All components run without external utilities such as compressed air or three-phase electricity, and require just three mains sockets for operation. The system can be manually transported in a lightweight van and rapidly installed on a small estates footprint in a hospital setting.The polariser system consists of six main functional components: (i) a laser diode array and optics; (ii) a B129Xe for routine clinical lung MRI. To test the concept of portability and rapid deployment, it was transported 200 km, installed at a hospital with no previous experience with the technology and 129Xe MR images of a diagnostic quality were acquired the day after system transport and installation.The polariser routinely provides polarised 129Xe hyperpolariser system could form the basis of a cost-effective platform for wider clinical dissemination and multicentre evaluation of 129Xe lung MR imaging.This portable 129Xe MRI with a portable hyperpolariser system.Our work successfully demonstrates the feasibility of multicentre clinical  Additionally, hyperpolarised 129Xe MRI has been demonstrated in other organs (e.g. the brain and kidneys) where dissolved-phase xenon can be used to study tissue perfusion and gas exchange. While conventional proton (1H) MRI makes use of proton spin densities in tissues and blood in the lungs, owing to the comparatively lower density of 129Xe in the lung airspaces or dissolved in blood and other tissues, hyperpolarised 129Xe MRI relies on exogenous enhancement (“hyperpolarisation”) of the magnetisation of the 129Xe nuclei using laser polarisation techniques. This enhancement leads to a five orders of magnitude MR signal enhancement for hyperpolarised 129Xe when compared to thermally polarised129Xe. This is critical in enabling MR images of 129Xe in the lung airspaces, as well as in well-perfused organs such as the kidneys and the brain. Early development of noble gas MRI of human lungs focussed on polarised helium-3 (3He) because of its isotopic purity and higher intrinsic MR signal. In recent years, however, noble gas lung MRI research has moved from 3He towards 129Xe due to reasons of greater availability and thus lower cost of xenon, which can be extracted from the air.Magnetic resonance imaging (MRI) of hyperpolarised xenon-129  with sufficient 129Xe polarisation levels (>20%) to enable clinical interpretation of 129Xe MR images.For clinical applications of hyperpolarised 129Xe is spin-exchange optical pumping. Two methods of spin-exchange optical pumping have emerged from physics research for clinical 129Xe MRI applications: batch mode and continuous-flow mode. The main operational difference between batch and continuous-flow mode is that batch mode does not require cryogenic accumulation of xenon, whereas continuous-flow mode does. This difference generally leads to two distinct performance regimes for batch and continuous-flow mode, namely (i) batch mode yields higher 129Xe polarisation (50%–90%) with slower xenon production rates (100–1000 mL/h) and (ii) continuous-flow mode yields lower 129Xe polarisation (20%–50%) but with faster production rates (1000–3500 mL/h). The faster production rates currently obtainable on continuous-flow hyperpolarisers offer increased potential for routine clinical 129Xe MRI when compared to batch mode systems given typical xenon throughput demands. For example, at the University of Sheffield, where the modality is being used clinically for lung disease assessment, the throughput is as many as five patients/day, each requiring between 1,000 mL and 3,500 mL of hyperpolarised 129Xe for a given MR scan protocol. While modern continuous-flow polarisers demonstrate the feasibility of clinical 129Xe MRI, widespread clinical uptake is still restricted by the need for additional lab space and/or the electrical and mechanical estates utilities necessary to install, house and run a commercially-available 129Xe polariser.The most common technique used to magnetise 129Xe polariser for rapid low-cost deployment for clinical 129Xe lung MRI. The design is a self-contained, stand-alone and transportable device that can quickly be installed on-site or be taken temporarily to another facility where there is interest in using hyperpolarised 129Xe. Thus removing the need for costly stationary equipment installation and not interfering with existing infrastructure required for operation. In this work we build on previous work aimed at increasing portability of 129Xe polariser technology, by developing a compact 129Xe hyperpolariser optimised for high-volume xenon production for clinical lung MRI in a hospital setting. The design criteria of the system were as follows: (i) readily transportable to an external clinical MRI facility; (ii) compact with no need for additional site infrastructure (i.e. it should run on mains electricity without the need for compressed air supplies), (iii) regulatory approved for in vivo use; and (iv) provide doses of 129Xe for high-quality clinical lung MRI in <20 min.Our solution was to design and build a portable B0 coil assembly; (iii) an oven containing a cylindrical optical cell; (iv) NMR and optical spectrometers; (v) a laser power meter; (vi) a gas-handling manifold; and (vii) a cryostat within a permanent magnetic field. See ®, QPC Lasers, Sylmar, US) providing ~50 W of laser light tuned to the rubidium valence electron D1 line (795 nm). The class IV laser diode is coupled to an optical fibre to custom optics that produce left circularly polarised light in a homogeneous circular beam that is incident on the oven window. Stray laser light is mitigated by enclosing the beam, optics and oven with front and back plates, however laser safety goggles are worn as an additional precautionary measure.The polariser comprises the following components: (i) a laser diode array with beam shaping optics; (ii) an electromagnetic static 3  loaded with ~1 g rubidium. The oven is heated using a bespoke air compressor / heating element combination. The oven and cell are contained within a magnetic field generated by a four-coil electromagnet using square coils with a side-length of ~60 cm for optimised field homogeneity [B0 field is maintained at 2.4 mT, corresponding to a 28 kHz resonance frequency for 129Xe nuclei. The gas manifold directs gas flow from a cylinder (located in a gas trolley ~1 m away from the system to prevent magnetic field distortion) containing 3% isotopically enriched xenon (86% 129Xe), 10% N2 and 87% He . After passing through a gas getter and pressure regulator set for a cell pressure of 2 bar, the gas enters the optical cell via high-vacuum PTFE stop-cock valves.The oven is made of compressed calcium silicate ceramic with antireflection glass windows and a cylindrical optical cell made from borosilicate glass with volume 1767 cm2, whereupon the xenon in the flowing gas mixture is cryogenically separated from He and N2, which are removed as exhaust gases through a vacuum line. The gas flow rate is controlled using a mass flow meter in line with a diaphragm vacuum pump which generates a downstream pressure of ~2 mbar. Once sufficient xenon snow is deposited in the spiral, it is defrosted by immersion in room-temperature water where the frozen xenon sublimates to the gas phase and flows into a Tedlar bag  ready for QC release to the subject. In situ laser monitoring is provided by an optical spectrometer and a photon power meter. A home-built low-field nuclear magnetic resonance spectrometer using a LabVIEW software interface and a National Instruments data acquisition board (PCIe-6351 X-series multifunction DAQ) is used for in-system signal monitoring of the polarised 129Xe. The signals from 129Xe is acquired using a 100-turn surface B1 coil located on top of the optical cell inside the oven.During continuous-flow operation, the xenon gas mixture is flowed through the cell in a direction counter to the incident laser light at a rate of 1000 sccm (standard cubic cm per minute), corresponding to a xenon flow rate of 1800 mL/h. The polarised gas exiting the cell is flowed through Tygon  tubing towards spiral glassware. The glassware is held within a field of 250 mT from a NdBFe horseshoe permanent magnet and submerged in a dewar containing liquid N129Xe polariser  under the Manufacturing Specials licence number MS-18739.The assembled 129Xe MRI. Lung imaging was performed using a transmit-receive vest coil  on a healthy male volunteer the next day after 3 h of commissioning on site on a 1.5 T MR scanner . Prior to running the ventilation sequence, we calibrated the flip angle and centre frequency from an inhaled dose of 1L of a 3% xenon gas mixture. Sequence details are shown within the caption of To test the concept of portability of the polariser, it was transported 200 km in a van to a novice clinical radiology site with no previous in-house experience of 129Xe acquired from the lungs of patients with the polariser in routine use at our MR imaging centre; inhaled dose 500 mL of xenon topped up to 1 litre with medical grade nitrogen acquired on a GE 450W 1.5 T MRI scanner housed in a relocatable mobile