Malaria immunity in man and mosquito: insights into unsolved mysteries of a deadly infectious disease.

Malaria is a mosquito-borne disease caused by parasites of the obligate intracellular Apicomplexa phylum the most deadly of which, Plasmodium falciparum, prevails in Africa. Malaria imposes a huge health burden on the world's most vulnerable populations, claiming the lives of nearly one million children and pregnant women each year. Although there is keen interest in eradicating malaria, we do not yet have the necessary tools to meet this challenge, including an effective malaria vaccine and adequate vector control strategies. Here we review what is known about the mechanisms at play in immune resistance to malaria in both the human and mosquito hosts at each step in the parasite's complex life cycle with a view toward developing the tools that will contribute to the prevention of disease and death and, ultimately, to the goal of malaria eradication. In so doing, we hope to inspire immunologists to participate in defeating this devastating disease.

A high internal heat flux and large core in a warm Neptune exoplanet.

Interactions between exoplanetary atmospheres and internal properties have long been proposed to be drivers of the inflation mechanisms of gaseous planets and apparent atmospheric chemical disequilibrium conditions1. However, transmission spectra of exoplanets have been limited in their ability to observationally confirm these theories owing to the limited wavelength coverage of the Hubble Space Telescope (HST) and inferences of single molecules, mostly H2O (ref. 2). In this work, we present the panchromatic transmission spectrum of the approximately 750 K, low-density, Neptune-sized exoplanet WASP-107b using a combination of HST Wide Field Camera 3 (WFC3) and JWST Near-Infrared Camera (NIRCam) and Mid-Infrared Instrument (MIRI). From this spectrum, we detect spectroscopic features resulting from H2O (21σ), CH4 (5σ), CO (7σ), CO2 (29σ), SO2 (9σ) and NH3 (6σ). The presence of these molecules enables constraints on the atmospheric metal enrichment (M/H is 10-18× solar3), vertical mixing strength (log10Kzz = 8.4-9.0 cm2 s-1) and internal temperature (>345 K). The high internal temperature is suggestive of tidally driven inflation4 acting on a Neptune-like internal structure, which can naturally explain the large radius and low density of the planet. These findings suggest that eccentricity-driven tidal heating is a critical process governing atmospheric chemistry and interior-structure inferences for most of the cool (<1,000 K) super-Earth-to-Saturn-mass exoplanet population.

Methane throughout the atmosphere of the warm exoplanet WASP-80b.

The abundances of main carbon- and oxygen-bearing gases in the atmospheres of giant exoplanets provide insights into atmospheric chemistry and planet formation processes1,2. Thermochemistry suggests that methane (CH4) should be the dominant carbon-bearing species below about 1,000 K over a range of plausible atmospheric compositions3; this is the case for the solar system planets4 and has been confirmed in the atmospheres of brown dwarfs and self-luminous, directly imaged exoplanets5. However, CH4 has not yet been definitively detected with space-based spectroscopy in the atmosphere of a transiting exoplanet6-11, but a few detections have been made with ground-based, high-resolution transit spectroscopy12,13 including a tentative detection for WASP-80b (ref. 14). Here we report transmission and emission spectra spanning 2.4-4.0 µm of the 825 K warm Jupiter WASP-80b taken with the NIRCam instrument of the JWST, both of which show strong evidence of CH4 at greater than 6σ significance. The derived CH4 abundances from both viewing geometries are consistent with each other and with solar to sub-solar C/O and around five times solar metallicity, which is consistent with theoretical predictions15-17.

A broadband thermal emission spectrum of the ultra-hot Jupiter WASP-18b.

Close-in giant exoplanets with temperatures greater than 2,000 K ('ultra-hot Jupiters') have been the subject of extensive efforts to determine their atmospheric properties using thermal emission measurements from the Hubble Space Telescope (HST) and Spitzer Space Telescope1-3. However, previous studies have yielded inconsistent results because the small sizes of the spectral features and the limited information content of the data resulted in high sensitivity to the varying assumptions made in the treatment of instrument systematics and the atmospheric retrieval analysis3-12. Here we present a dayside thermal emission spectrum of the ultra-hot Jupiter WASP-18b obtained with the NIRISS13 instrument on the JWST. The data span 0.85 to 2.85 µm in wavelength at an average resolving power of 400 and exhibit minimal systematics. The spectrum shows three water emission features (at >6σ confidence) and evidence for optical opacity, possibly attributable to H-, TiO and VO (combined significance of 3.8σ). Models that fit the data require a thermal inversion, molecular dissociation as predicted by chemical equilibrium, a solar heavy-element abundance ('metallicity', [Formula: see text] times solar) and a carbon-to-oxygen (C/O) ratio less than unity. The data also yield a dayside brightness temperature map, which shows a peak in temperature near the substellar point that decreases steeply and symmetrically with longitude towards the terminators.

