A hypervariable intron of the STAYGREEN locus provides excellent discrimination among Pisum fulvum accessions and reveals evidence for a relatively recent hybridization event with Pisum sativum.

An analysis of 82 non-synonymous Pisum fulvum accessions for sequence variation in a fragment of the STAYGREEN (SGR) locus revealed 57 alleles, most of which differed in indel structure. Eight additional P. fulvum accessions, each supposedly synonymous with a different accession of the initial group, were also analyzed. In every case the paired synonymous accessions possessed the same SGR sequence but varied slightly for a 6-trait morphological phenotype, indicating that SGR sequence is a much more reliable indicator of accession identity than is a morphological characterization. SGR sequence analysis confirmed our previous finding that P. fulvum accessions separate into two allele groups. This division was not supported by results of previous studies that were based on sequences distributed across the entire genome, suggesting that the division may have been produced by selection at a nearby locus and that the SGR phylogeny may not be good indicator of overall relationships within the species. One P. fulvum accession, PI 595941 (=JI1796), displayed an SGR sequence outside the variation typical of the species. Instead, its allele resembled alleles limited to a set of Pisum sativum landraces from the Middle East, suggesting hybridization between ancestors of PI 595941 and some primitive form of domesticated P. sativum. With one exception from the extreme northwest corner of Israel, P. fulvum accessions collected north of latitude 35.5° N were fixed for alleles from group A. These northern accessions also displayed greatly reduced SGR sequence diversity compared to group A accessions collected from other regions, suggesting that the northern populations may represent recent extensions of the range of the species. Group B accessions were distributed from Lake Tiberias south and were generally sympatric with the southern group A accessions. Although group B accessions occupied a smaller area than group A, the SGR sequence diversity in this group (28 alleles in 33 accessions) exceeded that for group A.

Endothelial cell activation, Weibel-Palade body secretion, and enhanced angiogenesis in severe COVID-19.

Background: Severe COVID-19 is associated with marked endothelial cell (EC) activation that plays a key role in immunothrombosis and pulmonary microvascular occlusion. However, the biological mechanisms through which SARS-CoV-2 causes EC activation and damage remain poorly defined. Objectives: We investigated EC activation in patients with acute COVID-19, and specifically focused on how proteins stored within Weibel-Palade bodies may impact key aspects of disease pathogenesis. Methods: Thirty-nine patients with confirmed COVID-19 were recruited. Weibel-Palade body biomarkers (von Willebrand factor [VWF], angiopoietin-2 [Angpt-2], and osteoprotegerin) and soluble thrombomodulin (sTM) levels were determined. In addition, EC activation and angiogenesis were assessed in the presence or absence of COVID-19 plasma incubation. Results: Markedly elevated plasma VWF antigen, Angpt-2, osteoprotegerin, and sTM levels were observed in patients with acute COVID-19. The increased levels of both sTM and Weibel-Palade body components (VWF, osteoprotegerin, and Angpt-2) correlated with COVID-19 severity. Incubation of COVID-19 plasma with ECs triggered enhanced VWF secretion and increased Angpt-2 expression, as well as significantly enhanced in vitro EC tube formation and angiogenesis. Conclusion: We propose that acute SARS-CoV-2 infection leads to a complex and multifactorial EC activation, progressive loss of thrombomodulin, and increased Angpt-2 expression, which collectively serve to promote a local proangiogenic state.

First wave of SARS-CoV-2 in Santiago Chile: Seroprevalence, asymptomatic infection and infection fatality rate.

