Surveillance of Mycoplasma agalactiae and Mycoplasma mycoides subsp. capri in dairy goat herds

This study was designed to monitor the presence of Mycoplasma agalactiae and Mycoplasma mycoides subsp. capri (Mmc) in 66 dairy goat herds of a genetic improvement programme in a region of Spain where contagious agalactia is endemic. Over a whole lactation period, 300 bulk tank milk and 381 milk samples from goats with clinical mastitis were subjected to polymerase chain reaction (PCR) to detect the two mycoplasma species. The presence of mycoplasmas (either species or both) was detected in 66.7% of the herds and M. agalactiae was identified in 95.45% of these positives herds. In a given infected herd, mycoplasmas were not continuously detected over the whole study period. Our findings indicate that in an endemic area, M. agalactiae and Mmc can be monitored through PCR analysis of mastitic milk and bulk tank milk (BTM) samples. Over a lactation period we recommend testing multiple BTM samples on a herd. No relationship was observed between the use of inactivated mycoplasma vaccines and the PCR detection of both mycoplasmas.     

Mechanisms involved in testosterone-induced relaxation to the pig urinary bladder neck

ObjectivesTestosterone replacement therapy improves bladder capacity in urinary tract dysfunction. There is no information, however, about the role of this steroid hormone on the muscle tension of the bladder outflow region. The current study investigated the mechanisms underlying the testosterone-induced action in the pig bladder neck.MethodsUrothelium-denuded bladder neck strips were mounted in myographs for isometric force recordings and for simultaneous measurements of intracellular Ca²⁺ concentration ([Ca²⁺]_i) and tension. The relaxations to testosterone, the non-aromatizable metabolite 4,5α-dihydrotestosterone (DHT) and electrical field stimulation (EFS) were carried out on phenylephrine (PhE)-precontracted strips.ResultsTestosterone and DHT evoked similar concentration-dependent relaxations only at very high pharmacological concentrations. The presence of the urothelium and the inhibition of intracellular androgenic receptor (AR), aromatase, 5α-reductase, nitric oxide (NO) synthase, guanylyl cyclase, cyclooxygenase (COX), large-, intermediate- and small-Ca²⁺-activated K⁺ channels or ATP-dependent K⁺ channels failed to modify the testosterone relaxations. Neuronal voltage-gated Ca²⁺ (VOC) channels and voltage-gated K⁺ (K_V) channel blockers potentiated these responses. EFS evoked frequency-dependent relaxations, which were not changed by threshold concentrations of testosterone. In Ca²⁺-free potassium rich physiological saline solution, testosterone inhibited the contractions induced by CaCl₂ and the L-type VOC channel activator (±)-BAY K 8644. Relaxations elicited by testosterone were accompanied by simultaneous decreases in smooth muscle [Ca²⁺]_i.ConclusionsTestosterone produces relaxation of the pig urinary bladder neck through mechanisms independent of urothelium, AR, aromatase, 5α-reductase, NO synthase, guanylyl cyclase, COX and K⁺ channels. Testosterone-induced relaxation is produced via the inhibition of the extracellular Ca²⁺ entry through L-type VOC channels.     

Anthracene Removal using Activated Soil Reactors

The aim of this study was to evaluate anthracene removal using activated soil reactors, previously inoculated, under both aerobic and anaerobic conditions. In the reactors, the soil was maintained at 60% moisture (weight basis), room temperature, in the dark, and under constant agitation at 100 rpm. Two experiments were run during and after acclimatization to evaluate anthracene removal under both aerobic and anaerobic conditions. The first one took place during inoculum acclimatization using three different concentrations of anthracene (50, 100, and 500 mg anthracene/L per day) during 90 days. The second experiment took place after acclimatization (during 132 days). The results of anthracene removal were compared with controls in which no additional inoculum was added. During the two experiments, the behavior of pH, chemical oxygen demand (COD), and biogas production was evaluated. Results indicate that the bacterial community adapted for removal of anthracene became enriched through the acclimatization process. Anthracene biodegradation occurred in the soil model with both types of reactors (aerobic and anaerobic), but the rates and extent of biodegradation in the aerobic reactor were higher (95%) than those in anaerobic conditions (74%). Microbial activity also contributed to enhancing bioremediation in the soil by reducing anthracene sorption.     

