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Abstract Introduction International guidelines recommend routine screening for syphilis (aetiological agent: Treponema pallidum subspecies pallidum) amongst key populations and vulnerable populations using tests detecting treponemal and non-treponemal antibodies. Whilst treponemal tests have high sensitivities and specificities, they differ regarding subjective or objective interpretation, throughput and workload. Chemiluminescence immunoassays (CLIAs) are cost- and time-effective automated methods for detecting treponemal antibodies. The Treponema pallidum particle agglutination assay (TPPA) has been considered the “gold standard” treponemal assay, however, this includes a highly manual procedure, low throughput and subjective interpretation. The present multi-country study evaluated the ADVIA Centaur® Syphilis CLIA (Siemens Healthcare) assay compared to the reference SERODIA-TP·PA® (Fujirebio Diagnostics) for the serodiagnosis of syphilis amongst men who have sex with men (MSM). Method 1,485 MSM were enrolled in Brighton (UK), Malta, and Verona (Italy) as part of a larger WHO multi-country and multi-site ProSPeRo study. Ethical approval was obtained. Serum was tested with the ADVIA Centaur® Syphilis CLIA assay and SERODIA-TP·PA®, in accordance with the manufacturers’ instructions, for a first round of validation. A second round of validation was carried out for discrepant results that were additionally tested with both Western Blot (Westernblot EUROIMMUN®) and an Immunoblot (INNO-LIA, Fujirebio Diagnostics). Sensitivity, specificity, positive and negative predictive value (PPV and NPV), likelihood ratios (positive/negative), and the Diagnostic Odds Ratio (DOR)/pre-post-test probability (Fagan's nomogram) were calculated. Results Out of 1,485 eligible samples analysed in the first phase, the SERODIA-TP·PA® identified 360 positive and 1,125 negative cases. The ADVIA Centaur® Syphilis CLIA assay (Siemens) identified 366 positives, missclassifying one TPPA-positive sample. In the second phase, the ADVIA Centaur® Syphilis CLIA resulted in 1 false negative and 4 false positive results. Considering the syphilis study prevalence of 24% (95% CI: 22–26.7), The sensitivity of the ADVIA Centaur® Syphilis CLIA assay was 99.7% (95% CI: 98.5–100), and the specificity was 99.4% (95% CI: 98.7–99.7). The ROC area values were 0.996 (95% CI: 0.992–0.999), and both the PPV and NPV values were above 98% (PPV 98.1%, 95% CI: 96.1–99.2; NPV 99.9%, 95% CI: 99.5–100). Conclusions The ADVIA Centaur® Syphilis CLIA assay showed similar performance compared to the SERODIA-TP·PA®. Considering the study is based on QUADAS principles and with a homogeneous population, results are also likely to be generalisable to MSM population but potentially not applicable to lower prevalence populations routinely screened for syphilis. The automated CLIA treponemal assay confirmed to be accurate and appropriate for routine initial syphilis screening, i.e. when the reverse testing algorithm is applied.

Marine regions are undergoing rapid evolution, primarily driven by natural and anthropogenic activities. Safeguarding these ecosystems necessitates the ability to observe their physical features and control processes with precision in both space and time. This demands the acquisition of precise and up-to-date information regarding several marine parameters. Thus, to gain a comprehensive understanding of these ecosystems, this study employs remote sensing techniques, Machine Learning algorithms and traditional in situ approaches. Together, these serve as valuable tools to help comprehend the distinctive parametric characteristics and mechanisms occurring within these regions of the Maltese archipelago. An empirical workflow was implemented to predict the spatial and temporal variations in sea surface salinity and sea surface temperature from 2022 to 2024. This was achieved by leveraging Sentinel-2 satellite platforms, the random forest Machine Learning algorithm, and in situ data collected from sea gliders and floats. Subsequently, the numerical data generated by the random forest algorithm were validated with different error metrics and converted into visual representations to illustrate the sea surface salinity and sea surface temperature variations across the Maltese Islands. The random forest algorithm demonstrated strong performance in predicting sea surface salinity and sea surface temperature, indicating its capability to handle dynamic parameters effectively. Additionally, the parametric maps generated for all three years provided a clear understanding of both the spatial and temporal changes for these two parameters.