BACKGROUND: The first wave of SARS-CoV-2 infection in Chile occurred during the cold season reaching a peak by the end of June 2020, with 80 % of the cases concentrated in its capital, Santiago. The main objective of this study was to estimate the attack rate during this first wave of SARS-CoV-2 in a large, densely populated city with more than seven million inhabitants. Since the number of confirmed cases provides biased information due to individuals' potential self-selection, mostly related to asymptomatic patients and testing access, we measured antibodies against SARS-CoV-2 to assess infection prevalence during the first wave in the city, as well as estimate asymptomatic cases, and infection fatality ratio. To our knowledge this is one of the few population-based cross-sectional serosurvey during the first wave in a highly affected emerging country. The challenges of pandemic response in urban settings in a capital city like Santiago, with heterogeneous subpopulations and high mobility through public transportation, highlight the necessity of more accurate information regarding the first waves of new emerging diseases. METHODS: From April 24 to June 21, 2020, 1326 individuals were sampled from a long-standing panel of household representatives of Santiago. Immunochromatographic assays were used to detect IgM and IgG antibody isotypes. RESULTS: Seroprevalence reached 6.79 % (95 %CI 5.58 %-8.26 %) in the first 107 days of the pandemic, without significant differences among sex and age groups; this figure indicates an attack rate 2.8 times higher than the one calculated with registered cases. It also changes the fatality rate estimates, from a 2.33 % case fatality rate reported by MOH to an estimated crude 1.00 % (CI95 % 0.97-1.03) infection fatality rate (adjusted for test performance 1.66 % [CI95 % 1.61-1.71]). Most seropositive were symptomatic (81,1 %). CONCLUSIONS: Despite the high number of cases registered, mortality rates, and the stress produced over the health system, the vast majority of the people remained susceptible to potential new epidemic waves. We contribute to the understanding of the initial spread of emerging epidemic threats. Consequently, our results provide better information to design early strategies that counterattack new health challenges in urban contexts.

An anaplerotic approach to correct the mitochondrial dysfunction in ataxia-telangiectasia (A-T).

BACKGROUND: ATM, the protein defective in the human genetic disorder, ataxia-telangiectasia (A-T) plays a central role in response to DNA double-strand breaks (DSBs) and in protecting the cell against oxidative stress. We showed that A-T cells are hypersensitive to metabolic stress which can be accounted for by a failure to exhibit efficient endoplasmic reticulum (ER)-mitochondrial signalling and Ca2+ transfer in response to nutrient deprivation resulting in mitochondrial dysfunction. The objective of the current study is to use an anaplerotic approach using the fatty acid, heptanoate (C7), a metabolic product of the triglyceride, triheptanoin to correct the defect in ER-mitochondrial signalling and enhance cell survival of A-T cells in response to metabolic stress. METHODS: We treated control cells and A-T cells with the anaplerotic agent, heptanoate to determine their sensitivity to metabolic stress induced by inhibition of glycolysis with 2- deoxyglucose (2DG) using live-cell imaging to monitor cell survival for 72 h using the Incucyte system. We examined ER-mitochondrial signalling in A-T cells exposed to metabolic stress using a suite of techniques including immunofluorescence staining of Grp75, ER-mitochondrial Ca2+ channel, the VAPB-PTPIP51 ER-mitochondrial tether complexes as well as proximity ligation assays between Grp75-IP3R1 and VAPB1-PTPIP51 to establish a functional interaction between ER and mitochondria. Finally, we also performed metabolomic analysis using LC-MS/MS assay to determine altered levels of TCA intermediates A-T cells compared to healthy control cells. RESULTS: We demonstrate that heptanoate corrects all aspects of the defective ER-mitochondrial signalling observed in A-T cells. Heptanoate enhances ER-mitochondrial contacts; increases the flow of calcium from the ER to the mitochondrion; restores normal mitochondrial function and mitophagy and increases the resistance of ATM-deficient cells and cells from A-T patients to metabolic stress-induced killing. The defect in mitochondrial function in ATM-deficient cells was accompanied by more reliance on aerobic glycolysis as shown by increased lactate dehydrogenase A (LDHA), accumulation of lactate, and reduced levels of both acetyl CoA and ATP which are all restored by heptanoate. CONCLUSIONS: We conclude that heptanoate corrects metabolic stress in A-T cells by restoring ER-mitochondria signalling and mitochondrial function and suggest that the parent compound, triheptanoin, has immense potential as a novel therapeutic agent for patients with A-T.

Premature ovarian ageing following heterozygous loss of Senataxin.

Premature loss of ovarian activity before 40 years of age is known as primary ovarian insufficiency (POI) and occurs in ∼1% of women. A more subtle decline in ovarian activity, known as premature ovarian ageing (POA), occurs in ∼10% of women. Despite the high prevalence of POA, very little is known regarding its genetic causation. Senataxin (SETX) is an RNA/DNA helicase involved in repair of oxidative stress-induced DNA damage. Homozygous mutation of SETX leads to the neurodegenerative disorder, ataxia oculomotor apraxia type 2 (AOA2). There have been reports of POI in AOA2 females suggesting a link between SETX and ovarian ageing. Here, we studied female mice lacking either one (Setx+/-) or both (Setx-/-) copies of SETX over a 12- to 14-month period. We find that DNA damage is increased in oocytes from 8-month-old Setx+/- and Setx-/- females compared with Setx+/+ oocytes leading to a marked reduction in all classes of ovarian follicles at least 4 months earlier than typically occurs in female mice. Furthermore, during a 12-month long mating trial, Setx+/- and Setx-/- females produced significantly fewer pups than Setx+/+ females from 7 months of age onwards. These data show that SETX is critical for preventing POA in mice, likely by preserving DNA integrity in oocytes. Intriguingly, heterozygous Setx loss causes an equally severe impact on ovarian ageing as homozygous Setx loss. Because heterozygous SETX disruption is less likely to produce systemic effects, SETX compromise could underpin some cases of insidious POA.

