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Biofilms are structured and densely packed communities of microbial cells attached to surfaces. They are considered the natural growth state for a vast majority of microorganisms. The ability to form biofilms is an important virulence factor for most pathogens, including the opportunistic human fungal pathogen Candida albicans. C. albicans is one of the most prevalent fungal species of the human microbiota. However, C. albicans can also cause severe and life-threatening infections when host conditions permit. Conventional C. albicans biofilms are often resistant to antifungal agents and the host immune response and can act as reservoirs of infectious cells, maintaining persistent infections and seeding new infections in a host. The majority of C. albicans clinical isolates are heterozygous (a/α) at the mating type-like (MTL) locus, which defines Candida mating types, and can form robust biofilms when cultured in vitro. These “conventional” biofilms, formed by MTL-heterozygous (a/α) cells, have been the primary focus of C. albicans biofilm research to date. Recent work in the field has uncovered novel mechanisms through which biofilms are generated by C. albicans cells that are homozygous or hemizygous (a/a, a/Δ, α/α, or α/Δ) at the MTL-locus. In these studies, the addition of mating pheromone of a particular mating type can induce cells of the other mating type to form specialized “sexual” biofilms. Although sexual biofilms are generally less robust than conventional biofilms, they could serve as a protective niche to promote the parasexual life cycle of mating-competent cells, and thus could be an adaptation which may increase population diversity in dynamic environments. In this work, we investigate the molecular and genetic mechanisms that enable sexual biofilm formation in C. albicans to gain insight into this unique biological process and establish a basis for comparison between sexual and conventional biofilms. Chapter 1 of this dissertation provides a detailed review of sexual biofilm formation in C. albicans. Chapter 2 of this dissertation provides detailed analyses and transcriptomics of several clinically isolated strains growing under sexual biofilm inducing conditions as well as co-cultures of white and opaque cells of various mating types. This led to the identification of several downstream genes that are involved in sexual biofilm formation. Chapter 3 of this dissertation identifies several TFs that are involved in regulating sexual biofilms. We find that many regulators of conventional biofilm formation are similarly involved in sexual biofilm formation. Interestingly, some transcription factors appear to switch from positive regulators of one system to negative regulators of the other. In addition, differential gene expression and gene ontology analyses reveal the unique role of these transcription factors in regulating sexual biofilm formation. Finally, Chapter 4 provides a detailed description of the conclusions and future directions for where other researchers can take this project, particularly to build clinical relevance for the sexual biofilm system and to uncover the transcription network of sexual biofilms and elucidate the evolutionary history of sexual and conventional biofilms.

16 p.-6 fig.-2 tab., The sustainability of control programs for the Mediterranean fruit fly, Ceratitis capitata, for citrus crops in Spain has been threatened by the development of resistance to malathion and lambda-cyhalothrin in recent years. Spinosad is widely used without apparent loss of efficacy. However, a highly resistant strain, JW-100s, has been obtained after laboratory selection. Spinosad resistance in JW-100s has been associated with different mutant alleles of the α6 subunit of the nicotinic acetylcholine receptor (Ccα6) including an isoform-specific truncation allele, Ccα63aQ68*. Using the GAL4 > UAS system in Drosophila melanogaster to demonstrate expression of this truncated α6 subunit, in a dα6 loss-of-function genetic background, does not rescue susceptibility to spinosad, while the expression of Ccα6 wild-type isoforms does. We have also generated C. capitata isolines from JW-100s homozygous for: (1) the Ccα63aQ68*Δ3b-4 allele, which contains the mutation 3aQ68*, and (2) the Ccα63aQ68*–K352* allele, which contains the mutations 3aQ68* and K352*. Neither of these produce complete Ccα6 transcripts. The frequency of resistant alleles declined when in competition with individuals carrying the wild-type allele. Through extensive testing of both biological and behavioral fitness traits, we identified a reduced ability of Ccα63aQ68*Δ3b-4 males to detect the parapheromone and to mate with females carrying the Ccα63aQ68*–K352* allele in competition experiments. Thus, not only the potential for spontaneous resistant mutations to arise in Ccα6 but also their fitness costs must be considered when planning resistance management strategies for C. capitata., This work received financial support from CICYT(AGL2016-76516-R). The Spanish MINECO granted A. Guillem-Amat a predoc (BES-C-2014-068937) and a mobility (EEBB-I-16-11336) fellowships.