The European X-ray Free Electron Laser (XFEL) and Linac Coherent Light Source (LCLS) II are extremely intense sources of X-rays capable of generating Serial Femtosecond Crystallography (SFX) data at megahertz (MHz) repetition rates. Previous work has shown that it is possible to use consecutive X-ray pulses to collect diffraction patterns from individual crystals. Here, we exploit the MHz pulse structure of the European XFEL to obtain two complete datasets from the same lysozyme crystal, first hit and the second hit, before it exits the beam. The two datasets, separated by

Osteosarcoma (OS) is a bone cancer exhibiting a 20% survival rate for metastatic patients, which motivates the development of new therapeutic options. Among the various new treatment approaches, modulation of autophagy is the subject of rising interest. In addition to its pro-survival role in established tumors, autophagy recently emerged as an active player in the crosstalk between tumor and stromal cells. In OS, although the knockdown of key autophagy genes in human cell lines demonstrates a protumoral role of autophagy, the analysis of patient tumors indicates that lack of LC3-positive punctae at resection following neoadjuvant chemotherapy is a poor prognostic marker, suggesting that loss of autophagy is not detrimental for the tumor. In the present work, we analyzed the consequences of autophagy inactivation in OS cells both on tumor development and on bone microenvironment in an orthotopic syngeneic model. We found that inactivation of the autophagy-essential gene Atg5 in OS cells decreases their tumorigenic properties in vitro. However, these effects were no longer observed in vivo, likely due to microenvironment modifications such as overexpression of the major OS-promoting factor TGF-β or increased infiltration of Foxp3-positive and CD31-positive cells in Atg5 KO tumors. In addition, autophagy-deficient tumor cells stimulate the in vitro formation of osteoclast, the cells in charge of bone resorption which can release bone matrix-embedded growth factors thereby stimulating tumor growth. Taken together, these results suggest that Atg5 inactivation in OS cells is associated with microenvironment modifications known as poor prognosis-associated factors in OS, and could thus balance the negative cell-autonomous effects of autophagy suppression. Abbreviations: ACTB -β-actin; Atg -autophagy-related; Baf-A1 - Bafilomycin-A1; CSC -cancer stem cells; Col1A -type 1a collagen; d -day; HBSS -Hank’s balanced salt solution; LC3 -microtubule-associated protein 1 light chain 3 protein; SQSTM1/p62 -sequestosome; OB -osteoblast; OC -osteoclast; OS -osteosarcoma; TEM -transmission electron microscopy; TGF-β -transforming growth factor β; TRAP -acid phosphatase 5, tartrate-resistant.

Free-electron lasers are capable of high repetition rates and it is assumed that protein crystals often do not survive the first X-ray pulse. Here the authors address these issues with a demonstration of multi-hit serial crystallography in which multiple FEL pulses interact with the sample without destroying it.

Monoolein-based liquid crystal phases are established media that are researched for various biological applications, including drug delivery. While water is the most common solvent for self-assembly, some ionic liquids (ILs) can support lipidic self-assembly. However, currently, there is limited knowledge of IL-lipid phase behavior in ILs. In this study, the lyotropic liquid crystal phase behavior of monoolein was investigated in six protic ILs known to support amphiphile self-assembly, namely ethylammonium nitrate, ethanolammonium nitrate, ethylammonium formate, ethanolammonium formate, ethylammonium acetate, and ethanolammonium acetate. These ILs were selected to identify specific ion effects on monoolein self-assembly, specifically increasing the alkyl chain length of the cation or anion, the presence of a hydroxyl group in the cation, and varying the anion. The lyotropic liquid crystal phases with 20–80 wt. % of monoolein were characterized over a temperature range from 25 to 65 °C using synchrotron small angle x-ray scattering and cross-polarized optical microscopy. These results were used to construct partial phase diagrams of monoolein in each of the six protic ILs, with inverse hexagonal, bicontinuous cubic, and lamellar phases observed. Protic ILs containing the ethylammonium cation led to monoolein forming lamellar and bicontinuous cubic phases, while those containing the ethanolammonium cation formed inverse hexagonal and bicontinuous cubic phases. Protic ILs containing formate and acetate anions favored bicontinuous cubic phases across a broader range of protic IL concentrations than those containing the nitrate anion. [ABSTRACT FROM AUTHOR]

