Evaluation of a novel liquid stabilised peste des petits ruminants vaccine: Safety and immunogenic efficacy in sheep and goats in the field in Jordan.

A novel liquid stabiliser was tested with the Nigeria 75/1 Peste des Petit Ruminants (PPR) vaccine over two field studies carried out in sheep and goats. PPR seronegative sheep and goats were selected from farms surrounding Amman, Jordan and were vaccinated with either a stabilised liquid PPR vaccine that had been formulated 3 months prior to use and stored at 2-8 °C or a reconstituted lyophilised PPRV vaccine reconstituted on the day of vaccination. Sera were taken immediately before vaccination and at approximately 1.5, 3 and 6 months following vaccination, then subsequently tested using IDVet ID Screen® PPR competition ELISA and Serum Neutralisation tests to determine the presence of PPRV anti-N antibodies and neutralising antibodies, respectively. It was observed that the liquid-stabilised vaccine was able to provide comparable antibody responses in both species to those induced by the lyophilized vaccine. The ability to store liquid stabilised PPRV vaccine for field use would positively impact PPRV eradication efforts.

International consensus conference recommendations on ultrasound education for undergraduate medical students.

OBJECTIVES: The purpose of this study is to provide expert consensus recommendations to establish a global ultrasound curriculum for undergraduate medical students. METHODS: 64 multi-disciplinary ultrasound experts from 16 countries, 50 multi-disciplinary ultrasound consultants, and 21 medical students and residents contributed to these recommendations. A modified Delphi consensus method was used that included a systematic literature search, evaluation of the quality of literature by the GRADE system, and the RAND appropriateness method for panel judgment and consensus decisions. The process included four in-person international discussion sessions and two rounds of online voting. RESULTS: A total of 332 consensus conference statements in four curricular domains were considered: (1) curricular scope (4 statements), (2) curricular rationale (10 statements), (3) curricular characteristics (14 statements), and (4) curricular content (304 statements). Of these 332 statements, 145 were recommended, 126 were strongly recommended, and 61 were not recommended. Important aspects of an undergraduate ultrasound curriculum identified include curricular integration across the basic and clinical sciences and a competency and entrustable professional activity-based model. The curriculum should form the foundation of a life-long continuum of ultrasound education that prepares students for advanced training and patient care. In addition, the curriculum should complement and support the medical school curriculum as a whole with enhanced understanding of anatomy, physiology, pathophysiological processes and clinical practice without displacing other important undergraduate learning. The content of the curriculum should be appropriate for the medical student level of training, evidence and expert opinion based, and include ongoing collaborative research and development to ensure optimum educational value and patient care. CONCLUSIONS: The international consensus conference has provided the first comprehensive document of recommendations for a basic ultrasound curriculum. The document reflects the opinion of a diverse and representative group of international expert ultrasound practitioners, educators, and learners. These recommendations can standardize undergraduate medical student ultrasound education while serving as a basis for additional research in medical education and the application of ultrasound in clinical practice.

Response of Key Metabolites during a UV-A Exposure Time-Series in the Cyanobacterium Chlorogloeopsis fritschii PCC 6912.