PURPOSE CD19-targeted chimeric antigen receptor T cells (CD19-CAR) and blinatumomab effectively induce remission in relapsed or refractory B-cell acute lymphoblastic leukemia (ALL) but are also associated with CD19 antigen modulation. There are limited data regarding the impact of prior blinatumomab exposure on subsequent CD19-CAR outcomes. PATIENTS AND METHODS We conducted a multicenter, retrospective review of children and young adults with relapsed or refractory ALL who received CD19-CAR between 2012 and 2019. Primary objectives addressed 6-month relapse-free survival (RFS) and event-free survival (EFS), stratified by blinatumomab use. Secondary objectives included comparison of longer-term survival outcomes, complete remission rates, CD19 modulation, and identification of factors associated with EFS. RESULTS Of 420 patients (median age, 12.7 years; interquartile range, 7.1-17.5) treated with commercial tisagenlecleucel or one of three investigational CD19-CAR constructs, 77 (18.3%) received prior blinatumomab. Blinatumomab-exposed patients more frequently harbored KMT2A rearrangements and underwent a prior stem-cell transplant than blinatumomab-naïve patients. Among patients evaluable for CD19-CAR response (n = 412), blinatumomab nonresponders had lower complete remission rates to CD19-CAR (20 of 31, 64.5%) than blinatumomab responders (39 of 42, 92.9%) or blinatumomab-naive patients (317 of 339, 93.5%), P < .0001. Following CD19-CAR, blinatumomab nonresponders had worse 6-month EFS (27.3%; 95% CI, 13.6 to 43.0) compared with blinatumomab responders (66.9%; 95% CI, 50.6 to 78.9; P < .0001) or blinatumomab-naïve patients (72.6%; 95% CI, 67.5 to 77; P < .0001) and worse RFS. High-disease burden independently associated with inferior EFS. CD19-dim or partial expression (preinfusion) was more frequently seen in blinatumomab-exposed patients (13.3% v 6.5%; P = .06) and associated with lower EFS and RFS. CONCLUSION With the largest series to date in pediatric CD19-CAR, and, to our knowledge, the first to study the impact of sequential CD19 targeting, we demonstrate that blinatumomab nonresponse and high-disease burden were independently associated with worse RFS and EFS, identifying important indicators of long-term outcomes following CD19-CAR.

The pathogenesis of Ebola virus disease (EVD) is still incomplete, although the non-human primate model has been studied for more than 4 decades. To further investigate EVD pathogenesis, a natural history study has been conducted using 27 Chinese-origin rhesus macaques. Of them, 24 macaques were exposed intramuscularly to Kikwit Ebola virus (EBOV) and euthanized at pre-determined timepoints or when end stage clinical disease criteria were met, while 3 other sham-exposed macaques were euthanized at the study day 0. This study demonstrates for the first time that Ebola virus causes uterine cervicitis, vaginitis, posthitis, and medullary adrenalitis. Not only is Ebola virus detected in the interstitial stromal cells of the genital tract, but it is also present in the epididymal and seminal vesicular tubular epithelial cells, ectocervical and vaginal squamous epithelial cells, and seminal fluid. Furthermore, as early as day 3 after exposure, EBOV replicative intermediate RNA was detected in Kupffer cells and hepatocytes. These findings in the nonhuman model provide additional insight into potential sexual transmission, possible disruption of sympathetic hormone production, and early virus replication sites in human EVD patients.

Hematuria or proteinuria may be just a normal transient finding in children, usually accompanying a non-specific viral infection, or it can be an indicator of a kidney or urinary tract disorder. In this chapter, the various causes and significance of discolored urine, hematuria and proteinuria in children are discussed. A pragmatic clinical approach to evaluate a child with isolated hematuria or isolated proteinuria is suggested to reduce unnecessary extensive workup in benign cases and yet not miss the cases with significant glomerular or urinary tract disease. Early detection and treatment of significant disorders would hopefully delay or prevent the onset of renal insufficiency in children.

Allergic asthma is a chronic inflammatory disorder of the airway associated with bronchial obstruction, airway hyper-reactivity (AHR), and mucus production. The epithelium may direct and propagate asthmatic-like responses. Central to this theory is the observation that viruses, air pollution, and allergens promote epithelial damage and trigger the generation of IL-25, IL-33, and TSLP via innate pathways such as TLRs and purinergic receptors. Similarly, engineered nanomaterials promote a Th2-associated pathophysiology. In this study, we tested the hypothesis that instillation of multi-walled carbon nanotubes (MWCNT) impair pulmonary function in C57Bl/6 mice due to the development of IL-33-dependent Th2-associated inflammation. MWCNT exposure resulted in elevated levels of IL-33 in the lavage fluid (likely originating from airway epithelial cells), enhanced AHR, eosinophil recruitment, and production of Th2-associated cytokines and chemokines. Moreover, these events were dependent on IL-13 signaling and the IL-33/ST2 axis, but independent of T and B cells. Finally, MWCNT exposure resulted in the recruitment of innate lymphoid cells. Collectively, our data suggest that MWCNT induce epithelial damage that results in release of IL-33, which in turn promotes innate lymphoid cell recruitment and the development of IL-13-dependent inflammatory response.

