Review: Recent advances in bovine in vitro embryo production: reproductive biotechnology history and methods

In vitro production (IVP) of embryos and associated technologies in cattle have shown significant progress in recent years, in part driven by a better understanding of the full potential of these tools by end users. The combination of IVP with sexed semen (SS) and genomic selection (GS) is being successfully and widely used in North America, South America and Europe. The main advantages offered by these technologies include a higher number of embryos and pregnancies per unit of time, and a wider range of potential female donors from which to retrieve oocytes (including open cyclic females and ones up to 3 months pregnant), including high index genomic calves, a reduced number of sperm required to produce embryos and increased chances of obtaining the desired sex of offspring. However, there are still unresolved aspects of IVP of embryos that limit a wider implementation of the technology, including potentially reduced fertility from the use of SS, reduced oocyte quality after in vitro oocyte maturation and lower embryo cryotolerance, resulting in reduced pregnancy rates compared to in vivo–produced embryos. Nevertheless, promising research results have been reported, and work is in progress to address current deficiencies. The combination of GS, IVP and SS has proven successful in the commercial field in several countries assisting practitioners and cattle producers to improve reproductive performance, efficiency and genetic gain.     

Synthesis of gold nanoparticles using Crinum macowanii bulb extracts and the application of these materials in blood detections at crime scenes

We here in report the synthesis of gold nanoparticles (AuNPs) using a Crinum macowanii bulb water extract. The as‐synthesized AuNPs were characterized using ultraviolet–visible spectroscopy, Fourier transform infrared spectroscopy, X‐ray diffraction, transmission electron microscopy, and a zeta potential‐sizer. The results showed that the as‐synthesized AuNPs were crystalline and mostly spherical in shape with a small mixture of triangular, tetrahedral, hexagonal, octagonal, and diamond shapes. The as‐synthesized AuNPs together with those synthesized by conventional methods were subsequently used as enhancers for the luminol signal in blood detection. It was noted that the AuNPs synthesized from the Crinum macowanii bulb water extract could enhance the chemiluminescence signal for blood detection by luminol to the same extent as AuNPs prepared by conventional methods. Furthermore, both types of AuNPs served as fluorescence enhancers for blood detection when luminol was replaced with the bulb water extract.     

Colorimetric paper bioassay by horseradish peroxidase for the detection of catechol and resorcinol in aqueous samples

Abstract

Phenolic compounds such as catechol and resorcinol are toxic and persistent pollutants in the aqueous environment. Detection procedures such as chromatographic and spectrophotometric methods are time-consuming and require sophisticated instruments with skilled manpower. Development of a simple, cost effective, portable and disposable paper based biosensor could be a better alternative to the conventional methods. The present study attempted to develop a paper based biosensor by immobilizing horseradish peroxidase enzyme to detect catechol and resorcinol in aqueous samples. Horseradish peroxidase catalyzes the oxidation of phenolic compounds to semiquinones, which on reaction with a chromogen, 3-methyl 2-benzothiazolinone hydrazine (MBTH) gives faint pink to red color depending on the compound and its concentration in the sample is the basis for biosensing application. Different methods of enzyme immobilization on filter paper like physical adsorption, covalent coupling, and polysaccharide entrapment were executed. The performance of the various enzyme immobilization methods was evaluated by analyzing the developed color intensity using ImageJ software. Entrapment technique is the most effective method of immobilizing enzyme on the filter paper that produces the highest color intensity with better stability. The visible limit of detection (LoD) was observed as 0.45?mM (50?mg/L) for catechol and 0.09?mM (10?mg/L) for resorcinol in aqueous samples.     