Intraspecific variation in crayfish behavioral traits affects leaf litter breakdown in streams.

Although intraspecific trait variation is increasingly recognized as affecting ecosystem processes, few studies have examined the ecological significance of among-population variation in behavioral traits in natural ecosystems. In freshwater habitats, crayfish are consumers that can influence ecosystem structure (e.g., macroinvertebrate communities) and function (e.g., leaf litter breakdown). To test whether crayfish behavioral traits (activity, boldness, and foraging voracity) are major contributors of leaf litter breakdown rates in the field, we collected rusty crayfish (Faxonius rusticus) from eight streams across the midwestern USA and measured behaviors using laboratory assays. At the same streams, we measured breakdown rates of leaf packs that were accessible or inaccessible to crayfish. Our results provide evidence that among-population variation in crayfish boldness and foraging voracity was a strong predictor of leaf litter breakdown rates, even after accounting for commonly appreciated environmental drivers (water temperature and human land use). Our results suggest that less bold rusty populations (i.e., emerged from shelter more slowly) had greater direct impacts on leaf litter breakdown than bold populations (P = 0.001, r2 = 0.85), potentially because leaf packs can be both a shelter and food resource to crayfish. Additionally, we found that foraging voracity was negatively related to breakdown rates in leaf packs that were inaccessible to crayfish (P = 0.025, r2 = 0.60), potentially due to a trophic cascade from crayfish preying on other invertebrates that consume leaf litter. Overall, our results add to the growing evidence that trait variation in animals may be important for understanding freshwater ecosystem functioning.

Developmental reprogramming of myometrial stem cells by endocrine disruptor linking to risk of uterine fibroids.

BACKGROUND: The stage, when tissues and organs are growing, is very vulnerable to environmental influences, but it's not clear how exposure during this time causes changes to the epigenome and increases the risk of hormone-related illnesses like uterine fibroids (UFs). METHODS: Developmental reprogramming of myometrial stem cells (MMSCs), the putative origin from which UFs originate, was investigated in vitro and in the Eker rat model by RNA-seq, ChIP-seq, RRBS, gain/loss of function analysis, and luciferase activity assays. RESULTS: When exposed to the endocrine-disrupting chemical (EDC) diethylstilbestrol during Eker rat development, MMSCs undergo a reprogramming of their estrogen-responsive transcriptome. The reprogrammed genes in MMSCs are known as estrogen-responsive genes (ERGs) and are activated by mixed lineage leukemia protein-1 (MLL1) and DNA hypo-methylation mechanisms. Additionally, we observed a notable elevation in the expression of ERGs in MMSCs from Eker rats exposed to natural steroids after developmental exposure to EDC, thereby augmenting estrogen activity. CONCLUSION: Our studies identify epigenetic mechanisms of MLL1/DNA hypo-methylation-mediated MMSC reprogramming. EDC exposure epigenetically targets MMSCs and leads to persistent changes in the expression of a subset of ERGs, imparting a hormonal imprint on the ERGs, resulting in a "hyper-estrogenic" phenotype, and increasing the hormone-dependent risk of UFs.

Research-Practice Partnership to Develop and Implement Routine Mental Health Symptom Tracking Tool Among Older Adults During COVID-19.