unit. 129Xe lung images from the healthy volunteer at the external imaging centre; inhaled dose 300 mL of xenon topped up to 1 litre with medical grade nitrogen. The 129Xe polarisation was measured to be ~30% (using the method described in ref.), and the 300 mL and 500 mL doses were generated in 10 min and 17 min (xenon production rate of 1800 mL/h). The signal-to-noise ratio over all image slices was measured to be >40.129Xe polariser for hyperpolarised gas lung MRI imaging. Portability of the system was demonstrated with a rapid turnkey installation at an external clinical imaging centre. From an installation and running perspective, the system fits easily in the back of a transit van, operates reliably on three 240V/50 Hz mains electricity sockets, and the on-board air compressor and heater is able to heat the oven to 140°C in ~20 min with <1°C fluctuation once thermal stability is reached. The system has been tested in various locations including in the close confines and noisy electrical environment of the equipment room of a relocatable mobile MRI scanner. The only additional requirements at the installation site are completion of risk assessments and also access to liquid nitrogen; this should not pose an impediment with respect to accessibility to other clinical MR centres, as all large hospitals should have access to liquid nitrogen storage.We have demonstrated the feasibility of a lightweight, high-volume production rate, portable  a metric that couples the quality and quantity of xenon gas available for patient inhalation within a given time period – i.e. it is a product of the operational 129Xe polarisation and the xenon volume production rate. It is quantified numerically as DER = f×P×Q, where f is the isotopic fraction of 129Xe , P is the fractional 129Xe polarisation and Q is the xenon volume production rate. Using this formula with a Q of 1800 mL/h and P of 0.30 for the system described here yields a DER equal to 540 mL/h, which enables acquisition of diagnostic-quality MR lung images using 500 mL of xenon gas produced in <20 min. The performance of our portable polariser is approaching that of a permanent clinical 129Xe polariser system, which has a DER value of ~1000 mL/h and is used routinely for diagnostic clinical lung imaging. It is worth noting that to facilitate multicentre applications of 129Xe MRI, as well as increasing a site’s accessibility to polariser technology, it is important to standardise xenon dosing and imaging protocols as outlined in ref. and to also consider the additional outlay on 129Xe RF coils and broadband RF amplifiers on the scanner itself.In order to compare operational performance against other polariser systems, we employ the metric dose equivalence rate (DER),129Xe gas. The system routinely provides high-SNR lung imaging, and can easily be transported and installed at novice clinical imaging centres where no specialist equipment is required for polariser installation. The system should form the basis of a cost-effective platform for wider dissemination and multicentre clinical evaluation of hyperpolarised 129Xe MR imaging. Increased access to 129Xe hyperpolarisation technology outlined in this work should create new opportunities to explore and utilise the clinical impact of 129Xe MRI for diagnosis, prognosis, and treatment response monitoring of a number of diseases within the lungs and beyond.In this work, we have demonstrated a self-contained, portable polariser that rapidly generates highly polarised


This is imaged with atomic and elementalresolution which reveal a prismatic configuration of the Eu planes.The results are supported by molecular dynamics simulations. Localmagnetic and susceptibility mappings show magnetic response in allnanowires, while a subset bearing a DC signal points to ferromagneticorder. These provide a mechanism for enhancing Zeeman responses, operationalat zero applied magnetic field. Such properties suggest that the obtainedstructures can serve as a preferred platform for time-reversal symmetrybroken one-dimensional states including intrinsic topological superconductivity.In the pursuit of magneto-electronic systems nonstoichiometricmagnetic elements commonly introduce disorder and enhance magneticscattering. We demonstrate the growth of (EuIn)As shells, with a uniquecrystal structure comprised of a dense net of Eu inversion planes,over InAs and InAs Therefore, magnetic topologyoffers unprecedented ability to control and manipulate exotic topologicalboundary modes, which are well beyond increased richness.Inducing magnetism into topologicalelectronic matter is a long-sought goal. It bears the promise to realizenovel time-reversal symmetry broken topological electronic statessuch as the anomalous quantum Hall insulator, axion insulator, andantiferromagnetic topological insulator, each hosting unique boundarymodes.3 Still, conflicting evidence fortime-reversal symmetry broken states has been reported. In transportmeasurements signatures of anomalous quantum Hall states have beenfound at low temperatures in magnetically doped Bi2Se3, Bi2Te3, and related ternary alloys.6 However, spectroscopically the induced time-reversal symmetry brokengap has been inconclusive.10However,the incorporation of magnetism into topologically classifiedmaterials produces a fundamental challenge, as the magnetic dopantsnecessarily introduce disorder. Indeed, initial attempts to dope topologicalinsulators with magnetic elements have resulted in strong spatialfluctuations in energy and momentum across the boundaries of the dopedsamples.2Se3.13 In general,those have not produced robust quantized responses. Nevertheless,zero bias conductance peaks at zero applied magnetic field, whichare believed to be a sign of topological superconductivity, were observedin semiconducting InAs nanowires (NWs) with EuS half-shells.14A different approach was taken to address theboundary modes aloneby growing magnetic layers over topologically classified materialssuch as EuS over Bi3Sn2S2,16 in which Fermi arc surface stateshave been imaged. In another antiferromagnetic topological insulator,MnBi2Te4,18 quantized Hall conductanceand the lack of it were demonstrated, hallmarking the realizationof anomalous quantum Hall and axion insulators, respectively.Recent efforts have thus concentratedon identifying stoichiometriccompounds that are intrinsically magnetic, as well as classified astopologically nontrivial. This approach has the dual benefit of affectingthe bulk topology class without introducing disorder. The classificationof magnetic topological compounds is fast-growing. Prime examplesare given by the ferromagnetic Weyl semimetal Cox1–Sbx NWcores. Bulk EuIn2As2 orders antiferromagneticallybelow a Néel temperature of 16 K.19 It is a type A antiferromagnet (AFM) with As-Eu-As ferromagneticlayers that order antiferromagnetically among themselves. It was predictedto become an axion insulator within the AFM phase.20 We obtain an initial success toward the demonstration ofintrinsically stoichiometric, magnetic semiconducting NWs. These (EuIn)Asshells over InAs and InAsx1–Sbx core NWs could serve as a superior platformfor induced topological superconductivity as well as other applicationsthat would benefit from the magnetic nature and the ability to controland manipulate it.Here we adopt a similar approach to semiconducting NWs. We groweuropium indium arsenide ((EuIn)As) shells over InAs and InAs23 on (001)-orientedsubstrates.24 The growth of InAs NWs ona (001) InAs substrate results in wurtzite (WZ) NWs that emerge from(111) nanofacets and are thus reclining with respect to the surface.Since there are two equivalent (111) directions, the reclining NWsoccasionally merge and form intersections. From the merging pointof such intersections a new stalactite NW emerges, which grows verticalto the substrate and has a pure zincblende (ZB) structure.25 Those stalactite ZB NWs provided an importantinsight into the growth of (EuIn)As shells, as will be discussed below.The (EuIn)As shell was subsequently evaporated on the pregrown InAsNWs. Although desirable, in all our trials we have found only negligible(EuIn)As axial growth of NWs, which was no more than 100 nm long.Instead, (EuIn)As forms a shell around the pregrown NWs.We start with a naive approach in which weattempted to grow a(EuIn)As shell over an InAs core NW. For this InAs core NWs were grownby molecular beam epitaxy (MBE) using the well-established gold-assistedvapor–liquid–solid (VLS) techniqueFigure SI(1)). We note that reducingthe (EuIn)As thickness was not sufficient for improving the surfacemorphology of the (EuIn)As shell grown on a WZ core. In Theresulting InAs/(EuIn)As core/shell NWs are presented in Figure SI(2)). The domain boundaries form a distinctivepattern in the TEM image with a network of triangular shapes in projection,which provide streaks along ⟨111⟩ directions in theFT. We find similar nanodomain structures within both the rough shellsthat grow on WZ cores and smooth shells that grow on the ZB stalactites.To resolve the crystalline structure of the (EuIn)Asshells, weimage