Ultraviolet A (UV-A) is the major component of UV radiation reaching the Earth's surface, causing indirect damage to photosynthetic organisms via the production of reactive oxygen species (ROS). In comparison, UV-B causes both direct damage to biomolecules and indirect damage. UV-B is well studied in cyanobacterial research due to their long evolutionary history and adaptation to high levels of UV, with less work on the effects of UV-A. In this study, the response of key metabolites in Chlorogloeopsis fritschii (C. fritschii) during 48 h of photosynthetically active radiation (PAR, 15 µmol·m-2·s-1) supplemented with UV-A (11 µmol·m-2·s-1) was investigated using gas chromatography- mass spectrometry (GC-MS). Results showed an overall significant increase in metabolite levels up to 24 h of UV-A exposure. Compared with previously reported UV-B (PAR + UV-B) and PAR only results, UV-A showed more similarity compared to PAR only exposure as opposed to supplemented UV-B. The amino acids glutamate, phenylalanine and leucine showed differences in levels between UV (both supplemented UV-A and supplemented UV-B) and PAR only (non-supplemented PAR), hinting to their relevance in UV stress response. The fatty acids, palmitic and stearic acid, showed positive log2 fold-change (FC) in supplemented UV-A and PAR only experiments but negative log2 FC in UV-B, indicating the more harmful effect of UV-B on primary metabolism. Less research has been conducted on UV-A exposure and cyanobacteria, a potential environmental stimuli for the optimisation of metabolites for industrial biotechnology. This study will add to the literature and knowledge on UV-A stress response at the metabolite level in cyanobacteria, especially within the less well-known species C. fritschii.

A Potent and Selective Kallikrein-5 Inhibitor Delivers High Pharmacological Activity in Skin from Patients with Netherton Syndrome.

Regulation of proteolytic activity in the skin plays a pivotal role in epidermal homeostasis. This is best exemplified in Netherton syndrome, a severe genetic skin condition caused by loss-of-function mutations in the gene serine protease inhibitor Kazal-type 5 encoding lympho-epithelial Kazal-type-related inhibitor, a serine protease inhibitor that regulates kallikrein (KLK)-related peptidase 5, 7, and 14 activities. KLK5 plays a central role in stratum corneum shedding and inflammatory cell signaling, activates KLK7 and KLK14, and is therefore an optimal therapeutic target. We aimed to identify a potent and selective small-molecule inhibitor of KLK5 amenable to epidermal delivery. GSK951 was identified using a structure-based design strategy and showed a half maximal inhibitory concentration of 250 pM for KLK5 and greater than 100-fold selectivity over KLK7 and KLK14. Cocrystal structure analysis identified the critical catalytic site interactions to a surrogate for KLK5. Topical application of GSK951-containing cream inhibited KLK5 activity in TgKLK5 mouse skin, reduced transepidermal water loss, and decreased proinflammatory cytokine expression. GSK951 achieved high concentrations in healthy human epidermis following topical application in a cream formulation. Finally, KLK5 protease activity was increased in stratum corneum of patients with Netherton syndrome and significantly inhibited by GSK951. These findings unveil a KLK5-specific small-molecule inhibitor with a high therapeutic potential for patients with Netherton syndrome.

In Vivo Half-Life Extension of BMP1/TLL Metalloproteinase Inhibitors Using Small-Molecule Human Serum Albumin Binders.

Reducing the required frequence of drug dosing can improve the adherence of patients to chronic treatments. Hence, drugs with longer in vivo half-lives are highly desirable. One of the most promising approaches to extend the in vivo half-life of drugs is conjugation to human serum albumin (HSA). In this work, we describe the use of AlbuBinder 1, a small-molecule noncovalent HSA binder, to extend the in vivo half-life and pharmacology of small-molecule BMP1/TLL inhibitors in humanized mice (HSA KI/KI). A series of conjugates of AlbuBinder 1 with BMP1/TLL inhibitors were prepared. In particular, conjugate c showed good solubility and a half-life extension of >20-fold versus the parent molecule in the HSA KI/KI mice, reaching half-lives of >48 h with maintained maximal inhibition of plasma BMP1/TLL. The same conjugate showed a half-life of only 3 h in the wild-type mice, suggesting that the half-life extension was principally due to specific interactions with HSA. It is envisioned that conjugation to AlbuBinder 1 should be applicable to a wide range of small molecule or peptide drugs with short half-lives. In this context, AlbuBinders represent a viable alternative to existing half-life extension technologies.

Dose-dependent response to infection with SARS-CoV-2 in the ferret model and evidence of protective immunity.