Objectives This report aims to review the management and outcomes of Brucella-associated mycotic aortic aneurysms. Methods This is a retrospective chart review at a tertiary-level healthcare system. IRB approval was waived per policy. Results We describe a case of Brucella aortitis acquired from habitual contact with wild hogs. Clinical presentation included lower back pain and elevated white blood cell count. Diagnosis was confirmed with imaging showing an infrarenal abdominal aortic aneurysm and serology revealing elevated Brucella antibodies titers. The patient was initially managed with endovascular aortic repair and combined oral and intravenous antibiotics therapy. He then underwent explanation and extra-anatomical bypass due to symptomatic periaortic infection and interval development of type I endoleak. The patient was asymptomatic after his final operation at 24 months of follow-up and remained on suppressive oral antibiotic therapy. Conclusions An aortic aneurysm secondary to Brucella is a rare entity. A detailed history of long-term exposure to animals may be a clue to obtain serologic testing. Operative debridement and re-establishing of reliable blood flow combined with long-term antibiotic suppression are the mainstay of treatment.

Diet-induced obesity has previously been shown to occur with the concomitant rise in the expression of proinflammatory cytokines and increases in collagen deposition. While it has been known that the regenerative process of skeletal muscle is altered in obese mice following an acute muscle injury, we sought to examine differences in the expression of various markers of extracellular matrix remodeling and repair. Our laboratory has previously reported an impaired inflammatory and protein synthetic signaling in these mice that may contribute negatively to the muscle regenerative process. To expand upon this previous investigation, tissues from these animals underwent further analysis to determine the extent of changes to the regenerative response within the extracellular matrix, including transcriptional changes in Collagen I, Collagen III, and Fibronectin. Here, we show that the expression of Collagen III:I is significantly increased at 3-days post-injury in obese injured animals compared to lean injured animals (p ​= ​0.0338), and by 28-days the obese injured animals exhibit a significantly lower Collagen III:I than their lean injured counterparts (p ​= ​0.0035). We demonstrate an impaired response to an acute muscle injury in obese mice when compared with lean counterparts. However, further studies are required to elucidate translational consequences of these changes, as well as to determine any causative mechanisms that may be driving this effect.

Zinc is an essential cofactor for bacterial metabolism, and many Enterobacteriaceae express the zinc transporters ZnuABC and ZupT to acquire this metal in the host. However, the probiotic bacterium Escherichia coli Nissle 1917 (or “Nissle”) exhibits appreciable growth in zinc-limited media even when these transporters are deleted. Here, we show that Nissle utilizes the siderophore yersiniabactin as a zincophore, enabling Nissle to grow in zinc-limited media, to tolerate calprotectin-mediated zinc sequestration, and to thrive in the inflamed gut. We also show that yersiniabactin’s affinity for iron or zinc changes in a pH-dependent manner, with increased relative zinc binding as the pH increases. Thus, our results indicate that siderophore metal affinity can be influenced by the local environment and reveal a mechanism of zinc acquisition available to commensal and pathogenic Enterobacteriaceae., Zinc is an essential cofactor for bacterial metabolism. Here, the authors show that the probiotic bacterium Escherichia coli Nissle 1917 utilizes the siderophore yersiniabactin as a zincophore, allowing the microbe to grow in zinc-limited media and to thrive in the inflamed gut.

Background: Abnormalities of mitochondrial fission and fusion, dynamic processes known to be essential for various aspects of mitochondrial function, have repeatedly been reported to be altered in Alzheimer’s disease (AD). Neurofibrillary tangles are known as a hallmark feature of AD and are commonly considered a likely cause of neurodegeneration in this devastating disease. Objective: To understand the pathological role of mitochondrial dynamics in the context of tauopathy. Methods: The widely used P301S transgenic mice of tauopathy (P301S mice) were crossed with transgenic TMFN mice with the forced expression of Mfn2 specifically in neurons to obtain double transgenic P301S/TMFN mice. Brain tissues from 11-month-old non-transgenic (NTG), TMFN, P301S, and P301S/TMFN mice were analyzed by electron microscopy, confocal microscopy, immunoblot, histological staining, and immunostaining for mitochondria, tau pathology, and tau pathology-induced neurodegeneration and gliosis. The cognitive function was assessed by the Barnes maze. Results: P301S mice exhibited mitochondrial fragmentation and a consistent decrease in Mfn2 compared to age-matched NTG mice. When P301S mice were crossed with TMFN mice (P301S/TMFN mice), neuronal loss, as well as mitochondria fragmentation were significantly attenuated. Greatly alleviated tau hyperphosphorylation, filamentous aggregates, and thioflavin-S positive tangles were also noted in P301S/TMFN mice. Furthermore, P301S/TMFN mice showed marked suppression of neuroinflammation and improved cognitive performance in contrast to P301S mice. Conclusion: These in vivo findings suggest that promoted mitochondrial fusion suppresses toxic tau accumulation and associated neurodegeneration, which may protect against the progression of AD and related tauopathies.