OBJECTIVE: Older adults are disproportionally impacted by the COVID-19 pandemic, causing a mental health crisis in late life, due to physical restrictions (e.g., quarantine), limited access to services, and lower literacy and access to technology. Despite established benefits, systematic screening of mental health needs of older adults in community and routine care settings is limited and presents multiple challenges. Cross-disciplinary collaborations are essential for identification and evaluation of mental health needs and service delivery. METHODS: Using a research-practice partnership model, we developed and implemented a routine mental health needs identification and tracking tool at a community-based social services organization. Repeated screenings were conducted remotely over 5 months and included depression, anxiety, perceived loneliness, social support, and related domains such as sleep quality, resilience, and trauma symptoms linked to COVID-19. We examined symptomatic distress levels and associations between different domains of functioning. RESULTS: Our project describes the process of establishing a research-practice partnership during the COVID-19 pandemic. We collected 292 screenings from 124 individuals; clients were mildly to moderately depressed and anxious, reporting large amounts of time alone and moderate levels of loneliness. Those reporting higher depressive symptoms reported higher anxiety symptoms, poorer sleep quality, lower quality of life, lower capacity to adapt to challenging situations, and greater trauma symptoms due to COVID-19. CONCLUSION: Our routine screening tool can serve as a blueprint for case management agencies and senior centers nationwide, beyond the pressing mental health crisis due to COVID-19, to continue identifying needs as they emerge in the community.

Toxic elements in arctic and sub-arctic brown bears: Blood concentrations of As, Cd, Hg and Pb in relation to diet, age, and human footprint.

Contamination with arsenic (As), cadmium (Cd), mercury (Hg) and lead (Pb) is a global concern impairing resilience of organisms and ecosystems. Proximity to emission sources increases exposure risk but remoteness does not alleviate it. These toxic elements are transported in atmospheric and oceanic pathways and accumulate in organisms. Mercury accumulates in higher trophic levels. Brown bears (Ursus arctos), which often live in remote areas, are long-lived omnivores, feeding on salmon (Oncorhynchus spp.) and berries (Vaccinium spp.), resources also consumed by humans. We measured blood concentrations of As, Cd, Hg and Pb in bears (n = 72) four years and older in Scandinavia and three national parks in Alaska, USA (Lake Clark, Katmai and Gates of the Arctic) using high-resolution, inductively-coupled plasma sector field mass spectrometry. Age and sex of the bears, as well as the typical population level diet was associated with blood element concentrations using generalized linear regression models. Alaskan bears consuming salmon had higher Hg blood concentrations compared to Scandinavian bears feeding on berries, ants (Formica spp.) and moose (Alces). Cadmium and Pb blood concentrations were higher in Scandinavian bears than in Alaskan bears. Bears using marine food sources, in addition to salmon in Katmai, had higher As blood concentrations than bears in Scandinavia. Blood concentrations of Cd and Pb, as well as for As in female bears increased with age. Arsenic in males and Hg concentrations decreased with age. We detected elevated levels of toxic elements in bears from landscapes that are among the most pristine on the planet. Sources are unknown but anthropogenic emissions are most likely involved. All study areas face upcoming change: Increasing tourism and mining in Alaska and more intensive forestry in Scandinavia, combined with global climate change in both regions. Baseline contaminant concentrations as presented here are important knowledge in our changing world.

Alternosema astaquatica n. sp. (Microsporidia: Enterocytozoonida), a systemic parasite of the crayfish Faxonius virilis.

Crayfish have strong ecological impacts in freshwater systems, yet our knowledge of their parasites is limited. This study describes the first systemic microsporidium (infects multiple tissue types) Alternosema astaquatica n. sp. (Enterocytozoonida) isolated from a crayfish host, Faxonius virilis, using histopathology, transmission electron microscopy, gene sequencing, and phylogenetics. The parasite develops in direct contact with the host cell cytoplasm producing mature spores that are monokaryotic and ellipsoid in shape. Spores have 9-10 coils of the polar filament and measure 3.07 ± 0.26 µm (SD) in length and 0.93 ± 0.08 µm (SD) in width. Our novel isolate has high genetic similarity to Alternosema bostrichidis isolated from terrestrial beetles; however, genetic data from this parasite is restricted to a small fragment (396 bp) of the SSU gene. Additional data related to spore morphology and development, host, environment, and ecology indicate that our novel isolate is distinct from A. bostrichidis, which supports a new species description. Alternosema astaquatica n. sp. represents a novel member of the Orthosomella-like group which appears to be a set of opportunists within the Enterocytozoonida. The presence of this microsporidium in F. virilis could be relevant for freshwater ecosystems across this crayfish's broad geographic range in North America and may affect interactions between F. virilis and invasive rusty crayfish Faxonius rusticus in the Midwest USA.