There is a vital need for authentic COVID-19 animal models to enable the pre-clinical evaluation of candidate vaccines and therapeutics. Here we report a dose titration study of SARS-CoV-2 in the ferret model. After a high (5 × 106 pfu) and medium (5 × 104 pfu) dose of virus is delivered, intranasally, viral RNA shedding in the upper respiratory tract (URT) is observed in 6/6 animals, however, only 1/6 ferrets show similar signs after low dose (5 × 102 pfu) challenge. Following sequential culls pathological signs of mild multifocal bronchopneumonia in approximately 5-15% of the lung is seen on day 3, in high and medium dosed groups. Ferrets re-challenged, after virus shedding ceased, are fully protected from acute lung pathology. The endpoints of URT viral RNA replication & distinct lung pathology are observed most consistently in the high dose group. This ferret model of SARS-CoV-2 infection presents a mild clinical disease.

Development of a small molecule that corrects misfolding and increases secretion of Z α1 -antitrypsin.

Severe α1 -antitrypsin deficiency results from the Z allele (Glu342Lys) that causes the accumulation of homopolymers of mutant α1 -antitrypsin within the endoplasmic reticulum of hepatocytes in association with liver disease. We have used a DNA-encoded chemical library to undertake a high-throughput screen to identify small molecules that bind to, and stabilise Z α1 -antitrypsin. The lead compound blocks Z α1 -antitrypsin polymerisation in vitro, reduces intracellular polymerisation and increases the secretion of Z α1 -antitrypsin threefold in an iPSC model of disease. Crystallographic and biophysical analyses demonstrate that GSK716 and related molecules bind to a cryptic binding pocket, negate the local effects of the Z mutation and stabilise the bound state against progression along the polymerisation pathway. Oral dosing of transgenic mice at 100 mg/kg three times a day for 20 days increased the secretion of Z α1 -antitrypsin into the plasma by sevenfold. There was no observable clearance of hepatic inclusions with respect to controls over the same time period. This study provides proof of principle that "mutation ameliorating" small molecules can block the aberrant polymerisation that underlies Z α1 -antitrypsin deficiency.

Myelination of peripheral nerves is controlled by PI4KB through regulation of Schwann cell Golgi function.

Better understanding myelination of peripheral nerves would benefit patients affected by peripheral neuropathies, including Charcot-Marie-Tooth disease. Little is known about the role the Golgi compartment plays in Schwann cell (SC) functions. Here, we studied the role of Golgi in myelination of peripheral nerves in mice through SC-specific genetic inactivation of phosphatidylinositol 4-kinase beta (PI4KB), a Golgi-associated lipid kinase. Sciatic nerves of such mice showed thinner myelin of large diameter axons and gross aberrations in myelin organization affecting the nodes of Ranvier, the Schmidt-Lanterman incisures, and Cajal bands. Nonmyelinating SCs showed a striking inability to engulf small diameter nerve fibers. SCs of mutant mice showed a distorted Golgi morphology and disappearance of OSBP at the cis-Golgi compartment, together with a complete loss of GOLPH3 from the entire Golgi. Accordingly, the cholesterol and sphingomyelin contents of sciatic nerves were greatly reduced and so was the number of caveolae observed in SCs. Although the conduction velocity of sciatic nerves of mutant mice showed an 80% decrease, the mice displayed only subtle impairment in their motor functions. Our analysis revealed that Golgi functions supported by PI4KB are critically important for proper myelination through control of lipid metabolism, protein glycosylation, and organization of microvilli in the nodes of Ranvier of peripheral nerves.

Ethologically based behavioural and neurochemical characterisation of mice with isoform-specific loss of dysbindin-1A in the context of schizophrenia.