them in transmission electron microscopy (TEM). An analysisof such (EuIn)As-coated WZ NWs exposes a particularly intriguing crystallinestructure. The unique crystalline structure has the basic periodicityof the ZB lattice. Yet, it is composed of nanodomains separated bya high density of {111} planes. This can be seen clearly in Figures SI(3) and SI(4) relating to the WZ and stalactite NWs, respectively.The WZ and stalactite NWs were carefully studied by energy-dispersiveX-ray spectroscopy (EDS) in order to verify the Eu composition. Eucompositions ranging from a low of few atomic percent reaching toas high as 15 atom % were found. The results for a couple of typicalsamples are given in 27 ZB InAsx1–Sbx NWs (Sb 5–7atom %) were grown on top of WZ InAs reclining stems. Remarkably,the ZB structure of the top InAsx1–Sbx cores indeed enabled the formationof a smooth (EuIn)As shell, as seen in x1–Sbx ZB NWs systematicallyproduced a smooth shell.The regular(EuIn)As shell that grew over the ZB InAs stalactitecore NWs strongly suggested that to obtain a smooth (EuIn)As coatingthe core should have a ZB rather than a WZ structure. To guaranteethe growth of NWs with a ZB core, we have thus added a low flux ofSb during the core growth, which has been demonstrated previouslyto enforce a ZB structure.x1–Sbx ZBcore NWs before and after (EuIn)As coating, as well as the correspondingEDS data, are presented in Figures SI(5) and SI(6), respectively). The bending of such NWs is presumably related tonon-negligible lattice strain between the core and the (EuIn)As shellas well as a long migration length. It likely originates from theasymmetric evaporation of a lattice mismatched material on the recliningNWs thoroughly studied previously.30 Finally, in Due to the reclining of the NWs grown on (001)substrates, half-shellgrowth also induces significant bending, which can be seen in 2As2 crystal.19 This position is available only at one polarity of the (111) surface.InAs NWs have no such surfaces as side facets, in neither the WZ northe ZB structure. Still, seeds of such surfaces can appear in theZB structures in steps on the {211} type facets. Since, for this favorablepolarity, the positions of the Eu atoms and their distance from thearsenic layer are different from those of In atoms in InAs, in orderto avoid defect formation Eu should embed into InAs over areas ofthe {111} surface as large as possible. To check if small {111} stepsactually appear, if they can become larger as the NW grows, and thusunderstand why the shell with Eu ions forms much better on the ZBcore structures than on WZ, the Lammps Molecular Dynamics simulationpackage was used to model Eu incorporation at the side facets of theNW. The initial simulated core NW has a WZ and a ZB section, and twoseparate Eu and As reservoirs are placed at opposite sides of theNW. While the temperature was slowly lowered, attachment of As atomswas first seen, and then Eu atoms stick to the side surfaces. Thesimulation proceeded until a stable core/shell NW structure was formed,as shown in Supporting Information including methods Figures SI(7) and SI(9) as well as the simulation Movie 1). Indeed, in the ZB part of the NW {2̅11}-orientedsurfaces with large {111} terraces appear between the {11̅0}side facets. On such terraces regular layers of Eu can develop ). Thus, the calculations have shownthat the ZB crystal structure enables Eu incorporation and the growthof a regular shell on the NW side facets, with an atomic structuresimilar to that of a EuIn2As2 crystal. In theWZ structure polar surfaces of {111} type appear only in the growthdirection. The WZ structure has neither such side facets nor suchsurfaces lying at an angle to the growth direction. Thus, the atomswhich hit the NWs can only form EuAs droplets on the NW side walls.In InAs the most advantageous positions for Eu ions incorporationare on {111} surfaces, above the triangles formed by As ions, as inthe EuInx1–Sbx cores. EDS chemical maps ofsuch NWs in a side view and in a cross-section are displayed in x1–Sbx core and the Eu-richshell ). Quantificationof the EDS data reveals that Eu is built into the shell with up to12 atom %, at the expense of indium. The elemental composition ofa lamella prepared from a core/shell NW with a shell of (EuIn)As grownon InAsx1–Sbx is shown in Figure SI(12). It shows arather uniform distribution of Eu throughout the shell diameter. Similarconcentrations of Eu were found also in (EuIn)As shells grown overWZ InAs cores, as well as ZB stalactite cores.Next, we resolve the incorporation of Eu within the (EuIn)As shellsand the nanodomain structures that form in them microscopically. TEMimages expose the same ZB structure comprised of a high density ofnanodomains as observed in the (EuIn)As shells grown on a WZ or astalactite InAs core. High-resolution scanning transmission electronmicroscope (HRSTEM) data shown in We now focus on the triangular mosaic structureand the mannerEu incorporates in it. The triangular mosaic pattern is shown in high-angleannular dark-field (HAADF) images in 2O3 compounds32 with WZ symmetry. Amodel of the structure of the Eu inversion planes derived from ourfindings is shown in The atomicallyresolved image in Figure SI(13)). The full 3D structure of a singlepolar domain of the InAs is prismatic; its shape is determined by{111} facets and by the surface facets of the NW, when the domaincuts the surface. A 3D model of such a prismatic domain is depictedin Next, we analyze how the 2D Eu inversion planes forma 3D networkand eventually lead to the characteristic mosaic structure seen inthe TEM images, such as in Figures SI(9), SI(13), and SI(14), where the apex of the triangular projection of all prismatic domainspoints to one common ⟨100⟩ lattice direction and hencethe appearance of the pattern of scales. Since Eu creates the inversionplanes, the density of the prismatic domains is a simple functionof the Eu content, under the constraint of perfect termination ofinverted polarity without coordination conflict.The octahedral coordination of Eu ions with As happens only on{111}B planes terminated by As. Consequently, the orientation of allprismatic domains is aligned relative to the dominant matrix orientationof the NW. This alignment of the prismatic domains is evident fromimages such as in Figure SI(15)). EDS mapping of a lamellacross section, cut and polished from such core/shell NW, shows clearlythe incorporation of the Eu in the shell, which just barely coatsalso the shadowed side of the NW. Occasional InAsx1–Sbx NWs that grow verticalto the substrate have a full shell similarly to the stalactite NWsextending from the intersections downward. As mentioned above, ina few samples slight axial growth (∼100 nm) bearing a homogeneous(EuIn)As composition was observed at the NW tips, as can clearly beseen in Figure SI(16). It is related tothe vertical growth taking place during the shell side growth.Notably, theEu incorporation takes place at the expense of indiumin the lattice, whereas arsenic remains mostly ∼50%, as isevident from a scatter plot of EDS data across various NW samples–SI(19)). Since InAs has no magnetic moment, we conclude that this strongparamagnetic response arises from the (EuIn)As shell. We note thatinterfaces with the InAs nanocrystallites in the As-Eu-As layer network,but also with the InAsx1–Sbx core, may enrich the magnetic phenomenology.Having obtained NWs with regular (EuIn)As shells, we turn to theirmagnetic characterization. We have employed scanning superconductingquantum interference devices (SQUIDs) operated at 4.2 K to map thelocal DC magnetization of the NWs. Our sensors are further equippedwith a local loop that allows us to apply local AC excitation andmap in addition the AC susceptibility. The core/shell (EuIn)As/InAsFigures SI(17) and SI(18)).This signifies that intrinsic magnetic order is established in such(EuIn)As shells which would allow magnetic response without applicationof an external magnetic field. We could not identify the structuralorigin that distinguishes the magnetically ordered subset of (EuIn)As/InAsx1–Sbx NWs.However, their existence suggests that intrinsic magnetism of stoichiometricNWs is within reach.Furthermore, a subset of all imaged NWs has also produced a clearsignal in DC magnetization. We compare those magnetic mappings toSEM images to resolve the origin of the magnetic signals we detect.SEM images of two pairs of such NWs are shown in 2As2 has anAFM order among its As-Eu-As ferromagnetic layers. It may very wellbe that the Eu-As-Eu in our (EuIn)As shells are ordered ferromagneticallyas well. However, the relative order of those layers is highly sensitiveto the interlayer separation, which is varying within the networkstructure realized in our NWs. Their magnetic nature thus remainsambiguous. Nevertheless, the intrinsic paramagnetic response alonewould substantially enhance the magnetic response to an externallyapplied magnetic field, rendering such NWs superior platforms forapplications requiring magnetism.Lack of a DC magnetic signal in the presence ofstrong AC susceptibility,which we find in the majority of the NWs, could signify either a paramagneticstate or an AFM order. Bulk EuIn