Cambaraspora faxoni n. sp. (Microsporidia: Glugeida) from native and invasive crayfish in the USA and a novel host of Cambaraspora floridanus.

Crayfishes are among the most widely introduced freshwater taxa and can have extensive ecological impacts. Knowledge of the parasites crayfish harbor is limited, yet co-invasion of parasites is a significant risk associated with invasions. In this study, we describe a novel microsporidium, Cambaraspora faxoni n. sp. (Glugeida: Tuzetiidae), from two crayfish hosts in the Midwest USA, Faxonius virilis and Faxonius rusticus. We also expand the known host range of Cambaraspora floridanus to include Procambarus spiculifer. Cambaraspora faxoni infects muscle and heart tissue of F. rusticus and develops within a sporophorous vesicle. The mature spore measures 3.22 ± 0.14 µm in length and 1.45 ± 0.13 µm in width, with 8-9 turns of the polar filament. SSU sequencing indicates the isolates from F. virilis and F. rusticus were identical (100%) and 93.49% similar to C. floridanus, supporting the erection of a new species within the Cambaraspora genus. The novel parasite was discovered within the native range of F. rusticus (Ohio, USA) and within a native congeneric (F. virilis) in the invasive range of F. rusticus (Wisconsin, USA). Faxonius virilis is invasive in other regions. This new parasite could have been introduced to Wisconsin with F. rusticus or it may be a generalist species with a broad distribution. In either case, this parasite infects two crayfish species that have been widely introduced to new drainages throughout North America and could have future effects on invasion dynamics or impacts.

A conserved subcomplex within the bacterial cytokinetic ring activates cell wall synthesis by the FtsW-FtsI synthase.

Cell division in bacteria is mediated by a multiprotein assembly called the divisome. A major function of this machinery is the synthesis of the peptidoglycan (PG) cell wall that caps the daughter poles and prevents osmotic lysis of the newborn cells. Recent studies have implicated a complex of FtsW and FtsI (FtsWI) as the essential PG synthase within the divisome; however, how PG polymerization by this synthase is regulated and coordinated with other activities within the machinery is not well understood. Previous results have implicated a conserved subcomplex of division proteins composed of FtsQ, FtsL, and FtsB (FtsQLB) in the regulation of FtsWI, but whether these proteins act directly as positive or negative regulators of the synthase has been unclear. To address this question, we purified a five-member Pseudomonas aeruginosa division complex consisting of FtsQLB-FtsWI. The PG polymerase activity of this complex was found to be greatly stimulated relative to FtsWI alone. Purification of complexes lacking individual components indicated that FtsL and FtsB are sufficient for FtsW activation. Furthermore, support for this activity being important for the cellular function of FtsQLB was provided by the identification of two division-defective variants of FtsL that still form normal FtsQLB-FtsWI complexes but fail to activate PG synthesis. Thus, our results indicate that the conserved FtsQLB complex is a direct activator of PG polymerization by the FtsWI synthase and thereby define an essential regulatory step in the process of bacterial cell division.

Posterior cruciate-retaining total knee arthroplasty exhibits small kinematic changes in the first postoperative year.