Vaccine-induced thrombosis and thrombocytopenia (VITT) in Ireland: A review of cases and current practices.

Since the beginning of the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS CoV-2) virus pandemic, several highly effective and safe vaccines have been produced at remarkable speed. Following global implementation of vaccination programmes, cases of thrombosis with thrombocytopenia following administration of adenoviral vector-based vaccines started being reported. In this review we discuss the known pathogenesis and epidemiology of so-called vaccine induced thrombocytopenia and thrombosis (VITT). We consider the available guidelines, diagnostic laboratory tests and management options for these patients. Finally, we discuss important unanswered questions and areas for future research in this novel pathoclinical entity.

Prolonged and large outbreak of invasive group A Streptococcus disease within a nursing home: repeated intrafacility transmission of a single strain.

OBJECTIVES: Multiple invasive group A Streptococcus (GAS) infections were reported to public health by a skilled nursing facility (facility A) in Illinois between May 2014 and August 2016. Cases continued despite interventions including antibiotic prophylaxis for all residents and staff. Two other geographically close facilities reported contemporaneous outbreaks of GAS. We investigated potential reasons for ongoing transmission. METHODS: We obtained epidemiologic data from chart review of cases and review of facility and public health records from previous investigations into the outbreak. Infection control practices at facility A were observed and evaluated. Whole genome sequencing followed by phylogenetic analysis was performed on available isolates from the three facilities. RESULTS: From 2014 to 2016, 19 invasive and 60 noninvasive GAS infections were identified at facility A occurring in three clusters. Infection control evaluations during clusters 2 and 3 identified hand hygiene compliance rates of 14% to 25%, appropriate personal protective equipment use in only 33% of observed instances, and deficient wound-care practices. GAS isolates from residents and staff of all three facilities were subtype emm89.0; on phylogenetic analysis, facility A isolates were monophyletic and distinct. CONCLUSIONS: Inadequate infection control and improper wound-care practices likely led to this 28-month-long outbreak of severe infections in a skilled nursing facility. Whole genome sequencing and phylogenetic analysis suggested that intrafacility transmission of a single highly transmissible GAS strain was responsible for the outbreak in facility A. Integration of genomic epidemiology tools with traditional epidemiology and infection control assessments was helpful in investigation of a facility-wide outbreak.

Acquired Factor Xiii Deficiency: An Uncommon But Easily Missed Cause Of Severe Bleeding

Factor XIII (FXIII) is a plasma clotting protein involved in clot stabilization. Severe FXIII deficiency may present with severe, even fatal bleeding. Critically however, routine coagulation assays may be normal and only specific FXIII assays will detect the abnormality. Herein we discuss a case report of a patient with acquired FXIII deficiency in order to highlight the clinical challenges associated with establishing the diagnosis and discuss the treatment approach. A 70-year-old man presented with a gluteal haematoma despite no preceding personal history of bleeding. Extensive initial haemostatic investigations were normal until a specific FXIII assay showed a marked reduction in FXIII levels. With directed treatment, bleeding episodes ceased and remission was achieved. Clinical awareness of FXIII deficiency is important, so appropriate testing can be implemented in patients with unexplained bleeding diatheses, particularly those in whom bleeding responds poorly to standard replacement therapy.

Invited review: Current production trends, farm structures, and economics of the dairy sheep and goat sectors.