Dysbindin-1 is implicated in several aspects of schizophrenia, including cognition and both glutamatergic and dopaminergic neurotransmission. Targeted knockout of dysbindin-1A (Dys-1A KO), the most abundant and widely expressed isoform in the brain, is associated with deficits in delay/interference-dependent working memory. Using an ethologically based approach, the following behavioural phenotypes were examined in Dys-1A KO mice: exploratory activity, social interaction, anxiety and problem-solving ability. Levels of monoamines and their metabolites were measured in striatum, hippocampus and prefrontal cortex using high-performance liquid chromatography with electrochemical detection. The ethogram of initial exploration in Dys-1A KO mice was characterised by increased rearing from a seated position; over subsequent habituation, stillness was decreased relative to wildtype. In a test of dyadic social interaction with an unfamiliar conspecific in a novel environment, female KO mice showed an increase in investigative social behaviours. Marble burying behaviour was unchanged. Using the puzzle-box test to measure general problem-solving performance, no effect of genotype was observed across nine trials of increasing complexity. Dys-1A KO demonstrated lower levels of 5-HT in ratio to its metabolite 5-HIAA in the prefrontal cortex. These studies elaborate the behavioural and neurochemical phenotype of Dys-1A KO mice, revealing subtle genotype-related differences in non-social and social exploratory behaviours and habituation of exploration in a novel environment, as well as changes in 5-HT activity in brain areas related to schizophrenia.

Disease Modification in Alzheimer's Disease: Current Thinking.

Alzheimer's disease (AD) has increasingly been recognized as a huge unmet medical need. Currently, there is no approved drug to cure, prevent, or even slow down the disease. It is imperative to develop disease-modifying treatments for AD to alter the underlying disease progression. This paper reviews the most up-to-date regulatory guidance on how to demonstrate disease modification and provides an overview of available methodologies and applications to clinical trials. The intent is to assist the field with future clinical trials designed to demonstrate disease-modifying effect in AD. The methodologies may be generalizable to broader neurodegenerative diseases.

Inflatable penile prosthesis implantation into scarred corporal bodies: timing may decrease postoperative problems.

OBJECTIVES: To compare the rates of intra- and postoperative complications between early (<4 months) or delayed (>4 months) inflatable penile prosthesis (IPP) re-implantation performed after a fibrosis-inducing event. PATIENTS AND METHODS: Cases of IPP re-implantation were performed by a single, high-volume prosthetic surgeon in hospitals domestically and abroad over a 15-year period (2001-2016). The time interval from the precipitating event to operation, intra-operative complications and postoperative complications for up to 1 year after implantation, were recorded. RESULTS: A total of 72 cases of IPP re-implantation were recorded. There were 16 (53%) intraoperative complications in patients who had surgery at <4 months (Group 1) and 23 (55%) in patients operated at >4 months (Group 2). There were four (13%) postoperative complications in Group 1 and 14 (33%) in Group 2. CONCLUSION: Re-implantation of an IPP into fibrotic corporal bodies is associated with less serious postoperative complications when performed within 4 months of the fibrosis-inducing event.

Acute stress in adolescence vs early adulthood following selective deletion of dysbindin-1A: Effects on anxiety, cognition and other schizophrenia-related phenotypes.

BACKGROUND: As exposure to stress has been linked to the onset and maintenance of psychotic illness, its pathogenesis may involve environmental stressors interacting with genetic vulnerability. AIM: To establish whether acute stress interacts with a targeted mutation of the gene encoding the neurodevelopmental factor dystrobrevin-binding protein 1 (DTNBP1), resulting in a specific loss of the isoform dysbindin-1A, to influence schizophrenia-relevant phenotypes in mice during adolescence and adulthood. METHODS: Male and female mice with a heterozygous or homozygous deletion of DTNBP1 were assessed in the open field test following acute restraint stress in adolescence (Day 35) and young adulthood (Day 60-70). Effects of acute restraint stress on memory retention in the novel object recognition test was also assessed in adulthood. Baseline corticosterone was measured in serum samples and, brain-derived neurotrophic factor (BDNF), glucocorticoid and mineralocorticoid receptor gene expression levels were measured in the hippocampus of adult mice. RESULTS: In the open field, deletion of dysbindin-1A induced hyperactivity and attenuated the action of stress to reduce hyperactivity in adolescence but not in adulthood; in females deletion of dysbindin-1A attenuated the effect of acute stress to increase anxiety-related behaviour in adolescence but not in adulthood. In the novel object recognition test, deletion of dysbindin-1A impaired memory and also revealed an increase in anxiety-related behaviour and a decrease in hippocampal BDNF gene expression in males. CONCLUSIONS: These data suggest that deletion of dysbindin-1A influences behaviours related to schizophrenia and anxiety more robustly in adolescence than in adulthood and that dysbindin-1A influences stress-related responses in a sex-dependent manner.