PURPOSE: Fluoroscopic knee kinematics have historically been quantified at least 1 year after total knee arthroplasty (TKA). The purpose of this study was to longitudinally assess knee kinematics at 6-12 weeks, 6 months, and 1 year after TKA to determine if earlier evaluation may be justified. METHODS: Twenty-one patients participated after undergoing TKA with a posterior cruciate ligament-retaining fixed-bearing prosthesis. Fluoroscopic examinations of lunge, kneel, and step-up activities were performed at 12 ± 4 weeks (V1), 7 ± 2 months (V2), and 13 ± 2 months (V3) postoperatively. Images were analyzed using a three-dimensional to two-dimensional image registration technique. Maximum flexion poses for lunging and kneeling were compared between visits with repeated-measures statistical tests. For the step-up activity, mixed-effects linear models were constructed for condylar anteroposterior (AP) contact points and tibial internal rotation throughout flexion. Estimated marginal means of fitted values were plotted with 95% confidence intervals and used to compare mean kinematics between visits. RESULTS: There were no significant changes in maximum lunging flexion over time (p = 0.405), though significant increases in maximum kneeling flexion were observed between V1 (106 ± 8°) and V2 (110 ± 9°) (p = 0.006), and V1 and V3 (113 ± 9°) (p = 0.0003). While statistical differences were calculated for lunging medial condyle AP translation and kneeling tibial internal rotation, absolute differences in condylar AP contact locations were less than ~ 2 mm between all visits during both movements. For the step-up activity, tibial internal rotation increased with flexion, and there were pair-wise significant differences at all flexion angles between V1-V2 (p < 0.001) and V1-V3 (p < 0.001). Anterior medial condylar translation was observed with flexion, with pair-wise significant differences present for V1-V3 (p = 0.005) and V2-V3 (p < 0.001). The lateral condyle exhibited initial posterior translation followed by anterior translation with increasing flexion, with pair-wise differences between all visits (p < 0.005 for all comparisons). CONCLUSION: Though statistical differences were observed between visits for all activities, variations in estimated mean condylar positions were within ~ 2 mm from ~ 12 weeks to 1 year. Considering measurement error averages approximately 1 mm for sagittal plane translations, these results indicate that knee kinematics during kneel, lunge, and step-up activities may be sustained from as early as 12 weeks after TKA.

Joint Track Machine Learning: An Autonomous Method of Measuring Total Knee Arthroplasty Kinematics From Single-Plane X-Ray Images.

BACKGROUND: Dynamic radiographic measurements of 3-dimensional (3-D) total knee arthroplasty (TKA) kinematics have provided important information for implant design and surgical technique for over 30 years. However, current methods of measuring TKA kinematics are too cumbersome, inaccurate, or time-consuming for practical clinical application. Even state-of-the-art techniques require human-supervision to obtain clinically reliable kinematics. Eliminating human supervision could potentially make this technology practical for clinical use. METHODS: We demonstrate a fully autonomous pipeline for quantifying 3D-TKA kinematics from single-plane radiographic imaging. First, a convolutional neural network (CNN) segmented the femoral and tibial implants from the image. Second, those segmented images were compared to precomputed shape libraries for initial pose estimates. Lastly, a numerical optimization routine aligned 3D implant contours and fluoroscopic images to obtain the final implant poses. RESULTS: The autonomous technique reliably produces kinematic measurements comparable to human-supervised measures, with root-mean-squared differences of less than 0.7 mm and 4° for our test data, and 0.8 mm and 1.7° for external validation studies. CONCLUSION: A fully autonomous method to measure 3D-TKA kinematics from single-plane radiographic images produces results equivalent to a human-supervised method, and may soon make it practical to perform these measurements in a clinical setting.

Epigenetic Modulation of Inflammatory Pathways in Myometrial Stem Cells and Risk of Uterine Fibroids.

The period during which tissue and organ development occurs is particularly vulnerable to the influence of environmental exposures. However, the specific mechanisms through which biological pathways are disrupted in response to developmental insults, consequently elevating the risk of hormone-dependent diseases, such as uterine fibroids (UFs), remain poorly understood. Here, we show that developmental exposure to the endocrine-disrupting chemical (EDC), diethylstilbestrol (DES), activates the inflammatory pathways in myometrial stem cells (MMSCs), which are the origin of UFs. Significantly, the secretome of reprogrammed MMSCs enhances the expression of critical inflammation-related genes in differentiated myometrial cells through the paracrine mechanism, which amplifies pro-inflammatory and immune suppression signaling in the myometrium. The expression of reprogrammed inflammatory responsive genes (IRGs) is driven by activated mixed-lineage leukemia protein-1 (MLL1) in MMSCs. The deactivation of MLL reverses the reprogramming of IRG expression. In addition, the inhibition of histone deacetylases (HDACs) also reversed the reprogrammed IRG expression induced by EDC exposure. This work identifies the epigenetic mechanisms of MLL1/HDAC-mediated MMSC reprogramming, and EDC exposure epigenetically targets MMSCs and imparts an IRG expression pattern, which may result in a "hyper-inflammatory phenotype" and an increased hormone-dependent risk of UFs later in life.

Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene cluster.