Dairy small ruminants account for approximately 21% of all sheep and goats in the world, produce around 3.5% of the world's milk, and are mainly located in subtropical-temperate areas of Asia, Europe, and Africa. Dairy sheep are concentrated around the Mediterranean and Black Sea regions, where their dairy products are typical ingredients of the human diet. Dairy goats are concentrated in low-income, food-deficit countries of the Indian subcontinent, where their products are a key food source, but are also present in high-income, technologically developed countries. This review evaluates the status of the dairy sheep and goat sectors in the world, with special focus on the commercially and technically developed industries in France, Greece, Italy, and Spain (FGIS). Dairy small ruminants account for a minor part of the total agricultural output in France, Italy, and Spain (0.9 to 1.8%) and a larger part in Greece (8.8%). In FGIS, the dairy sheep industry is based on local breeds and crossbreeds raised under semi-intensive and intensive systems and is concentrated in a few regions in these countries. Average flock size varies from small to medium (140 to 333 ewes/farm), and milk yield from low to medium (85 to 216 L/ewe), showing substantial room for improvement. Most sheep milk is sold to industries and processed into traditional cheese types, many of which are Protected Denomination of Origin (PDO) cheeses for gourmet and export markets (e.g., Pecorino, Manchego, and Roquefort). By comparing break-even milk price among FGIS countries, we observed the following: (1) most Greek and French dairy sheep farms were unprofitable, with the exception of the intensive Chios farms of Greece; (2) milk price was aligned with cost of production in Italy; and (3) profitable farms coexisted with unprofitable farms in Spain. In FGIS, dairy goat production is based on local breeds raised under more extensive systems than sheep. Compared with sheep, average dairy goat herds are smaller (36 to 190 does/farm) but milk yield is greater (153 to 589 L/doe), showing room for improvement. Goat milk is mainly processed on-farm into dairy products for national markets, but some PDO goat milk cheeses (e.g., Murcia al Vino) are exported. Processed goat milk is sold for local human consumption or dehydrated for export. Mixed sheep-goat (e.g., Feta) and cow-sheep-goat milk cheeses are common in many countries. Strategies to improve the dairy sheep and goat sectors in these 4 countries are proposed and discussed.

Lenalidomide as a novel treatment for refractory acquired von Willebrand syndrome associated with monoclonal gammopathy.

UNLABELLED: Essentials Treatment options are limited for refractory bleeding in acquired von Willebrand Syndrome (AVWS). Lenalidomide therapy was studied in two patients with AVWS due to monoclonal gammopathy (MG). Lenalidomide increased von Willebrand factor (VWF), lowered VWF clearance and resolved bleeding. Lenalidomide is a potential treatment option for refractory bleeding in AVWS secondary to MG. SUMMARY: Background Acquired von Willebrand syndrome (AVWS) is associated with lymphoproliferative disorders, including monoclonal gammopathy (MG) of undetermined significance (MGUS) and multiple myeloma. Patients commonly present with significant bleeding complications that are difficult to manage, owing to a markedly reduced von Willebrand factor (VWF) half-life. Objectives To investigate the use of the immunomodulatory drug lenalidomide in two patients with severe refractory bleeding caused by AVWS associated with MGs. Results In both patients, lenalidomide treatment resulted in significant clinical improvement, and marked increases in plasma VWF antigen (VWF:Ag) and VWF ristocetin cofactor levels. This normalization in plasma VWF levels was sustained for > 2 years in both patients. Furthermore, in one patient, plasma VWF levels remain normal for at least 14 months following discontinuation of lenalidomide treatment. To investigate the molecular mechanisms underlying these observations, VWF propeptide (VWFpp)/VWF:Ag ratios were analyzed to assess VWF clearance. At enrolment, plasma VWFpp/VWF:Ag ratios were significantly elevated in both patients. Importantly, lenalidomide treatment resulted in normalization of VWFpp/VWF:Ag ratios in both patients. These novel data suggest that lenalidomide functions to attenuate enhanced VWF clearance in AVWS. Interestingly, in a patient with MGUS, lenalidomide treatment was associated with a significant increase in plasma VWF levels, despite no major change in paraprotein level. Conclusions Collectively, our findings suggest that lenalidomide constitutes a novel therapeutic option for the management of AVWS associated with MG. The biological mechanism(s) through which lenalidomide causes a sustained increase in plasma VWF levels in AVWS independently of paraprotein level requires further study, but is in part modulated through inhibition of enhanced VWF clearance.

Fibromyalgia beyond reductionism. Heart rhythm fractal analysis to assess autonomic nervous system resilience.