Direct measurement of swimming and diving kinematics of giant Atlantic bluefin tuna (Thunnus thynnus).

Tunas possess a range of physiological and mechanical adaptations geared towards high-performance swimming that are of considerable interest to physiologists, ecologists and engineers. Advances in biologging have provided significant improvements in understanding tuna migrations and vertical movement patterns, yet our understanding of the locomotion and swimming mechanics of these fish under natural conditions is limited. We equipped Atlantic bluefin tuna (Thunnus thynnus) with motion-sensitive tags and video cameras to quantify the gaits and kinematics used by wild fish. Our data reveal significant variety in the locomotory kinematics of Atlantic bluefin tuna, ranging from continuous locomotion to two types of intermittent locomotion. The tuna sustained swimming speeds in excess of 1.5 m s-1 (0.6 body lengths s-1), while beating their tail at a frequency of approximately 1 Hz. While diving, some descents were entirely composed of passive glides, with slower descent rates featuring more gliding, while ascents were primarily composed of active swimming. The observed swimming behaviour of Atlantic bluefin tuna is consistent with theoretical models predicting such intermittent locomotion to result in mechanical and physiological advantages. Our results confirm that Atlantic bluefin tuna possess behavioural specializations to increase their locomotory performance, which together with their unique physiology improve their capacity to use pelagic and mesopelagic habitats.

Intracellular and Extracellular Metabolites from the Cyanobacterium Chlorogloeopsis fritschii, PCC 6912, During 48 Hours of UV-B Exposure.

Cyanobacteria have many defence strategies to overcome harmful ultraviolet (UV) stress including the production of secondary metabolites. Metabolomics can be used to investigate this altered metabolism via targeted and untargeted techniques. In this study we assessed the changes in the intra- and extracellular low molecular weight metabolite levels of Chlorogloeopsis fritschii (C. fritschii) during 48 h of photosynthetically active radiation (PAR) supplemented with UV-B (15 µmol m-2 s-1 of PAR plus 3 µmol m-2 s-1 of UV-B) and intracellular levels during 48 h of PAR only (15 µmol m-2 s-1) with sampling points at 0, 2, 6, 12, 24 and 48 h. Gas chromatography-mass spectrometry (GC-MS) was used as a metabolite profiling tool to investigate the global changes in metabolite levels. The UV-B time series experiment showed an overall significant reduction in intracellular metabolites involved with carbon and nitrogen metabolism such as the amino acids tyrosine and phenylalanine which have a role in secondary metabolite production. Significant accumulation of proline was observed with a potential role in stress mitigation as seen in other photosynthetic organisms. 12 commonly identified metabolites were measured in both UV-B exposed (PAR + UV-B) and PAR only experiments with differences in significance observed. Extracellular metabolites (PAR + UV-B) showed accumulation of sugars as seen in other cyanobacterial species as a stress response to UV-B. In conclusion, a snapshot of the metabolome of C. fritschii was measured. Little work has been undertaken on C. fritschii, a novel candidate for use in industrial biotechnology, with, to our knowledge, no previous literature on combined intra- and extracellular analysis during a UV-B treatment time-series. This study is important to build on experimental data already available for cyanobacteria and other photosynthetic organisms exposed to UV-B.