Marine ecosystems on continental shelves face multiple challenges due to anthropogenic disturbances, many of which can change the seawater stoichiometry (C:N:P) and consequently elemental ratios of phytoplankton. This change in food quality may not be tolerated by all grazers and predators. Gelatinous and soft-bodied zooplankton (GZ) might be more resilient to such changes. We sampled GZ species in neritic and oceanic waters of the Northeast Pacific off British Columbia, Canada, determined their phosphorus (P) content and elemental ratios (C/P, N/P), and analysed intraspecific variability associated with size and ontogeny. P content was measured for twelve GZ taxa. P % DW (dry weight) decreased with size for the hydrozoan Aequorea sp., scyphozoans Aurelia labiata, Cyanea capillata, and the thaliacean Salpa aspera, and differed significantly for two development stages of the salp S. aspera. While C/P and N/P were mostly size and stage independent, they were highly variable. C/P values of GZ were generally higher than values of crustacean zooplankton, indicating that GZ represent poor-quality prey for non-GZ predators, and that GZ may have a higher resilience towards P-limited (low quality) prey. Changing ocean conditions and nutrient stoichiometry of prey may favour GZ, although large variability in P dynamics among GZ taxa and uncertainty about future ocean stoichiometry changes make generalisations difficult.

Several large-scale Illumina whole-genome sequencing (WGS) and whole-exome sequencing (WES) projects have emerged recently that have provided exceptional opportunities to discover mobile element insertions (MEIs) and study the impact of these MEIs on human genomes. However, these projects also have presented major challenges with respect to the scalability and computational costs associated with performing MEI discovery on tens or even hundreds of thousands of samples. To meet these challenges, we have developed a more efficient and scalable version of our mobile element locator tool (MELT) called CloudMELT. We then used MELT and CloudMELT to perform MEI discovery in 57,919 human genomes and exomes, leading to the discovery of 104,350 nonredundant MEIs. We leveraged this collection (1) to examine potentially active L1 source elements that drive the mobilization of new Alu, L1, and SVA MEIs in humans; (2) to examine the population distributions and subfamilies of these MEIs; and (3) to examine the mutagenesis of GENCODE genes, ENCODE-annotated features, and disease genes by these MEIs. Our study provides new insights on the L1 source elements that drive MEI mutagenesis and brings forth a better understanding of how this mutagenesis impacts human genomes.

The state of somatic energy stores in metazoans is communicated to the brain, which regulates key aspects of behaviour, growth, nutrient partitioning and development1. The central melanocortin system acts through melanocortin 4 receptor (MC4R) to control appetite, food intake and energy expenditure2. Here we present evidence that MC3R regulates the timing of sexual maturation, the rate of linear growth and the accrual of lean mass, which are all energy-sensitive processes. We found that humans who carry loss-of-function mutations in MC3R, including a rare homozygote individual, have a later onset of puberty. Consistent with previous findings in mice, they also had reduced linear growth, lean mass and circulating levels of IGF1. Mice lacking Mc3r had delayed sexual maturation and an insensitivity of reproductive cycle length to nutritional perturbation. The expression of Mc3r is enriched in hypothalamic neurons that control reproduction and growth, and expression increases during postnatal development in a manner that is consistent with a role in the regulation of sexual maturation. These findings suggest a bifurcating model of nutrient sensing by the central melanocortin pathway with signalling through MC4R controlling the acquisition and retention of calories, whereas signalling through MC3R primarily regulates the disposition of calories into growth, lean mass and the timing of sexual maturation. MC3R deficiency is associated with a delay in the onset of puberty, and a reduction in growth and lean mass.

