethal temperatures were ?9.85?±?0.06°C for N. affinis and ?6.85?±?0.13°C for S. rufinasus, and the upper lethal temperatures were 42.7?±?0.85°C for N. affinis and 41.9?±?2.52°C S. rufinasus. Using the reduced major axis regression method, the development for N. affinis was described using the formula y?=?12.976x?+?435.24, while the development of S. rufinasus was described by y?=?13.6x?+?222.45. These results showed that S. rufinasus develops twice as fast as N. affinis. Using these formulae and temperature data obtained from the South African Weather Service, N. affinis was predicted to complete between 4 and 9 generations per year in South Africa, while S. rufinasus was predicted to complete between 5 and 14 generations per year around the country. These results suggest that both species should not be limited by cold winter, nor warm summer temperatures, and should establish throughout the ranges of the weeds in South Africa.     

Firstborn sex defines early childhood growth of subsequent siblings

Animal studies have shown that maternal resource allocation can be sex-biased in order to maximize reproductive success, yet this basic concept has not been investigated in humans. In this study, we explored relationships between maternal factors, offspring sex and prenatal and postnatal weight gain. Sex-specific regression models not only indicated that maternal ethnicity impacted male (n = 2456) and female (n = 1871) childrens postnatal weight gain differently but also that parity and mode of feeding influenced weight velocity of female (β ± s.e. = −0.31 ± 0.11 kg, p = 0.005; β ± s.e. = −0.37 ± 0.11 kg, p < 0.001) but not male offspring. Collectively, our findings imply that maternal resource allocation to consecutive offspring increases after a male firstborn. The absence of this finding in formula fed children suggests that this observation could be mediated by breast milk. Our results warrant further mechanistic and epidemiological studies to elucidate the role of breastfeeding on the programming of infant growth as well as of metabolic and cardiovascular diseases, with potential implications for tailoring infant formulae according to sex and birth order.     

Patterns of nucleotide and haplotype diversity at ICAM-1 across global human populations with varying levels of malaria exposure

Malaria is one of the strongest selective pressures in recent human evolution. African populations have been and continue to be at risk for malarial infections. However, few studies have re-sequenced malaria susceptibility loci across geographically and genetically diverse groups in Africa. We examined nucleotide diversity at Intercellular adhesion molecule-1 (ICAM-1), a malaria susceptibility candidate locus, in a number of human populations with a specific focus on diverse African ethnic groups. We used tests of neutrality to assess whether natural selection has impacted this locus and tested whether SNP variation at ICAM-1 is correlated with malaria endemicity. We observe differing patterns of nucleotide and haplotype variation in global populations and higher levels of diversity in Africa. Although we do not observe a deviation from neutrality based on the allele frequency distribution, we do observe several alleles at ICAM-1, including the ICAM-1 ^Kilifi allele, that are correlated with malaria endemicity. We show that the ICAM-1 ^Kilifi allele, which is common in Africa and Asia, exists on distinct haplotype backgrounds and is likely to have arisen more recently in Asia. Our results suggest that correlation analyses of allele frequencies and malaria endemicity may be useful for identifying candidate functional variants that play a role in malaria resistance and susceptibility.