Our understanding of the biofilm matrix components utilized by Gram-positive bacteria, and the signalling pathways that regulate their production are largely unknown. In a companion study, we developed a computational pipeline for the unbiased identification of homologous bacterial operons and applied this algorithm to the analysis of synthase-dependent exopolysaccharide biosynthetic systems. Here, we explore the finding that many species of Gram-positive bacteria have operons with similarity to the Pseudomonas aeruginosa pel locus. Our characterization of the pelDEADAFG operon from Bacillus cereus ATCC 10987, presented herein, demonstrates that this locus is required for biofilm formation and produces a polysaccharide structurally similar to Pel. We show that the degenerate GGDEF domain of the B. cereus PelD ortholog binds cyclic-3',5'-dimeric guanosine monophosphate (c-di-GMP), and that this binding is required for biofilm formation. Finally, we identify a diguanylate cyclase, CdgF, and a c-di-GMP phosphodiesterase, CdgE, that reciprocally regulate the production of Pel. The discovery of this novel c-di-GMP regulatory circuit significantly contributes to our limited understanding of c-di-GMP signalling in Gram-positive organisms. Furthermore, conservation of the core pelDEADAFG locus amongst many species of bacilli, clostridia, streptococci, and actinobacteria suggests that Pel may be a common biofilm matrix component in many Gram-positive bacteria.

Diagnosis and Follow-up of Incidental Liver Lesions in Children.

ABSTRACT: Incidental liver lesions are identified in children without underlying liver disease or increased risk of hepatic malignancy in childhood. Clinical and imaging evaluation of incidental liver lesions can be complex and may require a multidisciplinary approach. This review aims to summarize the diagnostic process and follow-up of incidental liver lesions based on review of the literature, use of state-of-the-art imaging, and our institutional experience. Age at presentation, gender, alpha fetoprotein levels, tumor size, and imaging characteristics should all be taken into consideration to optimize diagnosis process. Some lesions, such as simple liver cyst, infantile hemangioma, focal nodular hyperplasia (FNH), and focal fatty lesions, have specific imaging characteristics. Recently, contrast-enhanced ultrasound (CEUS) was Food and Drug Administration (FDA)-approved for the evaluation of pediatric liver lesions. CEUS is most specific in lesions smaller than 3 cm and is most useful in the diagnosis of infantile hemangioma, FNH, and focal fatty lesions. The use of hepatobiliary contrast in MRI increases specificity in the diagnosis of FNH. Recently, lesion characteristics in MRI were found to correlate with subtypes of hepatocellular adenomas and associated risk for hemorrhage and malignant transformation. Biopsy should be considered when there are no specific imaging characteristics of a benign lesion. Surveillance with imaging and alpha fetoprotein (AFP) should be performed to confirm the stability of lesions when the diagnosis cannot be determined, and whenever biopsy is not feasible.

The plot thickens: Ovipleistophora diplostomuri infects two additional species of Florida crayfish.

Ovipleistophora (Microsporidia) is a globally distributed genus of obligate parasites that infect fish, Crustacea, and trematodes. We report on two additional crayfish hosts, Procambarus pictus (Simms Creek, Florida) and Procambarus fallax (Santa Fe River, Florida), that exhibited signs of high-intensity microsporidian infection in the musculature. Sequence data (SSU) for the isolates were 99.79% and 99.97% similar to Ovipleistophora diplostomuri parasitizing Procambarus bivittatus. Additional screening of regional fish species (Lepomis macrochirus) revealed additional microsporidian diversity (Potaspora) but did not include Ovipleistophora. Detecting Clade V isolates in multiple host groups provides further evidence for trophic transmission among Clade V microsporidia.

College student motivations for and perceived impacts of volunteering with a nutrition and cooking education program for children.

To assess among college students their motivations for and perceived impacts of volunteering with Cooking Matters for Kids as part of No Kid Hungry NC. Seventeen college student volunteers responded to an online survey questionnaire assessing their motivations for volunteering and how they were impacted by their experiences. Motivational functions for volunteering (values, understanding, social, career, protective, enhancement) were assessed using the Volunteer Functions Inventory. The strongest motivational functions for volunteering were values and understanding. Students were also strongly motivated to volunteer related to the areas specifically addressed by the program (i.e., nutrition, public health, working with children). Perceived impacts of volunteering included being more comfortable working with children, improving knowledge/skills, gaining experience related to their future career, and having fun. Volunteering with Cooking Matters for Kids benefited college students. Issues addressed by the program and the desire to gain experience motivated students to volunteer.