OBJECTIVES: The prevailing linear reductionist medical model seems unable to explain complex multisymptomatic illnesses such as fibromyalgia (FM) and similar maladies. Paradigms derived from the complexity theory may provide a coherent framework for these elusive illnesses. Along these lines is the proposal that FM represents a degradation of our main complex adaptive system (the autonomic nervous system, ANS), in a failed effort to adjust to a hostile environment. Healthy complex systems have fractal structures. Heart rate fractal-like variability reflects resilient ANS performance. Our aim was to measure the heart rate variability (HRV) fractal scaling index in FM patients and to correlate this index with clinical symptoms. METHOD: We studied 30 women with FM and 30 controls. All participants filled out questionnaires assessing the severity of FM. The HRV fractal scaling index was estimated during 24 h using detrended fluctuation analysis (DFA). RESULTS: The fractal scaling index alpha-1 was higher in FM patients than in controls (mean ± sd: 1.22 ± 0.10 vs. 1.16 ± 0.09; p = 0.031). There was a positive correlation between the fractal scaling index alpha-1 and the visual analogue scale (VAS) for depression (Spearman's ρ = 0.36, p = 0.04). CONCLUSIONS: The heart rate fractal exponent alpha-1 is altered in FM patients, suggesting a rigid ANS performance. This tangible non-linear finding supports the notion that FM may represent a degradation of our main complex adaptive system, namely the ANS.

von Willebrand factor arginine 1205 substitution results in accelerated macrophage-dependent clearance in vivo.

BACKGROUND: Enhanced von Willebrand factor (VWF) clearance is important in the etiology of type 1 and type 2 von Willebrand disease (VWD). More than 20 different VWF point mutations have already been reported in patients with enhanced clearance. These include the VWD-Vicenza variant, which is characterized by an Arg1205His substitution in the VWF D3 domain. Critically, however, the molecular mechanisms through which single amino acid substitutions in VWF result in enhanced clearance of this complex multimeric glycoprotein have not been defined. OBJECTIVES: In this study, we have investigated the biological basis underlying the enhanced clearance of the VWF-R1205H variant. METHODS: Using VWF(-/-) mice, in vivo clearance rates were determined for a series of full-length and truncated recombinant VWF variants. In addition, the role of macrophages in modulating enhanced VWD-Vicenza clearance was investigated using clodronate liposome administration. RESULTS: Our findings demonstrate that substitutions of R1205 with histidine, cysteine or serine all result in markedly reduced survival of full-length recombinant VWF. Importantly, D'A3 fragments containing these same R1205 substitutions also demonstrated significantly enhanced clearance. In contrast to the reduced in vivo survival observed with R1205H, clearance of R1204H was not enhanced. Recent studies have demonstrated that hepatic and splenic macrophages play key roles in regulating VWF clearance. Importantly, macrophage-depletion also served to markedly attenuate the enhanced clearance phenotypes associated with VWF-R1205H, VWF-R1205S and VWF-R1205C. CONCLUSIONS: Collectively, these novel findings demonstrate a specific and critical role for the R1205 residue in modulating macrophage-mediated clearance of VWF in vivo.

A late methanogen origin for molybdenum-dependent nitrogenase.

Mounting evidence indicates the presence of a near complete biological nitrogen cycle in redox-stratified oceans during the late Archean to early Proterozoic (c. 2.5-2.0 Ga). It has been suggested that the iron (Fe)- or vanadium (V)-dependent nitrogenase rather than molybdenum (Mo)-dependent form was responsible for dinitrogen fixation during this time because oceans were depleted in Mo and rich in Fe. We evaluated this hypothesis by examining the phylogenetic relationships of proteins that are required for the biosynthesis of the active site cofactor of Mo-nitrogenase in relation to structural proteins required for Fe-, V- and Mo-nitrogenase. The results are highly suggestive that among extant nitrogen-fixing organisms for which genomic information exists, Mo-nitrogenase is unlikely to have been associated with the Last Universal Common Ancestor. Rather, the origin of Mo-nitrogenase can be traced to an ancestor of the anaerobic and hydrogenotrophic methanogens with acquisition in the bacterial domain via lateral gene transfer involving an anaerobic member of the Firmicutes. A comparison of substitution rates estimated for proteins required for the biosynthesis of the nitrogenase active site cofactor and for a set of paralogous proteins required for the biosynthesis of bacteriochlorophyll suggests that Nif emerged from a nitrogenase-like ancestor approximately 1.5-2.2 Ga. An origin and ensuing proliferation of Mo-nitrogenase under anoxic conditions would likely have occurred in an environment where anaerobic methanogens and Firmicutes coexisted and where Mo was at least episodically available, such as in a redox-stratified Proterozoic ocean basin.