We retrospectively evaluated [18F]FDG PET scans of 171 patients with NP  and 48 controls . Mean normalized bilateral [18F]FDG uptake in the SN was calculated and compared between groups with covariance and receiver operating characteristic (ROC) analyses . PD patients were additionally stratified by the expression of the well-established PD-related metabolic pattern . [18F]FDG uptake was significantly lower in NP  and its subgroups  than in controls. ROC analysis for discriminating NP vs. controls revealed an area under the curve of 0.81 and a sensitivity and specificity of 56 and 92%. Moreover, nigral metabolism was below the cut-off in 60% of PD patients without elevated PDRP expression. Glucose metabolism of the SN can distinguish patients with NP from controls with good diagnostic accuracy and can be used as a marker of nigral degeneration. Its evaluation is particularly valuable in PD patients without elevated PDRP expression and may thus help to narrow the diagnostic gap of [18F]FDG PET in neurodegenerative parkinsonism .Parkinson’s disease (PD), multiple system atrophy (MSA), and progressive supranuclear palsy (PSP) are characterized by nigrostriatal degeneration. We used [ Current guidelines account for this by recommending dopaminergic imaging for the clinical work-up of parkinsonian syndromes, but they do not support its use to distinguish between different neurodegenerative parkinsonian syndromes4. The evaluation of cerebral glucose metabolism with [18F]fluorodeoxyglucose (FDG) positron emission tomography (PET) instead allows to differentiate between Parkinson’s disease (PD), multiple system atrophy (MSA), and progressive supranuclear palsy (PSP) with high accuracy5. Patients with PD with cortical involvement , especially those with mild cognitive impairment [PD-MCI] and PD dementia [PDD]), MSA, and PSP can often be easily diagnosed by means of typical (sub-)cortical metabolic patterns. In turn, patients with PD without cortical involvement may present with only subtle metabolic abnormalities like relative striatal hypermetabolism. These may be hardly distinguishable from normality and constitute a clinical challenge6. In those cases, the differential diagnosis with [18F]FDG PET relies on the exclusion of other forms of NP and should be supported by the proof of nigrostriatal degeneration .Nigrostriatal degeneration is the histopathological hallmark of neurodegenerative parkinsonian syndromes (NP) and can be detected by means of presynaptic dopaminergic neuroimaging8. This suggests that nigral glucose metabolism may be a suitable measure for nigral degeneration. The aim of this study was, therefore, to examine the diagnostic accuracy of nigral metabolism measured with [18F]FDG PET in delineating NP from controls . Furthermore, we assessed the diagnostic value of reduced metabolism in the substantia nigra in a subgroup of PD patients who did not show an elevated expression of the well-characterized PD-related metabolic pattern (PDRP)9).Recent findings in PD patients revealed that metabolism in the substantia nigra pars compacta is reduced and correlates with putaminal dopamine synthesis capacityp > 0.1). However, patients with PSP were older than controls (p < 0.001), MSA and PD (both p < 0.01) and prevalence of male patients was lower in MSA compared to PD (p < 0.05). As expected, we observed significant differences in disease duration between groups: PD patients had the longest disease duration, and PSP patients had the shortest disease duration  = 3), for details, see Table There were no group differences in terms of sex and age at PET imaging between patients with NP, PD, or MSA and controls  and control patients (0.86 ± 0.05), that is why we performed all subsequent analyses with one combined control group only. [18F]FDG uptake was significantly different among patient groups and controls . Post hoc analyses indicated that nigral uptake was significantly lower in NP  and its subgroups  compared to controls . For subgroups, ROC analyses showed discrimination between patients and controls with a ROC AUC of 0.80, 0.77, and 0.90 for PD, MSA, and PSP, respectively , the aforementioned cut-off of normalized [18F]FDG uptake of the substantia nigra (0.80) allowed for separation from controls with 60% sensitivity and 92% specificity , low PDRP score , early  and late  PD stages .Elevated expression of the PDRP  was detected in 67 (58%) PD patients . In those PD patients without elevated PDRP expression (n = 111). Forty-five of these patients underwent [123I]FP-CIT SPECT within 3 months of PET imaging. No significant association was found between mean normalized [18F]FDG uptake of the substantia nigra and disease duration or motor scores, whereas [123I]FP-CIT specific binding potential in putamen showed a moderate negative correlation with UPDRS scores .At the time point of PET imaging, UPDRS motor part 3 scores in medication-off condition were available in 97% of PD patients cortical metabolic patterns5, the combined assessment of nigral and cerebral metabolism enables both evaluation of the nigrostriatal system and differential diagnosis of NP. The clinical benefit of confirming pathological DAT availability is negligible in patients with a clear metabolic pattern of PD , MSA , or PSP 5. However, in the subgroup of PD patients without elevated PDRP expression, dopaminergic imaging substantially increases diagnostic confidence. Thus, the present study suggests that evaluating nigral metabolism may supersede dopaminergic imaging in up to 60% of patients without elevated PDRP expression and thereby considerably improve the diagnostic value of [18F]FDG PET in the differential diagnosis of NP. Overall, 95 out of 115 PD patients (83%) showed metabolic abnormalities detectable with [18F]FDG PET  in the present study. Consequently, a diagnostic algorithm that favors [18F]FDG PET over DAT imaging for the initial evaluation of clinically uncertain parkinsonian syndromes may reduce the number of scans, associated costs, and burden to the patient. Further prospective studies are needed to define optimal clinical algorithms, confirm the present findings also at disease onset, and assess the clinical impact of the evaluation of nigral metabolism.DAT SPECT is a well-established imaging standard for identifying NP by means of striatal DAT loss (sensitivity of 78–97% at a specificity >95%)18F]FDG uptake: 0.80) and MSA (0.81), while the reduction was slightly more pronounced in patients with PSP (0.77). This is consistent with neuropathological findings of nigral cell loss in these three diagnostic entities15. However, the variance of nigral metabolism in each group and the overlap with the control cohort is noticeable and might be partially related to the clinical features of included subjects. Although we did not observe an association between nigral metabolism and disease duration or motor scores in the PD group, heterogeneity in terms of disease stages within each group may cause variable severity of brainstem involvement16. Furthermore, disease variants, like MSA with predominant parkinsonism or MSA with cerebellar ataxia and PSP Richardson syndrome, PSP with predominant parkinsonism or PSP with frontal presentation exhibit varying degrees of brainstem pathology and nigral cell loss17. This may, to a certain degree, explain differences in the level of glucose metabolism observed in our PSP cohort, in which PSP with predominant frontal presentation (n = 3) and corticobasal syndrome (n = 3) showed a less pronounced decrease compared to PSP Richardson syndrome (data not shown). However, due to the small sample sizes of the subgroups, we cannot draw firm conclusions and more data are needed, particularly of patients early during the disease course, to elucidate when metabolic changes in the substantia nigra start to occur.Reduced metabolism of the substantia nigra was observed in each subgroup of NP with comparable reductions in patients with PD  and the pons (for PD and controls) as alternative reference regions, as well as the evaluation of side-separated nigral metabolism, did not overcome this issue -based stereotactical normalization in those patients and controls with available T1 MPRAGE sequence supports the validity of the current PET-based approach .In summary, the present study shows in a large and clinically well-characterized cohort that glucose metabolism of the substantia nigra can distinguish patients with NP from controls with good diagnostic accuracy and can be used as a marker of nigral degeneration. Its evaluation is particularly valuable in PD patients without elevated PDRP expression and may thus help to narrow the diagnostic gap of [18F]FDG PET on a fully digital PET/CT system for the differential diagnosis of NP. Two movement disorder specialists  made a consensus diagnosis based on all available anamnestic, clinical, and diagnostic information. One hundred seventy one patients met the criteria of probable PD (n = 115), MSA (n = 35), or PSP (n = 21) and were therefore enrolled in this study4. We excluded patients with corticobasal degeneration because of the small sample size (n = 5). The consensus diagnosis was not blinded to the clinical reports of PET imaging  and integrated the results of [123I]FP-CIT SPECT in 75 patients . The latter consistently showed a pathologically reduced dopamine transporter (DAT) availability in all patients. Thirteen HC subjects  recruited by local advertisement combined with 35 control patients  derived from a prior study28 served as the control cohort. HC were healthy according to medical history (no neurologic or psychiatric condition or any other relevant comorbidity) and unimpaired on neuropsychological evaluation, had no neurological deficit on clinical examination and had normal MRI findings of the brain. For demographics and patient characteristics, see Table We retrospectively screened the records of the Department of Nuclear Medicine, Medical Center—the University of Freiburg (from January 2018 until April 2021) for patients who had undergone brain [18F]FDG for 10 min on a fully digital PET/CT system . Fully-corrected emission datasets were reconstructed with low-dose CT-based attenuation correction and the vendor-specific, line-of-response time-of-flight ordered subsets 3D iterative reconstruction algorithm employing spherically symmetric basis functions . This yielded a reconstructed, isotropic image resolution of approximately 4.5–5 mm full width at half maximum. Of note, we did not employ resolution recovery to avoid Gibb's artifacts.PET emission data were acquired 50 min after intravenous injection of 209 ± 12 MBq FDG PET scans for those individuals who had an available T1 MPRAGE MRI sequence . For this, individual MRI scans were segmented and stereotactically normalized to MNI space using CAT12 software in SPM12. [18F]FDG PET scans were co-registered to MRI and then transformed into MNI space by applying an MRI-defined transformation matrix. Mean [18F]FDG uptake in SN from MRI-based stereotactical normalized data was correlated to the uptake derived from PET-based stereotactical normalized data by using Spearman’s r. Moreover, between-group differences were assessed. Absolute mean counts in the substantia nigra from PET- and MRI-based stereotactically normalized scans showed a strong correlation . Normalized mean FDG uptake in the substantia nigra derived from scans stereotactically normalized using MRI was significantly different among diagnostic groups and controls, supporting the results based on PET-based stereotactical normalization  from a brain parenchyma mask  and the pons  as alternative reference regions. We observed significant differences in the mean bilateral [18F]FDG uptake in substantia nigra normalized to individual white matter uptake  confirming the results with the cohort-specific reference region.To assess metabolism in the substantia nigra, we normalized the PET data to the uptake within a cohort-specific composite reference region . The cohort-specific reference region was defined by excluding the brainstem but including the substantia nigra, as well as regions with PD-, MSA-, and PSP-specific hypometabolism . Mean [18F]FDG uptake of substantia nigra was significantly reduced in NP groups compared to controls (at least p < 0.01). Minimum [18F]FDG uptake of the both sides, showed similar significant results . For mean values of groups, please refer to Supplemental Table.When assessing FDG uptake of the substantia nigra  were assessed using age-adjusted analysis of covariance (ANCOVA) followed by Tukey’s honest significance test. We estimated the effect sizes for pairwise comparisons with Cohen’s d. Receiver operating characteristics (ROC) analysis was employed to assess the diagnostic performance of nigral [18F]FDG uptake for differentiation between NP and its subgroups and controls by means of the area under the ROC curve (AUC). Furthermore, we determined cut-off values for mean normalized [18F]FDG uptake of the substantia nigra with a requested minimum specificity of 90%, in line with the clinical need for a confirmatory test . We also performed ROC analyses for PD subgroups with and without elevated expression of the PDRP. Furthermore, we assessed group differences between controls and PD patients with early (Hoehn & Yahr ≤2.5) and late (Hoehn & Yahr >2.5) disease stages and short (<5 years) and long (>5 years) disease duration. For PD patients, the relationship between mean normalized [18F]FDG uptake, [123I]FP-CIT specific binding potential  in the putamen ipsilateral to the clinically most affected side , disease duration and Unified Parkinson’s Disease Rating Scale (UPDRS) motor part 3 (off medication) was assessed by using age-adjusted Pearson’s correlation coefficient. The significance threshold was set to Bonferroni-corrected p < 0.05.Statistical analysis was performed with R software  approved the retrospective analysis.All patients and controls gave written informed consent to [Supplemental material


The human mitochondrial genome is transcribed as long strands of RNA containing multiple genes, which require post-transcriptional cleavage and processing to release functional gene products that play vital roles in cellular energy production. Despite knowledge implicating mitochondrial post-transcriptional processes in pathologies such as cancer, cardiovascular disease and diabetes, very little is known about the way their function varies on a human population level and what drives changes in these processes to ultimately influence disease risk. Here, we develop a method to detect and quantify mitochondrial RNA cleavage events from standard RNA sequencing data and apply this approach to human whole blood data from > 1000 samples across independent cohorts.MRPP3, TBRG4 and FASTKD5), including a potentially novel role for RPS19 in influencing cleavage rates at a site near to the MTATP6-COX3 junction that we validate using shRNA knock down data.We detect 54 putative mitochondrial RNA cleavage sites that not only map to known gene boundaries, short RNA ends and RNA modification sites, but also occur at internal gene positions, suggesting novel mitochondrial RNA cleavage junctions. Inferred RNA cleavage rates correlate with mitochondrial-encoded gene expression across individuals, suggesting an impact on downstream processes. Furthermore, by comparing inferred cleavage rates to nuclear genetic variation and gene expression, we implicate multiple genes in modulating mitochondrial RNA cleavage (e.g. We identify novel cleavage junctions associated with mitochondrial RNA processing, as well as genes newly implicated in these processes, and detect the potential impact of variation in cleavage rates on downstream phenotypes and disease processes. These results highlight the complexity of the mitochondrial transcriptome and point to novel mechanisms through which nuclear-encoded genes can potentially influence key mitochondrial processes.The online version contains supplementary material available at 10.1186/s12915-022-01373-5. In humans, mitochondria play important roles in many fundamental and interconnected cellular processes, such as thermogenesis, cellular energy production and cell death , and mitMitochondria are unique organelles that contain their own independent genome, a remnant of their ancestral bacterial origin . The humThe human mitochondrial genome itself is transcribed as polycistronic RNA containing multiple genes, which is then processed under the ‘punctuation model’ whereby tRNAs that intersperse coding and ribosomal sequences are targeted and cleaved by nuclear-encoded proteins to release gene products . CanonicMTATP6 and MTCO3), and thus, other proteins and mechanisms are needed to cleave RNA. For example, FASTKD family proteins have been associated with RNA processing at some gene boundaries   using a stringency parameter value of 3  annotated boundary for ribosomal and messenger RNA mitochondrial genes. Within each region, we then identified the position with the highest cleavage ratio as representative of the gene boundary and then obtained the distribution of cleavage ratios for this site across all individuals. For the background rate, we randomly selected 100 sites from locations at least 50bp away from a gene boundary region and followed the procedure as above for each site. We used one-sided https://github.com/nanopore-wgs-consortium/NA12878). Within this study, data was aligned with minimap2 to the GRCh38 human genome reference, which contains the exact same mitochondrial sequence as the reference used here (1000G GRCh37 reference) and data was merged across all sequencing runs to create a single alignment file for each of the native RNA and cDNA data. For each alignment file, we extracted sequencing reads that mapped to the mitochondrial genome, were labelled as the primary alignment and had mapping quality greater than 30, and then removed reads that had segments that aligned elsewhere in the mitochondrial genome and those that aligned elsewhere in the nuclear genome with equal or greater mapping quality