Quality evaluation of Saudi honey harvested from the Asir province by using high-performance liquid chromatography (HPLC)

Sugar profile and hydroxymethylfurfural (HMF) of Saudi honey were examined through high-performance liquid chromatography (HPLC) system equipped with refractive index and diode array detectors. The work was designed to assess the quality of various types of blossom honey i.e. Sider (Ziziphus spina-christi), Dhuhyana (Acacia asak), Sumra (Acacia tortilis), Qatada (Acacia hamulosa), Dhurum (Lavandula dentata), multiflora with majra (Hypoestes forskaolii), multiflora with herbs, Keena (Eucalyptus spp.) produced in the southwestern areas of the kingdom. Hierarchical cluster analysis (HCA), principal cluster analysis (PCA), and similarity and difference indices (SDI) were also applied to examine the possible grouping based on the studied quality parameters. Four main sugars (two monosaccharides i.e. fructose and glucose, two disaccharides i.e. sucrose and maltose) and HMF were investigated . The average values of fructose and glucose were in the range 33.10%–44.77% and 26.68%–37.91%, respectively. The maltose was present in all types of honey and its mean values were in the range of 0.37%–2.97%, while sucrose was absent in six types of honey, 0.25% in one unifloral honey, and 3.25% in one multi-floral honey. HMF was not detected in seven types of honey but was below the limit of quantification (0.13 mg/kg) in one type of honey. PCA displayed the accumulative variance of 79.96% for the initial two PCs suggesting that honey samples were not well distinguished by their sugar profile. Based on the sucrose and HMF contents, it was concluded that all types of blossom honey from the Asir province were of the best quality in the kingdom and met the international quality parameters.     

Evaluation of quenching methods for metabolite recovery in photoautotrophic Synechococcus sp. PCC 7002

The first step of many metabolomics studies is quenching, a technique vital for rapidly halting metabolism and ensuring that the metabolite profile remains unchanging during sample processing. The most widely used approach is to plunge the sample into prechilled cold methanol; however, this led to significant metabolite loss in Synecheococcus sp. PCC 7002. Here we describe our analysis of the impacts of cold methanol quenching on the model marine cyanobacterium Synechococcus sp. PCC 7002, as well as our brief investigation of alternative quenching methods. We tested several methods including cold methanol, cold saline, and two filtration approaches. Targeted central metabolites were extracted and metabolomic profiles were generated using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results indicate that cold methanol quenching induces dramatic metabolite leakage in Synechococcus, resulting in a majority of central metabolites being lost prior to extraction. Alternatively, usage of a chilled saline quenching solution mitigates metabolite leakage and improves sample recovery without sacrificing rapid quenching of cellular metabolism. Finally, we illustrate that metabolite leakage can be assessed, and subsequently accounted for, in order to determine absolute metabolite pool sizes; however, our results show that metabolite leakage is inconsistent across various metabolite pools and therefore must be determined for each individually measured metabolite.     

An Integrative Structural Biology Analysis of Von Willebrand Factor Binding and Processing by ADAMTS-13 in Solution

Von Willebrand Factor (vWF), a 300-kDa plasma protein key to homeostasis, is cleaved at a single site by multi-domain metallopeptidase ADAMTS-13. vWF is the only known substrate of this peptidase, which circulates in a latent form and becomes allosterically activated by substrate binding. Herein, we characterised the complex formed by a competent peptidase construct (AD13-MDTCS) comprising metallopeptidase (M), disintegrin-like (D), thrombospondin (T), cysteine-rich (C), and spacer (S) domains, with a 73-residue functionally relevant vWF-peptide, using nine complementary techniques. Pull-down assays, gel electrophoresis, and surface plasmon resonance revealed tight binding with sub-micromolar affinity. Cross-linking mass spectrometry with four reagents showed that, within the peptidase, domain D approaches M, C, and S. S is positioned close to M and C, and the peptide contacts all domains. Hydrogen/deuterium exchange mass spectrometry revealed strong and weak protection for C/D and M/S, respectively. Structural analysis by multi-angle laser light scattering and small-angle X-ray scattering in solution revealed that the enzyme adopted highly flexible unbound, latent structures and peptide-bound, active structures that differed from the AD13-MDTCS crystal structure. Moreover, the peptide behaved like a self-avoiding random chain. We integrated the results with computational approaches, derived an ensemble of structures that collectively satisfied all experimental restraints, and discussed the functional implications. The interaction conforms to a ‘fuzzy complex’ that follows a ‘dynamic zipper’ mechanism involving numerous reversible, weak but additive interactions that result in strong binding and cleavage. Our findings contribute to illuminating the biochemistry of the vWF:ADAMTS-13 axis.     