Demodex blepharitis is a common ophthalmologic disease that is often overlooked in the workup of blepharitis. Demodex infestation occurs in both symptomatic and asymptomatic individuals at similar rates; consequently, its role in the development of blepharitis has not been well elucidated. Two species have been confirmed to inhabit the human eyelid- Demodex folliculorum and Demodex brevis. These species differ in their preferred location of infestation, with D. folliculorum occupying the base of the eyelash and D. brevis inhabiting the meibomian glands, contributing to anterior and posterior Demodex blepharitis, respectively. A clinical index of suspicion must be developed from the history when blepharitis, conjunctivitis, or keratitis in adults and blepharoconjunctivitis or chalazia in children are resistant to treatment. The diagnosis can be strongly suspected by the presence of cylindrical dandruff at the base of the eyelash and confirmed by light microscopy of an epilated lash or by in vivo confocal microscopy. No cure is currently available. Management most frequently includes topical tea tree oil and its active ingredient terpinen-4-ol, both of which have produced good clinical outcomes. Topical tea tree oil is typically applied by a professional due to risk of toxicity. Several second-line treatment options have been studied, including ivermectin, metronidazole, selenium sulfide, microblepharoexfoliation, and lid hygiene. Novel, targeted therapies such as TP-03 (Tarsus Pharmaceuticals) are also currently being investigated in phase 2b/3 clinical trials. The purpose of this review purpose is to characterize Demodex blepharitis in detail, including its historical perspective and various classifications, and describe the latest diagnostic and management strategies.

There is an urgent need for a malaria vaccine that can prevent severe disease in young children and adults. Despite earlier work showing an immunological mechanism for preventing infection and reducing disease severity, there is currently no reliable vaccine that can provide durable protection. In part, this may reflect a limited number of ways that the host can respond to the NANP repeat sequences of circumsporozoite protein (CSP) in the parasite. In addition, it may reflect antigenic escape by the parasite from protective antibodies. To be successful, a vaccine must protect against repeated exposure to infected mosquitoes in endemic areas. We have created a series of live viral vectors based on the rubella vaccine strain that express multiple tandem repeats of NANP, and we demonstrate immunogenicity in a rhesus macaque model. We tested the vectors in a sequential immunization strategy. In the first step, the animals were primed with CSP-DNA vaccine and boosted with rubella/CSP vectors. In the second step, we gave rubella/CSP vectors again, followed by recombinant CSP protein. Following the second step, antibody titers were comparable to adult exposure to malaria in an endemic area. The antibodies were specific for native CSP protein on sporozoites, and they persisted for at least 1½ years in two out of three macaques. Given the safety profile of rubella vaccine in children, these vectors could be most useful in protecting young children, who are at greatest risk of severe malarial disease.

Peroxynitrite is a reactive intermediate formed in vivo through uncatalysed reaction of superoxide and nitric oxide radicals. Despite significant interest in detecting peroxynitrite in vivo and understanding its production, little attention has been given to the evolutionary origins of peroxynitrite signalling. Herein we focus on two enzymes that are key to the biosynthesis of superoxide and nitric oxide, NADPH oxidase 5 (NOX5) and endothelial nitric oxide synthase (eNOS), respectively. Multiple sequence alignments of both enzymes including homologues from all domains of life, coupled with a phylogenetic analysis of NOX5, suggest eNOS and NOX5 are present in animals as the result of horizontal gene transfer from ancestral cyanobacteria to ancestral eukaryotes. Therefore, biochemical studies from other laboratories on a NOX5 homologue in Cylindrospermum stagnale and an eNOS homologue in Synechococcus sp. PCC 7335 are likely to be of relevance to human NOX5 and eNOS and to the production of superoxide, nitric oxide and peroxynitrite in humans.

Coral reef fish perform numerous important functional roles on coral reefs. Of these, carbonate sediment production, as a by‐product of parrotfish feeding, is especially important for contributing to reef framework construction and reef‐associated landform development. However, only limited data exist on: (i) how production rates vary among reef habitats as a function of parrotfish assemblages, (ii) the relative importance of sediment produced from eroded, reworked, and endogenous sources, or (iii) the size fractions of sediment generated by different parrotfish species and size classes. These parameters influence not only overall reef‐derived sediment supply, but also influence the transport potential and depositional fate of this sedimentary material. Here, we show that parrotfish sediment production varies significantly between reef‐platform habitats on an atoll‐margin Maldivian reef. Highest rates of production (over 0.8 kg m−2 year−1) were calculated in three of the eight platform habitats; a rubble‐dominated zone, an Acropora spp. dominated zone, and a patch reef zone. Habitat spatial extent and differences in associated parrotfish assemblages strongly influenced the total quantities of sediment generated within each habitat. Nearly half of total parrotfish sediment production occurred in the rubble habitat, which comprised only 8% of the total platform area. Over 90% of this sedimentary material originated from eroded reef framework as opposed to being reworked existing or endogenously produced sediment, and comprised predominantly coral sands (predominantly 125–1000 µm in diameter). This is comparable to the dominant sand types and size fractions found on Maldivian reef islands. By contrast, nearly half of the sediment egested by parrotfish in the Acropora spp. dominated and patch reef habitats resulted from reworked existing sediments. These differences between habitats are a result of the different parrotfish assemblages supported. Endogenous carbonate production was found to be insignificant compared to the quantity of eroded and reworked material. Our findings have important implications for identifying key habitats and species which act as major sources of sediment for reef‐island systems., Our findings show that parrotfish sediment production varies between reef habitat types as a function of the species assemblages and fish sizes supported. The origins of this material (i.e., bioerosion of reef substrate, sediment reworking, or endogenous production) also varies between habitat types because of the species and sizes of parrotfish present. In addition, we show that all species studied produce sediment size fractions suitable for shoreline maintenance.