score. For the remaining data, we calculated the start and end positions of each read using the CIGAR string (which contains information on sequence matches and insertions/deletions for each read versus the reference sequence). To test for an overlap with putative RNA cleavage sites identified in short read data, we identified start and end positions in long read data that had at least 200 supporting reads and were in the top 1/50 of sites across the mitochondrial genome in terms of the number of reads that started or ended at that position. For validation, we required these positions to be within 3bp of the putative RNA cleavage site.To validate mitochondrial RNA cleavage sites using data generated with other library preparation and/or sequencing techniques, we first obtained an additional 16 whole blood RNA sequencing datasets from healthy controls, generated after ribosomal RNA depletion (rather than polyA enrichment) and sequenced on the Illumina Hiseq 4000 platform  (GEO acchttps://qtltools.github.io/qtltools/) was used to check for consistent sample labelling between genotype and RNAseq data. Quality control was carried out using PLINK [v1.90b3.44]  [QTLtools [v1.0]  with a minimum calling threshold of 0.9 and then filtered for having an IMPUTE2 info score <0.8, genotype missingness > 5%, MAF < 5%, HWE P < 0.001 or for being multi-allelic. GTEx data, which was genotyped on two different arrays, were imputed separately and then merged.Phased chromosomes were imputed using IMPUTE2 [v2.3.2] , 63 usinQuantitative trait loci mapping was carried out for the 54 reproducible cleavage sites identified in both CARTaGENE and GTEx datasets. Analyses were carried out separately for each position (therefore comparing samples that were generated using the same library preparation and sequencing protocols), using linear models in PLINK [v1.9]. Covariates used in the linear model included 5 study-specific genetic PCs and 10 PEER factors calculated from RNAseq data using PEER [v1.0] . PEER faP-value was adjusted for Bonferroni correction using the p.adjust function and the two lists of significant associations we intersected , identifying 52 significant associations . Since the discovery dataset identified a significantly larger number of associations  when compared to the replication dataset (465), we checked the influence of sample size on these results by randomly resampling the discovery dataset without replacement down to the same size as the replication set (n = 344) and repeating the analysis as above. In doing so, we find 2259 significant associations (after Bonferroni correction), which more closely matches the number of associations found in the replication dataset. The remaining differences may be driven by random variation between the samples or systematic differences that may include RNA degradation levels, read length or sequencing depth.Mitochondrial and nuclear gene expression were generated as in . The lm P-value as a representative cis-eQTL and only used associations that were significant after correction for multiple tests (FDR 5%). For nuclear genes/genetic variants that pass these criteria, we tested whether the expression of the nuclear gene significantly mediated the relationship between the peak nuclear variant and associated inferred mitochondrial RNA cleavage ratios using 1000 bootstrapping simulations with the ‘Mediation’ package in R, correcting the P-value for the number of genes tested using Bonferroni correction. To test whether nuclear genes might be influencing mitochondrial-encoded gene expression levels through mitochondrial RNA cleavage, we obtained all mitochondrial RNA cleavage sites that were significantly associated with the expression levels of both a nuclear- and mitochondrial-encoded gene  and then performed mediation analysis by first testing whether the nuclear- and mitochondrial-encoded genes were correlated in discovery data (P < 0.05) and then second whether this relationship is significantly mediated by the inferred cleavage rate of the associated site in discovery data .To identify the direction of effect between associated nuclear gene expression values and mitochondrial RNA cleavage rates, we carried out mediation analysis. First, we used PLINK to identify cis-eQTLs within 1MB of the start and end for each of the genes identified as significant in the comparison of nuclear gene expression and mitochondrial RNA cleavage rates using the CARTaGENE dataset, including 5 study-specific genetic PCs and 10 PEER factors as covariates. We then selected the SNP with the lowest TBRG4 and RPS19) had RNA sequencing data available after shRNA knock down (KD) as part of the ENCODE project. For each gene, there were 8 RNA sequencing datasets available in total (4 from KD and 4 from controls) in 2 different cell lines (HepG2 and K562). We obtained raw RNA sequencing datasets for each sample via the ENCODE portal  data for any gene that has been implicated in mitochondrial RNA processing in our analyses, including those linked through quantitative trait loci mapping (four unique genes) and expression correlation analyses  down experiments in the ENCODE portal.


The admixture of domestic pig into wild boar populations is controlled until now, by cytogenetic analysis. Even if a first-generation hybrid animal is discernable because of its 37-chromosome karyotype, the cytogenetic method is not applicable in the case of advanced intercrosses. The aim of this study is therefore to evaluate the use of SNP (Single Nucleotide Polymorphism) markers as an alternative technology to characterize recent or past hybridization between the two sub-species. The final goal would be to develop a molecular diagnostic tool.The Geneseek Genomic Profiler High-Density porcine beadchip , comprising 68,516 porcine SNPs, was used on a set of 362 wild boars with diverse chromosomal statuses collected from different areas and breeding environments in France. We generated approximately 62,192–64,046 genotypes per wild boar. The present dataset might be useful for the community (i) for developing molecular tools to evaluate the admixture of domestic pig into wild boar populations, and (ii) for genetic diversity studies including wild boar species or phylogeny analyses of Suidae populations.Raw data files and a processed matrix data file were deposited in the ArrayExpress at European Molecular Biology Laboratory-European Bioinformatics Institute (EMBL-EBI) data portal under accession number E-MTAB-10591. Different studies have reported that wild boar populations have significantly increased in France, like in other parts of the world . Among tThe data set is still confidential but a scientific publication is in progress and initial analyses have already been presented in an oral communication . In the The present data set might be useful for genetic diversity studies to improve wild species preservation measures in our country. In addition, these all-genome genotyping data from the French wild boar populations could be useful for phylogeny studies of Suidae populations.The experiment included 362 French wild boars from different French regions and environments. Most of the animals (n = 252) were bred in enclosed wild boar farms spread out over 38 French administrative subdivisions. The other animals were bred in free-range conditions, either in a protected area , or in an unprotected area . There were 223 females and 139 males. According to the cytogenetic analysis, there were 203 animals with chromosome numbers equal to 2n = 36, 70 animals with 2n = 37, 10 animals with 2n = 38, and 79 animals with unknown chromosomal status. No animals in our study were bred/killed/taken specifically for the needs of our project, which therefore did not require explicit authorization (in accordance with the 2010/63/EU European Directive).A total of 362 biological samples were collected. The biological samples of animals from free ranging “Ardèche” were ear biopsies n = 79) collected between the years 2014–2016 for an edema disease project [9 collectDNA samples were extracted for the 362 biological samples. Genomic DNA of 283 heparin blood samples were extracted using the Blood DNA Isolation (Norgen) kit. Genomic DNA of 79 ear biopsies were extracted with an in-house protocol . Total genomic DNA quality was determined using the Nanodrop 8000 spectrophotometer . Total genomic DNA concentration was determined using the Quant-iT picogreen dsDNA Assay Broad Range kit  with the QuantStudio6 instrument at the Genomic and Transcriptomic (GeT) platform [A total of 362 genomic DNA samples were genotyped in collaboration with the Cancer Research Center of Toulouse (CRCT) core facility platform . GenotypGenotypes were inferred from the raw fluorescence intensity data using the Genotyping Analysis Module included in GenomeStudio (version 2.0.5) software . We used a custom cluster from the commercial cluster file and Illumina guidelines . The calThe call rate of wild boar samples is approximately 0.91–0.93. Quality of wild boar genotyping using porcine array depends on the quality of heterologous hybridization.