Estimating Natural Mortality of Atlantic Bluefin Tuna Using Acoustic Telemetry.

Atlantic bluefin tuna (Thunnus thynnus) are highly migratory fish with a contemporary range spanning the North Atlantic Ocean. Bluefin tuna populations have undergone severe decline and the status of the fish within each population remains uncertain. Improved biological knowledge, particularly of natural mortality and rates of mixing of the western (GOM) and eastern (Mediterranean) populations, is key to resolving the current status of the Atlantic bluefin tuna. We evaluated the potential for acoustic tags to yield empirical estimates of mortality and migration rates for long-lived, highly migratory species such as Atlantic bluefin tuna. Bluefin tuna tagged in the Gulf of St. Lawrence (GSL) foraging ground (2009-2016) exhibited high detection rates post release, with 91% crossing receiver lines one year post tagging, 61% detected after year two at large, with detections up to ~1700 days post deployment. Acoustic detections per individual fish ranged from 3 to 4759 receptions. A spatially-structured Bayesian mark recapture model was applied to the acoustic detection data for Atlantic bluefin tuna electronically tagged in the GSL to estimate the rate of instantaneous annual natural mortality. We report a median estimate of 0.10 yr-1 for this experiment. Our results demonstrate that acoustic tags can provide vital fisheries independent estimates for life history parameters critical for improving stock assessment models.

Phos-tag analysis of Rab10 phosphorylation by LRRK2: a powerful assay for assessing kinase function and inhibitors.

Autosomal dominant mutations that activate the leucine-rich repeat kinase 2 (LRRK2) cause inherited Parkinson's disease. Recent work has revealed that LRRK2 directly phosphorylates a conserved threonine/serine residue in the effector-binding switch-II motif of a number of Rab GTPase proteins, including Rab10. Here we describe a facile and robust method to assess phosphorylation of endogenous Rab10 in mouse embryonic fibroblasts (MEFs), lung and spleen-derived B-cells, based on the ability of the Phos-tag reagent to retard the electrophoretic mobility of LRRK2-phosphorylated Rab10. We exploit this assay to show that phosphorylation of Rab10 is ablated in kinase-inactive LRRK2[D2017A] knockin MEFs and mouse lung, demonstrating that LRRK2 is the major Rab10 kinase in these cells/tissue. We also establish that the Phos-tag assay can be deployed to monitor the impact that activating LRRK2 pathogenic (G2019S and R1441G) knockin mutations have on stimulating Rab10 phosphorylation. We show that upon addition of LRRK2 inhibitors, Rab10 is dephosphorylated within 1-2 min, markedly more rapidly than the Ser(935) and Ser(1292) biomarker sites that require 40-80 min. Furthermore, we find that phosphorylation of Rab10 is suppressed in LRRK2[S910A+S935A] knockin MEFs indicating that phosphorylation of Ser(910) and Ser(935) and potentially 14-3-3 binding play a role in facilitating the phosphorylation of Rab10 by LRRK2 in vivo The Rab Phos-tag assay has the potential to significantly aid with evaluating the effect that inhibitors, mutations and other factors have on the LRRK2 signalling pathway.

Methods of conducting effective outreach to private well owners - a literature review and model approach.

Educational outreach programs have the potential to increase the occurrence of private well testing and maintenance behaviors, but are not always able to successfully engage the intended audience and overcome their barriers to change. We conducted a review of literature regarding behavior change and risk communication to identify common barriers to private well stewardship and motivational strategies to encourage change, as well as best practices for communicating with well owners. Results indicated that no specific strategy will be appropriate for all audiences, as different groups of well owners will have different barriers to change. For this reason, educators must develop an understanding of their audience so they are able to identify the most significant barriers to change and select motivational strategies that will directly reduce barriers. Implications for private well outreach programs are discussed.