Ultrasound shear wave elastography measurement of the deep posterior cervical muscles: Reliability and ability to differentiate between muscle contraction states

The deep posterior cervical muscles (DPCM), specifically the semispinalis cervicis and cervical multifidus, are often impaired in patients with neck disorders and have been assessed by several imaging techniques. Prior ultrasound shear wave elastography (SWE) imaging and reliability assessments of the DPCM were performed utilizing similar positioning as assessments for the more superficial cervical extensors. Our objectives were to describe an SWE imaging technique for the DPCM, establish intra-rater reliability of DPCM SWE, and compare DPCM shear modulus during rest and submaximal contraction in both prone and seated positions in individuals without spinal pain. In sixteen participants, the DPCM was located using B‐mode ultrasound, then muscle shear modulus was assessed via SWE at both rest and with contraction against a 2‐kg resistance applied at the C2 spinous process. Within‐day intra‐rater reliability was moderate to good (ICC = 0.70–0.88). The DPCM were stiffer during contraction than at rest in the prone position (p = 0.002), and at rest in sitting versus at rest in prone (p = 0.003). Further research is needed to assess DPCM-specific SWE in symptomatic individuals and compare DPCM shear modulus to electromyography across contraction intensities.     

Parallel speciation or long‐distance dispersal? Lessons from seaweeds (Fucus) in the Baltic Sea

Parallel evolution has been invoked as a forceful mechanism of ecotype and species formation in many animal taxa. However, parallelism may be difficult to separate from recently monophyletically diverged species that are likely to show complex genetic relationships as a result of considerable shared ancestral variation and secondary hybridization in local areas. Thus, species' degrees of reproductive isolation, barriers to dispersal and, in particular, limited capacities for long‐distance dispersal will affect demographical structures underlying mechanisms of divergent evolution. Here, we used nine microsatellite DNA markers to study intra‐ and interspecific genetic diversity of two recently diverged species of brown macroalgae, Fucus radicans (L. Bergström & L. Kautsky) and F. vesiculosus (Linnaeus), in the Baltic Sea. We further performed biophysical modelling to identify likely connectivity patterns influencing the species' genetic structures. For each species, we found intraspecific contrasting patterns of clonality incidence and population structure. In addition, strong genetic differentiation between the two species within each locality supported the existence of two distinct evolutionary lineages (F_ST = 0.15–0.41). However, overall genetic clustering analyses across both species' populations revealed that all populations from one region (Estonia) were more genetically similar to each other than to their own taxon from the other two regions (Sweden and Finland). Our data support a hypothesis of parallel speciation. Alternatively, Estonia may be the ancestral source of both species, but is presently isolated by oceanographic barriers to dispersal. Thus, a limited gene flow in combination with genetic drift could have shaped the seemingly parallel structure.     

Anisotropic adaptive finite element method for modelling blood flow

In this study, we present an adaptive anisotropic finite element method (FEM) and demonstrate how computational efficiency can be increased when applying the method to the simulation of blood flow in the cardiovascular system. We use the SUPG formulation for the transient 3D incompressible Navier–Stokes equations which are discretised by linear finite elements for both the pressure and the velocity field.

Given the pulsatile nature of the flow in blood vessels we have pursued adaptivity based on the average flow over a cardiac cycle. Error indicators are derived to define an anisotropic mesh metric field. Mesh modification algorithms are used to anisotropically adapt the mesh according to the desired size field. We demonstrate the efficiency of the method by first applying it to pulsatile flow in a straight cylindrical vessel and then to a porcine aorta with a stenosis bypassed by a graft. We demonstrate that the use of an anisotropic adaptive FEM can result in an order of magnitude reduction in computing time with no loss of accuracy compared to analyses obtained with uniform meshes.