Serial crystallography of membrane proteins often employs high-viscosity injectors (HVIs) to deliver micrometre-sized crystals to the X-ray beam. Typically, the carrier medium is a lipidic cubic phase (LCP) media, which can also be used to nucleate and grow the crystals. However, despite the fact that the LCP is widely used with HVIs, the potential impact of the injection process on the LCP structure has not been reported and hence is not yet well understood. The self-assembled structure of the LCP can be affected by pressure, dehydration and temperature changes, all of which occur during continuous flow injection. These changes to the LCP structure may in turn impact the results of X-ray diffraction measurements from membrane protein crystals. To investigate the influence of HVIs on the structure of the LCP we conducted a study of the phase changes in monoolein/water and monoolein/buffer mixtures during continuous flow injection, at both atmospheric pressure and under vacuum. The reservoir pressure in the HVI was tracked to determine if there is any correlation with the phase behaviour of the LCP. The results indicated that, even though the reservoir pressure underwent (at times) significant variation, this did not appear to correlate with observed phase changes in the sample stream or correspond to shifts in the LCP lattice parameter. During vacuum injection, there was a three-way coexistence of the gyroid cubic phase, diamond cubic phase and lamellar phase. During injection at atmospheric pressure, the coexistence of a cubic phase and lamellar phase in the monoolein/water mixtures was also observed. The degree to which the lamellar phase is formed was found to be strongly dependent on the co-flowing gas conditions used to stabilize the LCP stream. A combination of laboratory-based optical polarization microscopy and simulation studies was used to investigate these observations.

Marine regions are undergoing rapid evolution, primarily driven by natural and anthropogenic activities. Safeguarding these ecosystems necessitates the ability to observe their physical features and control processes with precision in both space and time. This demands the acquisition of precise and up-to-date information regarding several marine parameters. Thus, to gain a comprehensive understanding of these ecosystems, this study employs remote sensing techniques, Machine Learning algorithms and traditional in situ approaches. Together, these serve as valuable tools to help comprehend the distinctive parametric characteristics and mechanisms occurring within these regions of the Maltese archipelago. An empirical workflow was implemented to predict the spatial and temporal variations in sea surface salinity and sea surface temperature from 2022 to 2024. This was achieved by leveraging Sentinel-2 satellite platforms, the random forest Machine Learning algorithm, and in situ data collected from sea gliders and floats. Subsequently, the numerical data generated by the random forest algorithm were validated with different error metrics and converted into visual representations to illustrate the sea surface salinity and sea surface temperature variations across the Maltese Islands. The random forest algorithm demonstrated strong performance in predicting sea surface salinity and sea surface temperature, indicating its capability to handle dynamic parameters effectively. Additionally, the parametric maps generated for all three years provided a clear understanding of both the spatial and temporal changes for these two parameters. [ABSTRACT FROM AUTHOR]

The measurement of daily step counts is a widely adopted and efficacious approach for assessing children’s physical activity levels. With the aim of finding out the effect of daily PA sessions in schools on the total PA levels in terms of steps taken, two (treatment and control) Year Four classrooms (with students aged from eight to nine years old) in three participating schools in Malta participated in this study. A random sample of 45 children across all the treatment and control groups wore pedometers for five continuous school days. Additional data were logged to facilitate data analysis. Statistical analysis was conducted on data from 18 participants, as incomplete data were eliminated. The findings show that a daily physical activity programme was sufficient to make a significant difference in the number of steps taken at school. It also made the difference between Maltese schools being either under par (if not engaged in daily PA at school) or among high-performing nations (if engaged in daily PA at school) in terms of the proportional contribution of steps taken at school to children’s total PA. However, the capacity for school-based activity to exert wider effects on total steps taken throughout the entire day appears to be limited. Reported engagement in additional formal sports activities outside school also had limited effects on steps overall. Finally, a statistical model incorporating sex, daily PA, sports participation, and BMI as exogenous variables accounted only for around a quarter of the variation in average daily pedometer-measured PA, leaving approximately 75% of the variation unexplained. Implications of the findings are discussed, including recommendations for alternative ways of conceptualising and promoting health-related physical activity and exercise.