Rationale: Fetal growth and survival depend critically on proper development and integrity of the vascular system. Fli1 (Friend leukemia integration 1), a member of the Ets family of transcription factors, plays critical roles in vascular morphogenesis and homeostasis at mid-gestation, the developmental stage at which expression of its upstream regulator, Etv2, ceases. However, molecular mechanisms of Fli1 action in vascular morphogenesis remain incompletely understood. Objective: To dissect molecular mechanisms of vascular morphogenesis governed by Fli1. Methods and Results: Utilizing Fli1 promoter-driven lineage-specific LacZ expression, Fli1 loss-of-function strategies, and a series of molecular techniques, we demonstrate that Fli1 expression in multipotent myogenic progenitor cells (MPCs) occurs independent of Etv2, and loss of Fli1 expression results in a significant increase in LacZ + cells in mesoderm within somites and limb buds, leading to reciprocal regulation of the expression of several key endothelial and myogenic genes and vascular abnormalities. Conversely, embryos harboring conditional Fli1 gain-of-function in MPCs manifested aberrant vasculogenesis with lack of myogenesis. Mechanistically, elevated Fli1 activity in myoblasts and in adult MPCs (also called satellite cells) of X-linked muscular dystrophic mdx mice markedly induced endothelial, but attenuated myogenic, gene expression and differentiation. Importantly, ectopic expression of Myf5 or MyoD, two key myogenic regulators, in Fli1-expressing myoblasts restored their differentiation potential, indicating that levels of Fli1 and myogenic regulators in MPCs inversely regulate their endothelial versus myogenic potential. Conclusions: Fli1 governs vascular morphogenesis by regulating endothelial potential by inversely regulating endothelial versus myogenic programs in MPCs. Our data uncover an important and previously unrecognized mechanism of vascular morphogenesis governed by Fli1 and highlight the physiological significance of the fine tuning of Fli1 activity in multipotent progenitors for proper vascular and muscle morphogenesis during development and disease.

Background: Antenatal care for pregnant and postnatal mothers is the main entry gate for maternal Tetanus Toxoid (TT) vaccination. In developing countries including Tanzania, pregnant and postnatal mothers especially in rural communities have been experiencing barriers to maternal vaccination. The study aimed to explore perspectives of pregnant and postnatal mothers on accessing TT vaccine in Hai District Kilimanjaro Region Tanzania. Methods: A descriptive qualitative study was conducted during baseline assessment from October 2020 to May 2021, in 10 remote villages. Pregnant and postnatal mothers attending antenatal and postnatal services were selected purposively based on their availability and convenience, A semi-structured interview guide relevant to the study objectives was used to guide the interviews. Focus Group Discussion (FGD) and In-depth interviews (IDIs) were conducted; and thematic analysis was used to obtain themes and subthemes. Results: A total of 15 mothers aged 18 to 40 years participated in the study. Most of the mothers were Christians, married with 1 to 5 children. Majority had completed primary level education engaged in small-scale farming. Main themes were extracted namely: Unreliable vaccine information, Healthcare workers attitude, Overworked with household activities, adherence of appointment dates for vaccination after delivery and role of Traditional medicine during pregnancy. Conclusion: The role of healthcare providers are critical component for the appropriate transfer of vaccine information to the mothers and their spouses during clinic visits, to eliminate vaccine misconceptions hence improving overall tetanus vaccine uptake. Quality of information from reliable sources could be an important factor to upgrade the awareness of pregnant mothers on maternal vaccination.