To date, knowledge is limited regarding time-dependent suicide risk in the years following return from deployment and whether such rates vary by military rank  or component . To address these gaps in knowledge, the objectives of this study were to determine and compare postdeployment suicide rates and trends (percent change over time), and hazard rates for Army soldiers, by rank and component (measured at the end of the deployment).Longitudinal cohort study of 860,930 Army soldiers returning from Afghanistan/Iraq deployment in fiscal years 2008–2014 from the Substance Use and Psychological Injury Combat study. Death by suicide was observed from the end of the first deployment in the study period through 2018  for up to 11 years of follow-up. Analyses were conducted in 2021–2022.Adjusting for age, lowest-ranking Junior Enlisted (E1–E4) soldiers had a suicide rate 1.58 times higher than Senior Enlisted (E5–E9)/Warrant Officers  and 2.41 times higher than Officers . Suicide rates among lower-ranking enlisted soldiers remained elevated for 11 years postdeployment. Overall and annual postdeployment suicide rates did not differ significantly across components. Comparisons across rank and component for females were generally consistent with the full cohort results.Lower-ranking enlisted soldiers had the highest rate of suicide, underscoring the importance of understanding rank as it relates to social determinants of health. For over a decade following Afghanistan/Iraq deployment, lower-enlisted rank during deployment was associated with an elevated rate of suicide; thereby suggesting that postdeployment prevention interventions targeting lower-ranking military members are warranted.The online version contains supplementary material available at 10.1186/s40621-022-00410-9. Deployment is an experience unique to military members that may increase suicide risk  suicide surveillance and studies during the Afghanistan/Iraq conflicts, lowest-ranking Junior Enlisted (E1–E4) members were at highest suicide risk compared to Senior Enlisted or Officers  2008–2014 (the first deployment in this period was defined as the index deployment). Study objectives were to: (1) estimate average annual and time-dependent suicide rates and trends by rank and component; (2) estimate the relative hazard for suicide and compare suicide rates and trends across rank and component; (3) estimate and compare suicide rates and estimate the relative hazard for suicide between those returning from a first deployment versus those with multiple deployments, by rank and component; and (4) determine if results were different among female soldiers compared to the full cohort. For all objectives, rank and component were measured at the end of the index deployment. Knowledge generated may provide novel information about time-dependent suicide risk during the postdeployment window specific to rank and component, with the goal of preventing military and Veteran suicide.Study data were drawn from the Substance Use and Psychological Injury Combat study (SUPIC), an observational, population-based, longitudinal cohort study of all Army soldiers returning from an Afghanistan/Iraq deployment which ended in FYs 2008–2014 , soldiers were excluded if they did not have a usable social security number , required to search the NDI through the MDR. An additional 1123 soldiers were excluded because their NDI-documented death date was before the end of their index deployment. After these exclusions were applied, inclusion criteria for being in the analytic cohort included: (1) having military component data at the end of index deployment; (2) having an index deployment lasting ≤ 5 years; and (3) ensuring SSN and demographic data consistency within and between the SUPIC finder file, Veterans Health Administration medical record data, and the MDR . The application of these exclusion and inclusion criteria resulted in a final analytic cohort of 860,930  codes X60-X84 and Y87.0 as the underlying cause of death. Independent variables of interest included rank , and component . Deployment group was defined as first deployers  or 2 + deployers . Covariates included age , gender, race/ethnicity , and FY return from index deployment .F distribution (Fay Demographic and military characteristics were summarized for: (1) the overall analytic cohort; (2) by component; (3) by deployment group; and (4) by component within deployment group, using frequencies. The follow-up period for death by suicide was from October 1, 2007 – December 31, 2018 (range 0–11 years postdeployment). Crude and age-adjusted average annual suicide rates (using the direct standardization method) were calculated per 100,000 person-years over the follow-up period within rank groups, deployment groups and by component. Age-adjusted rates were standardized based on the 2000 US population Klein . Rate ration Fay .Suicide rates since return from index deployment were calculated for 1- or 2-year intervals (depending on cell sizes) and presented per 100,000 person years—within rank group and component. Annual percent change (APC) in suicide rates was estimated using trend analysis (joinpoint regression) , R v4.1.1 (R Core Team Most of the cohort was aged 18–29 at the end of their index deployment 62.4%), male (89%) and White non-Hispanic  and 2.41 times that of Officers . Senior Enlisted/Warrant Officers had an age-adjusted rate 1.53 times that of the Officers . When stratifying by deployment group, findings remained largely consistent with regards to both effect size and statistical significance , and Senior Enlisted/Warrant Officers had a higher hazard compared to Officers . The overall pattern revealed a larger magnitude of increased risk for Junior Enlisted and Senior Enlisted/Warrant Officers among first deployers than 2 + deployers.Suicide rates and unadjusted and adjusted hazard for suicide did not differ significantly across components  rank and (2) component for females were similar to those for the full cohort. Females comprise 11% of the SUPIC cohort and had a significantly lower age-adjusted suicide rate than their male counterparts . Females were 62% less likely to die by suicide than male soldiers . Given the combination of a lower overall sample size and a lower suicide rate, the number of events was small and limited the analyses that could be conducted in the female sample. As such, only the unadjusted and adjusted Cox proportional hazard models were run. Overall, estimates were less precise for the female models than those for the full cohort and mostly nonsignificant. However, the estimated hazard ratios were consistent in direction of association  between the female and full cohort models with the exception of the adjusted comparisons between Junior Enlisted and Senior Enlisted/Warrant Officers and the unadjusted comparison between Junior Enlisted and Senior Enlisted/Warrant Officers for 2 + deployers   due to small samples sizes, particularly in the female and 2 + deployer models. We were unable to adjust for or make additional comparisons by race and ethnicity in this paper, due to sample size limitations when examining the associations between rank and component and suicide risk. Additional analyses are underway with this cohort to examine the association between race and ethnicity and suicide risk following index deployment, though these analyses do not evaluate rank and component differences simultaneously due to same sample size limitations. Our study included soldiers returning from Afghanistan/Iraq deployments during FYs 2008–2014, and these findings may not generalize to other branches of service or other years of the Afghanistan/Iraq conflicts. While military rank/paygrade is a good proxy for the income of military members at the time of return from their index deployment, we did not have additional information pertinent to the socioeconomic status of soldiers in this study . We did not have access to Military Occupational Status information for the SUPIC cohort , which may be associated with different military exposures during deployment and has been shown to be associated with suicide risk  were at highest risk for suicide in the years following return from deployment, highlighting the importance of understanding rank as a potential social determinant of health which may increase postdeployment suicide risk. As lower-rank was associated with elevated risk for suicide for many years following deployment, prevention interventions targeting lower-ranking military members/Veterans following deployment may be required for decades to come to reduce the likelihood of this trend continuing. Similar to all enlisted members, Junior enlisted members are disproportionately recruited to serve in the military from communities with higher unemployment  and for those whose index deployment was not their first (2+ deployers), within the overall cohort and within each military component.Additional file 4: Female Sample Characteristics. Table of demographic and military characteristics only among female military members, for those whose index deployment was their first (first deployers) and for those whose index deployment was not their first (2+ deployers), within the overall cohort and within each military component.Additional file 5: Component Average Annual Suicide Rates per 100,000 Person Years . Table of crude and age-adjusted suicide rates, as well as rate ratios, by military component for the overall cohort and within deployment status (first deployers and 2+ deployers).Additional file 6: Hazard Ratios from Unadjusted and Adjusted Cox Proportional Hazards Models across Military Component. Table of hazard ratios from six Cox proportional hazards models: three unadjusted models, and three models adjusting for demographics, comparing military component within 1) the full cohort, 2) first deployers and 3) 2+ deployers.Additional file 7: Component Average Annual Suicide Rates with 95% CIs. Average annual suicide rates post index deployment per 100,000 person years by military component (years 8-11 combined). Trend analysis annual percent change over time is also presented.Additional file 8: Average Annual Suicide Rates per 100,000 Person Years with Trend Lines by Component. Figure with average annual suicide rates over time post index deployment  by military component with trend lines overlaid.Additional file 9: Hazard Ratios from Cox Proportional Hazards Models across Rank Groups within the Female Cohort. Table of hazard ratios from six Cox proportional hazards models among female military members: three unadjusted models, and three models adjusting for demographics, comparing rank within 1) the full cohort, 2) first deployers and 3) 2+ deployers.Additional file 10: Hazard Ratios from Cox Proportional Hazards Models by Military Component within the Female Cohort. Table of hazard ratios from six Cox proportional hazards models among female military members: three unadjusted models, and three models adjusting for demographics, comparing military component within 1) the full cohort, 2) first deployers and 3) 2+ deployers.