Canine Recombinant Adenovirus Vector Induces an Immunogenicity-Related Gene Expression Profile in Skin-Migrated CD11b+ -Type DCs

Gene expression profiling of the blood cell response induced early after vaccination has previously been demonstrated to predict the immunogenicity of vaccines. In this study, we evaluated whether the analysis of the gene expression profile of skin-migrated dendritic cells (DCs) could be informative for the in vitro prediction of immunogenicity of vaccine, using canine adenovirus serotype 2 (CAV2) as vaccine vector. CAV2 has been shown to induce immunity to transgenes in several species including sheep and is an interesting alternative to human adenovirus-based vectors, based on the safety records of the parental strain in dogs and the lack of pre-existing immunity in non-host species. Skin-migrated DCs were collected from pseudo-afferent lymph in sheep. Both the CD11b⁺ -type and CD103⁺ -type skin-migrated DCs were transduced by CAV2. An analysis of the global gene response to CAV2 in the two skin DC subsets showed that the gene response in CD11b⁺ -type DCs was far higher and broader than in the CD103⁺ -type DCs. A newly released integrative analytic tool from Ingenuity systems revealed that the CAV2-modulated genes in the CD11b⁺ -type DCs clustered in several activated immunogenicity-related functions, such as immune response, immune cell trafficking and inflammation. Thus gene profiling in skin-migrated DC in vitro indicates that the CD11b⁺ DC type is more responsive to CAV2 than the CD103⁺ DC type, and provides valuable information to help in evaluating and possibly improving viral vector vaccine effectiveness.     

Sialyllactose in viral membrane gangliosides is a novel molecular recognition pattern for mature dendritic cell capture of HIV-1

HIV-1 is internalized into mature dendritic cells (mDCs) via an as yet undefined mechanism with subsequent transfer of stored, infectious virus to CD4+ T lymphocytes. Thus, HIV-1 subverts a DC antigen capture mechanism to promote viral spread. Here, we show that gangliosides in the HIV-1 membrane are the key molecules for mDC uptake. HIV-1 virus-like particles and liposomes mimicking the HIV-1 lipid composition were shown to use a common internalization pathway and the same trafficking route within mDCs. Hence, these results demonstrate that gangliosides can act as viral attachment factors, in addition to their well known function as cellular receptors for certain viruses. Furthermore, the sialyllactose molecule present in specific gangliosides was identified as the determinant moiety for mDC HIV-1 uptake. Thus, sialyllactose represents a novel molecular recognition pattern for mDC capture, and may be crucial both for antigen presentation leading to immunity against pathogens and for succumbing to subversion by HIV-1.     

Development of a panel of genome-wide ancestry informative markers to study admixture throughout the Americas

Most individuals throughout the Americas are admixed descendants of Native American, European, and African ancestors. Complex historical factors have resulted in varying proportions of ancestral contributions between individuals within and among ethnic groups. We developed a panel of 446 ancestry informative markers (AIMs) optimized to estimate ancestral proportions in individuals and populations throughout Latin America. We used genome-wide data from 953 individuals from diverse African, European, and Native American populations to select AIMs optimized for each of the three main continental populations that form the basis of modern Latin American populations. We selected markers on the basis of locus-specific branch length to be informative, well distributed throughout the genome, capable of being genotyped on widely available commercial platforms, and applicable throughout the Americas by minimizing within-continent heterogeneity. We then validated the panel in samples from four admixed populations by comparing ancestry estimates based on the AIMs panel to estimates based on genome-wide association study (GWAS) data. The panel provided balanced discriminatory power among the three ancestral populations and accurate estimates of individual ancestry proportions (R2 > 0.9 for ancestral components with significant between-subject variance). Finally, we genotyped samples from 18 populations from Latin America using the AIMs panel and estimated variability in ancestry within and between these populations. This panel and its reference genotype information will be useful resources to explore population history of admixture in Latin America and to correct for the potential effects of population stratification in admixed samples in the region.     