Intensity-correlation measurements allow access to nanostructural information on a range of ordered and disordered materials beyond traditional pair-correlation methods. In real space, this information can be expressed in terms of a pair-angle distribution function (PADF) which encodes three- and four-body distances and angles. To date, correlation-based techniques have not been applied to the analysis of microstructural effects, such as preferred orientation, which are typically investigated by texture analysis. Preferred orientation is regarded as a potential source of error in intensity-correlation experiments and complicates interpretation of the results. Here, the theory of preferred orientation in intensity-correlation techniques is developed, connecting it to the established theory of texture analysis. The preferred-orientation effect is found to scale with the number of crystalline domains in the beam, surpassing the nanostructural signal when the number of domains becomes large. Experimental demonstrations are presented of the orientation-dominant and nanostructure-dominant cases using PADF analysis. The results show that even minor deviations from uniform orientation produce the strongest angular correlation signals when the number of crystalline domains in the beam is large.

C-C motif chemokine receptor-like 2 (CCRL2) is a non-signaling 7 transmembrane receptor that binds chemotactic ligands to shape leukocyte recruitment to sites of inflammation. However, there is a lack of consensus on the ligands that directly bind CCRL2 or their functional impact. Studies with CCRL2 knockout mice have demonstrated that neutrophils have impaired degranulation and migration in response to CXCL8, where the underlying molecular mechanism is proposed to be due to the formation of CCRL2 heterodimers with the chemokine receptor CXCR2. Herein, we characterized the ligands that bind directly to CCRL2 and interrogated the impact of CCRL2 neutralization on CXCL8 signaling in neutrophils using pharmacological antibody tools. Using flow cytometry and Surface Plasmon Resonance microscopy (SPRm) cell binding experiments, we confirmed that chemerin, but not previously reported C-C chemokines, binds CCRL2. Furthermore, we identified human and mouse CCRL2 antibodies that neutralized chemerin binding to CCRL2. Unexpectedly, we found that neutralization of CCRL2 with these antibodies did not attenuate CXCL8-induced human neutrophil degranulation nor CXCL8-induced murine neutrophil recruitment to the peritoneum. Based on the observed differences in modulating CCRL2 function with neutralizing antibodies compared to the reported CCRL2 deficient murine models, we hypothesize that the ligand binding function of CCRL2 is dispensable for CXCL8 signaling in neutrophils. Finally, extensive profiling of CCRL2 expression on peripheral blood leukocytes revealed monocytes, dendritic cells (DC), and subpopulations of natural killer T (NKT) cells as additional targets, highlighting potential roles for CCRL2 in human cell types beyond neutrophils that warrants future investigation.

The new European X-ray Free-Electron Laser is the first X-ray free-electron laser capable of delivering X-ray pulses with a megahertz inter-pulse spacing, more than four orders of magnitude higher than previously possible. However, to date, it has been unclear whether it would indeed be possible to measure high-quality diffraction data at megahertz pulse repetition rates. Here, we show that high-quality structures can indeed be obtained using currently available operating conditions at the European XFEL. We present two complete data sets, one from the well-known model system lysozyme and the other from a so far unknown complex of a β-lactamase from K. pneumoniae involved in antibiotic resistance. This result opens up megahertz serial femtosecond crystallography (SFX) as a tool for reliable structure determination, substrate screening and the efficient measurement of the evolution and dynamics of molecular structures using megahertz repetition rate pulses available at this new class of X-ray laser source.