Global demand for seaweed is increasing, resulting in the rapid growth of the seaweed industry. To sustain and enhance the industry (seaweed growers and processors), it is essential to establish and optimize post-harvest food processing practices to extend the shelf life of the fresh, edible product. Blanching and freezing were applied in this study to offer longer-term storage to enable the distribution of sugar kelp (Saccharina latissima), farm-raised in Maine, USA. The objectives were to evaluate blanching, specifically the impacts of blanching method (direct immersion or vacuum packaged), temperature (80 or 100 °C) and time (5 or 30 s), kelp form (whole blade or shredded), and the effects of frozen storage on food product qualities including color, texture, drip loss, mineral content, total phenolic content, and the native microflora of sugar kelp after thawing. The fresh brown kelp developed a vivid green color immediately after blanching, especially with higher blanching temperature and longer blanching time. Drip loss was higher in blades than in shredded slaw, and vacuum packaged blanching produced higher moisture content, phenolic compounds, and antioxidant activity. Results substantiate the importance of the blanching method, temperature, and time on frozen kelp quality. Blanching prior to freezing resulted in relatively higher product quality than unblanched frozen kelp despite some losses in hardness and color. Frozen storage had minimal to no impact on texture, aerobic plate count, and psychrotrophic microorganisms. Higher temperature, longer time, and vacuum packaged blanching is suggested to extend the marketable shelf life of sugar kelp to at least 6 months at − 20 °C.

Despite the prevalence of high-throughput sequencing in phylogenetics, many relationships remain difficult to resolve because of conflicting signal among genomic regions. Selection of different types of molecular markers from different genomic regions is required to overcome these challenges. For evolutionary studies in frogs, we introduce the publicly available FrogCap suite of genomic resources, which is a large collection of ~15,000 markers that unifies previous genetic sequencing efforts. FrogCap is designed to be modular, such that subsets of markers and SNPs can be selected based on the desired phylogenetic scale. FrogCap uses a variety of marker types that include exons and introns, ultraconserved elements, and previously sequenced Sanger markers, which span up to 10,000 bp in alignment lengths; in addition, we demonstrate potential for SNP-based analyses. We tested FrogCap using 121 samples distributed across five phylogenetic scales, comparing probes designed using a consensus- or exemplar genome-based approach. Using the consensus design is more resilient to issues with sensitivity, specificity, and missing data than picking an exemplar genome sequence. We also tested the impact of different bait kit sizes (20,020 vs. 40,040) on depth of coverage and found triple the depth for the 20,020 bait kit. We observed sequence capture success (i.e., missing data, sequenced markers/bases, marker length, and informative sites) across phylogenetic scales. The incorporation of different marker types is effective for deep phylogenetic relationships and shallow population genetics studies. Having demonstrated FrogCap's utility and modularity, we conclude that these new resources are efficacious for high-throughput sequencing projects across variable timescales.

Establishment and maintenance of pregnancy in the pig is a complex process that relies on conceptus regulation of the maternal proinflammatory response to endometrial attachment. Following elongation, pig conceptuses secrete interferon gamma (IFNG) during attachment to the endometrial luminal epithelium. The objective here was to determine if conceptus production of IFNG is important for early development and establishment of pregnancy. CRISPR/Cas9 gene editing and somatic cell nuclear transfer technologies were used to create an IFNG loss-of-function study in pigs. Wild-type (IFNG+/+) and null (IFNG−/−) fibroblast cells were used to create embryos through somatic cell nuclear transfer. IFNG expression was not detected in IFNG−/− conceptuses on either day 15 or day 17 of pregnancy. Ablation of conceptus IFNG production resulted in the reduction of stromal CD3+ and mast cells, which localized to the site of conceptus attachment on day 15. The uteri of recipients with IFNG−/− conceptuses were inflamed, hyperemic and there was an abundance of erythrocytes in the uterine lumen associated with the degenerating conceptuses. The endometrial stromal extracellular matrix was altered in the IFNG−/− embryo pregnancies and there was an increased endometrial mRNA levels for collagen XVII (COL17A1), matrilin 1 (MATN1), secreted phosphoprotein 1 (SPP1), and cysteine-rich secretory protein 3 (CRISP3), which are involved with repair and remodeling of the extracellular matrix. These results indicate conceptus IFNG production is essential in modulating the endometrial proinflammatory response for conceptus attachment and survival in pigs.