Influence of level of zilpaterol chlorhydrate supplementation on growth performance and carcass characteristics of feedlot lambs

Forty-eight Pelibuey × Katahdin (38.8 ± 0.67 kg) crossbred male lambs were used in a 32-day feeding trial (four pens per treatment in a randomized complete block design), to evaluate the influence of zilpaterol (β₂-agonist) supplementation level on growth performance and carcass characteristics. Lambs were fed a dry-rolled corn-based finishing diet (3.04 Mcal/kg of ME) supplemented with 0, 0.15, 0.20, or 0.25 mg/kg of live weight d⁻¹ zilpaterol (as zilpaterol chlorhydrate, Zilmax^®, Intervet México, México City). DM intake averaged 1.099 ± 0.042 kg/d and was not affected (P = 0.40) by treatments. Compared with control lambs, zilpaterol supplementation increased gain efficiency (15.8%, P < 0.03), apparent energy retention per unit DMI (10.9%, P = 0.03), and tended to increased daily gain (16%, P < 0.07) and total gain (17.7%, P < 0.08). Zilpaterol supplementation did not affect (P = 0.20) carcass weight, longissimus muscle area (LM), or fat thickness, but increased (2.3%, P = 0.04) carcass dressing percentage and reduced (36%, P < 0.01) kidney-pelvic fat. Increasing level of zilpaterol supplementation increased total weight gain (linear component, P < 0.05), gain:feed (linear component, P < 0.01), and dressing percentage (linear component, P < 0.02), and decreased (linear component, P < 0.01) kidney-pelvic fat. We conclude that zilpaterol supplementation enhances growth performance and dressing percentage in lambs in a manner comparable to that of cattle (greater muscle accretion, reduced body fat). Responses to zilpaterol was optimal when supplemented at 0.20 mg of zilpaterol/kg of live weight d⁻¹.     

In silico protein fragmentation reveals the importance of critical nuclei on domain reassembly

Protein complementation assays (PCAs) based on split protein fragments have become powerful tools that facilitate the study and engineering of intracellular protein-protein interactions. These assays are based on the observation that a given protein can be split into two inactive fragments and these fragments can reassemble into the original properly folded and functional structure. However, one experimentally observed limitation of PCA systems is that the folding of a protein from its fragments is dramatically slower relative to that of the unsplit parent protein. This is due in part to a poor understanding of how PCA design parameters such as split site position in the primary sequence and size of the resulting fragments contribute to the efficiency of protein reassembly. We used a minimalist on-lattice model to analyze how the dynamics of the reassembly process for two model proteins was affected by the location of the split site. Our results demonstrate that the balanced distribution of the “folding nucleus,” a subset of residues that are critical to the formation of the transition state leading to productive folding, between protein fragments is key to their reassembly.     

The family 52 beta-xylosidase from Geobacillus stearothermophilus is a dimer: structural and biophysical characterization of a glycoside hydrolase

Xylans are the most abundant polysaccharides forming the plant cell wall hemicelluloses, and they are degraded, among other proteins, by beta-xylosidase enzymes. In this work, the structural and biophysical properties of the family 52 beta-xylosidase from Geobacillus stearothermophilus, XynB2, are described. Size exclusion chromatography, analytical centrifugation, ITC, CD, fluorescence (steady state and ANS-binding) and FTIR were used to obtain the structure, the oligomerization state and the conformational changes of XynB2, as pH, chemical denaturants or temperature were modified. This report describes the first extensive conformational characterization of a family 52 beta-xylosidase. The active protein was a highly hydrated dimer, whose active site was formed by the two protomers, and it probably involved aromatic residues. At low pH, the protein was not active and it populated a monomeric molten-globule-like species, which had a conformational transition with a pK(a) of approximately 4.0. Thermal and chemical-denaturations of the native protein showed hysteresis behaviour. The protein at physiological pH was formed by alpha-helix (30%) and beta-sheet (30%), as shown by CD and FTIR. Comparison with other xylosidases of the same family indicates that the percentages of secondary structure seem to be conserved among the members of the family.