Abstract Sprouts are particularly prone to microbial contamination due to their high nutrient content and the warm temperatures and humid conditions needed for their production. Therefore, disinfection is a crucial step in food processing as a means of preventing the transmission of bacterial, parasitic and viral pathogens. In this study, a dielectric coplanar surface barrier discharge (DCSBD) system was used for the application of cold atmospheric plasma (CAP), plasma activated water (PAW) and their combination on mung bean seeds. Germination assessments were performed in a test tube set-up filled with glass beads and the produced irrigation water. Overall, it was found that the combined seed treatment with direct air CAP (350 W) and air PAW had no negative impact on mung bean seed germination and growth, nor the concentration of secondary metabolites within the sprouts. These treatments also reduced the total microbial population in sprouts by 2.5 log CFU/g. This research reports for first time that aside from the stimulatory effect of plasma discharge on seed surface disinfection, sustained plasma treatment through irrigation of treated seeds with PAW can significantly enhance seedling growth. The positive outcome and further applications of different forms, of plasma i.e., gaseous and aqueous, in the agro-food industry is further supported by this research.

MAL and MyD88 are downstream adaptors of Toll-like receptors (TLR) and the MAL TIR domain forms filaments in vitro, which in turn nucleate the assembly of crystalline arrays of the MyD88 TIR domain. Here, the authors present the structure of these MyD88 TIR crystalline arrays solved by both microcrystal electron diffraction and serial femtosecond crystallography, and they show with mutagenesis experiments that MyD88 interface residues are important for TLR4 signaling in vivo.

The current study provides new data on the known records of the Atlantic blue crab, Callinectes sapidus Rathbun, 1896, from the Maltese Islands through two ovigerous female specimens collected from Salini (Malta), a Natura 2000 site, in August 2020. Additional new records of the species were also presented from the San Leonardo River estuary, Syracuse (Sicily, Italy), in August 2022. The specimens collected from both Malta and Sicily were identified using both morphological and molecular analyses. The latter has shown that all the specimens analysed share their mitochondrial DNA barcode region with the most recorded haplotype of the same species from the Mediterranean Sea. Knowing the new distribution and range expansion records of alien species is important for the continuous monitoring of marine alien species, which is essential for the implementation of the best possible mitigation measures. This is especially relevant for C. sapidus, as it is considered one of the worst invaders of the Mediterranean Sea. Therefore, given that this alien species is edible, fishing for it may allow some control over its population size and further expansion. Knowing its distribution is crucial to inform fishers about its exact location, making such a mitigation measure more effective.

We report a bioinspired approach to tune surface nanostructures by soft‐template electropolymerization in micellar conditions. Monomers highly favoring π‐stacking interactions are particularly interesting for favoring the polymer deposition in one direction. Here, original triphenylamine‐based monomers di‐substituted by thiophene and carbazole are investigated. Conjugated building blocks monomers are tested to favor deposition vs polymerization, even if the monomers are not perfectly planar. The carbazole derivatives have a much higher electrodeposition capacity than the thiophene derivatives, which is unexpected if thiophene and carbazole are taken alone. Moreover, in all electrodeposited films, monomer seems to be present as shown by cyclic voltammetry experiments, confirming previous works. The amount of monomer vs oligomers is highly dependent on the investigated monomer. For the resulting surface structures, hollow spheres and nanoribbons are particularly formed with some investigated monomers by cyclic voltammetry while nanotubes are observed at constant potential. The formation of nanotubes indicates a polymer growth more favored in one direction. These surfaces are less hydrophobic (water contact angle up to 111.5°) compared to films with spherical nanoparticles but these results can be explained only by the presence of air inside the surface roughness. [ABSTRACT FROM AUTHOR]

Bio‐inspired by natural species and previous research works, nanotubes are prepared by soft‐template electropolymerization in micellar solution. Various thiophene‐based and carbazole‐based monomers are designed from a triphenylamine core. With one of the carbazole‐based monomers, which is fully conjugated and with a star‐like structure, gecko‐like surface structures are obtained even if the structure is not perfectly planar. Extremely long and densely packed‐microtubes are reached especially at constant potential. However, the surfaces are not both highly hydrophobic and with strong water adhesion, but are superhydrophilic. These wetting properties can be explained by a high surface energy (γSV)=41.9 mN m−1 compared to gecko foot even if the natural gecko feet are also chemically inhomogeneous, or by differences in the arrangement of the surface structures. [ABSTRACT FROM AUTHOR]