PelX is a UDP-N-acetylglucosamine C4-epimerase involved in Pel polysaccharide-dependent biofilm formation.

Pel is a GalNAc-rich bacterial polysaccharide that contributes to the structure and function of Pseudomonas aeruginosa biofilms. The pelABCDEFG operon is highly conserved among diverse bacterial species, and Pel may therefore be a widespread biofilm determinant. Previous annotation of pel gene clusters has helped us identify an additional gene, pelX, that is present adjacent to pelABCDEFG in >100 different bacterial species. The pelX gene is predicted to encode a member of the short-chain dehydrogenase/reductase (SDR) superfamily, but its potential role in Pel-dependent biofilm formation is unknown. Herein, we have used Pseudomonas protegens Pf-5 as a model to elucidate PelX function as Pseudomonas aeruginosa lacks a pelX homologue in its pel gene cluster. We found that P. protegens forms Pel-dependent biofilms; however, despite expression of pelX under these conditions, biofilm formation was unaffected in a ΔpelX strain. This observation led us to identify a pelX paralogue, PFL_5533, which we designate here PgnE, that appears to be functionally redundant to pelX In line with this, a ΔpelX ΔpgnE double mutant was substantially impaired in its ability to form Pel-dependent biofilms. To understand the molecular basis for this observation, we determined the structure of PelX to 2.1 Å resolution. The structure revealed that PelX resembles UDP-GlcNAc C4-epimerases. Using 1H NMR analysis, we show that PelX catalyzes the epimerization between UDP-GlcNAc and UDP-GalNAc. Our results indicate that Pel-dependent biofilm formation requires a UDP-GlcNAc C4-epimerase that generates the UDP-GalNAc precursors required by the Pel synthase machinery for polymer production.

Multidose Priming and Delayed Boosting Improve Plasmodium falciparum Sporozoite Vaccine Efficacy Against Heterologous P. falciparum Controlled Human Malaria Infection.

BACKGROUND: A live-attenuated Plasmodium falciparum sporozoite (SPZ) vaccine (PfSPZ Vaccine) has shown up to 100% protection against controlled human malaria infection (CHMI) using homologous parasites (same P. falciparum strain as in the vaccine). Using a more stringent CHMI, with heterologous parasites (different P. falciparum strain), we assessed the impact of higher PfSPZ doses, a novel multi-dose prime regimen, and a delayed vaccine boost upon vaccine efficacy (VE). METHODS: We immunized 4 groups that each contained 15 healthy, malaria-naive adults. Group 1 received 5 doses of 4.5 x 105 PfSPZ (Days 1, 3, 5, and 7; Week 16). Groups 2, 3, and 4 received 3 doses (Weeks 0, 8, and 16), with Group 2 receiving 9.0 × 105/doses; Group 3 receiving 18.0 × 105/doses; and Group 4 receiving 27.0 × 105 for dose 1 and 9.0 × 105 for doses 2 and 3. VE was assessed by heterologous CHMI after 12 or 24 weeks. Volunteers not protected at 12 weeks were boosted prior to repeat CHMI at 24 weeks. RESULTS: At 12-week CHMI, 6/15 (40%) participants in Group 1 (P = .04) and 3/15 (20%) participants in Group 2 remained aparasitemic, as compared to 0/8 controls. At 24-week CHMI, 3/13 (23%) participants in Group 3 and 3/14 (21%) participants in Group 4 remained aparasitemic, versus 0/8 controls (Groups 2-4, VE not significant). Postboost, 9/14 (64%) participants versus 0/8 controls remained aparasitemic (3/6 in Group 1, P = .025; 6/8 in Group 2, P = .002). CONCLUSIONS: Administering 4 stacked priming injections (multi-dose priming) resulted in 40% VE against heterologous CHMI, while dose escalation of PfSPZ using single-dose priming was not significantly protective. Boosting unprotected subjects improved VE at 24 weeks, to 64%. CLINICAL TRIALS REGISTRATION: NCT02601716.