Jojoba (Simmondsia chinensis (Link) Schneider) holds high industrial value and an extended cultivation trend. Despite its increased importance, there is a lack of fundamental information about its metabolic reserves and development. Our objective was to characterise metabolite allocation and fluctuations in the carbohydrate and nutrient balance of jojoba plants, as affected by fruit load and the plant’s annual cycle. Metabolite profiles were performed for each organ. Soluble carbohydrates (SC) and starch concentrations were surveyed in underground and aboveground organs of high-yield and fruit-removed plants. Simultaneously, nitrogen, potassium and phosphorus were determined in the leaves to evaluate the plant’s nutritional status. We found that sucrose and pinitol were the most abundant sugars in all jojoba organs. Each sugar had a ‘preferred’ organ: glucose was accumulated mainly in the leaves, sucrose and pinitol in woody branches, and fructose in the trunk wood. We found that fruit load significantly influenced the carbohydrate levels in green branches, trunk wood and thin roots. The phenological stage strongly affected the SC–starch balance. Among the examined minerals, only the leaf potassium level was significantly influenced by fruit load. We conclude that jojoba’s nutrient and carbohydrate balance is affected by fruit load and the phenological stage, and describe the organ-specific metabolic reserves.

Distributions of freshwater fish species in Labrador are poorly documented as the region is remote and sparsely inhabited. Here, we update distributions of four species native to the Labrador Peninsula based on data collected over 10 years: Burbot (Lota lota), Round Whitefish (Prosopium cylindraceum), Lake Trout (Salvelinus namaycush), and Slimy Sculpin (Cottus cognatus). In northern Labrador, our findings extend their ranges inland and northwest of their formerly reported distributions. Their presence in previously unknown locations indicates an alternative post-glacial colonization pathway to one previously proposed that suggested an isolated pocket of Lake Trout in a northern coastal area colonized through marine invasion. Instead, we suggest that overland colonization occurred when glacial Lake Naskaupi withdrew across Quebec into several northern drainages. In southern Labrador, we found Lake Trout and Round Whitefish to the southeast of their previously reported ranges. The discovery of an isolated population of Lake Trout in a remote location of southeast Labrador implies that they may have existed in the area historically (6000 years ago), but have undergone a range contraction with a warming climate. In addition, 22 new locations are documented for Lake Trout within their established range.

Viruses and viroids prevalent in a population of 42 wild grapevines (i.e., free-living, uncultivated grapevines; Vitis spp.) were compared with those in a population of 85 cultivated grapevines collected in Tennessee, United States by RNA sequencing analysis of pools of ribosomal RNA-depleted total RNA. The sequences of 10 viruses (grapevine fleck virus, grapevine leafroll-associated virus 2, grapevine rupestris stem pitting-associated virus, grapevine Syrah virus 1, grapevine vein-clearing virus, grapevine virus B, grapevine virus E, tobacco ringspot virus, tomato ringspot virus, and a novel nano-like virus) and two viroids (hop stunt viroid and grapevine yellow speckle viroid 1) were detected in both grapevine populations. Sequences of four viruses (grapevine associated tymo-like virus, grapevine leafroll-associated virus 3, grapevine red blotch virus, and grapevine virus H) were identified only from cultivated grapevines. High, moderate, and low numbers of sequence reads were identified only from wild grapevines for a novel caulimovirus, an enamovirus, and alfalfa mosaic virus, respectively. The presence of most virus sequences and both viroids was verified independently in the original samples by reverse-transcription PCR followed by Sanger sequencing. Comparison of viral sequences shared by both populations showed that cultivated and wild grapevines harbored distinct sequence variants, which suggests that there was limited virus movement between the two populations. Collectively, this study represents the first unbiased survey of viruses and viroids in both cultivated and wild grapevines within a defined geographic region.

To provide protection against viral infection and limit the uptake of mobile genetic elements, bacteria and archaea have evolved many diverse defence systems. The discovery and application of CRISPR-Cas adaptive immune systems has spurred recent interest in the identification and classification of new types of defence systems. Many new defence systems have recently been reported but there is a lack of accessible tools available to identify homologs of these systems in different genomes. Here, we report the Prokaryotic Antiviral Defence LOCator (PADLOC), a flexible and scalable open-source tool for defence system identification. With PADLOC, defence system genes are identified using HMM-based homologue searches, followed by validation of system completeness using gene presence/absence and synteny criteria specified by customisable system classifications. We show that PADLOC identifies defence systems with high accuracy and sensitivity. Our modular approach to organising the HMMs and system classifications allows additional defence systems to be easily integrated into the PADLOC database. To demonstrate application of PADLOC to biological questions, we used PADLOC to identify six new subtypes of known defence systems and a putative novel defence system comprised of a helicase, methylase and ATPase. PADLOC is available as a standalone package (https://github.com/padlocbio/padloc) and as a webserver (https://padloc.otago.ac.nz).