For the period of predominance of SARS-CoV-2 Omicron variant in Slovenia, February to March 2022, we estimated mRNA vaccine effectiveness (VE) against severe acute respiratory infection (SARI) COVID-19 using surveillance data. In the most vulnerable age group comprising individuals aged 65 years and more, VE against SARI COVID-19 was 95% (95% CI: 95–96%) for those vaccinated with three doses, in comparison to 82% (95% CI: 79–84%) for those vaccinated with two doses. Such levels of protection were maintained for at least 6 months. We estimated vaccine effectiveness (VE) of coronavirus disease COVID-19) vaccines used in Slovenia against hospitalisation due to severe acute respiratory infection (SARI) COVID-19 for the period with Delta variant predominance in October 2021 [9 vaccineA SARI case was defined as any case of acute respiratory infection of such severity that it resulted in hospital admission . A SARI Supplement to this manuscript. By using unique identifiers, we linked the data from the three databases to ascertain the vaccination status of SARI COVID-19 cases admitted to hospitals and previous diagnosis of COVID-19.Three different sources were used to obtain the data for the analysis: weekly numbers of SARI COVID-19 cases admitted to all hospitals in Slovenia were extracted from EPISARI database , vaccinaTo estimate VE against hospitalisation due to SARI COVID-19 for the second and third dose of mRNA vaccines by age groups and by time since vaccination, we used (i) the respective rates of SARI COVID-19 cases among individuals vaccinated with the second or third dose who had not had a previous diagnosis of COVID-19 and (ii) the respective rates of SARI COVID-19 cases among unvaccinated individuals without previous diagnosis of COVID-19.VE was calculated using the formula: (1 − RR) × 100, where RR was the ratio of the SARI COVID-19 cases rate among those vaccinated with second/third dose, to the SARI COVID-19 cases rate among those unvaccinated.To obtain estimates of weekly denominators (for each Monday during 8 weeks under observation) we counted individuals vaccinated with a second or third dose and without previous diagnosis of COVID-19. To obtain estimates of weekly denominators for unvaccinated individuals without previous diagnosis of COVID-19, we subtracted the number of individuals who have received at least one dose of any vaccine against COVID-19, and the number of individuals with previous COVID-19 diagnosis by each Monday for all weeks under observation, from the total number of individuals in the Central Population Registry on 1 January 2021. Then average numbers of respective weekly estimates for all these denominators were used to estimate VE for the 8-week period under observation.During February and March weeks 5–12) 2022, the period of Omicron predominance [ 2022, thFor individuals without previously confirmed COVID-19, the Both older age groups (aged 50 to 64 years and 65 years and more) were better protected against hospitalisation due to SARI COVID-19, if they had already received three dose of mRNA vaccine in comparison to those who had received only two doses of mRNA vaccines. In the more vulnerable age group, aged 65 years and more, VE against SARI COVID-19 was 95% (95% CI: 95–96%) for those vaccinated with three doses in comparison to 82% (95% CI: 79–84%) for those vaccinated with two doses. For both these groups (two doses and three doses received), such level of VE was maintained for at least 6 months after the last dose.The emergence of SARS CoV-2 Omicron variant resulted in the fifth wave of the COVID-19 pandemic in Slovenia and by the end of January 2022 the predominance of Omicron was almost 100% [The risk of severe outcome following SARS-CoV-2 infection is substantially lower for Omicron variant vs Delta variant . Recent Our results also showed high effectiveness of two doses of mRNA vaccines and very high effectiveness of three doses against SARI COVID-19 hospitalisations across all age groups studied. To receive the third dose of mRNA vaccine is especially important for elderly people, aged 65 years or more, who are at the highest risk for serious disease that may result in hospitalisation. In this group, VE against SARI COVID-19 was 82% (95% CI: 79–84%), if vaccinated with two doses of mRNA vaccine, and 95% (95% CI: 95–96%) with three doses. It should be noted that for both groups, those who received two and those who received three doses, such high VE lasted for at least 6 months after the last dose received. This was unlike the SARI COVID-19 waning immunity we observed during the period of Delta predominance, in a previous VE study using the same methodology and data sources .In contrast to our results, waning of protection against Omicron-related COVID-19 hospitalisation with time since vaccination with either two or three doses was reported by some ,11,12, wSupplement to this manuscript. It was important to exclude prior COVID-19 confirmed cases. The analysis including prior COVID-19 confirmed cases underestimated VE (data not shown). We are aware that we could not exclude all prior COVID-19 cases, as some were not diagnosed. Asymptomatic individuals and those with mild symptoms might not have sought testing and all individuals with a positive self-test result for SARS-CoV-2 infection may have not asked for confirmatory testing although it was strongly recommended.While quantified validations of the three databases are not available, they provide real-world national surveillance data collected from all Slovenian hospitals (EPISARI), and data from official national registries (eRCO and COVID-19 databases) ,4. The tSurveillance of SARI COVID-19 cases within EPISARI has some limitations, which have been described in detail previously . HoweverOur results show that during the period of Omicron predominance, vaccination with two doses of mRNA vaccines offers good protection against hospitalisation due to SARI COVID-19, which is significantly enhanced by a third dose in both older (aged 50 to 64 years and 65 years and more) age groups. The third dose is especially important for protection of the most vulnerable group, individuals aged 65 years or more. It is relevant for national vaccination policy planning that in this age group such solid protection seems not to decline for at least 6 months after the last dose.


Morpho butterfly species into different vertical strata of the Amazonian forest affects the performance of upward escape flight manoeuvres. Using stereoscopic high-speed videography, we compared the climbing flight kinematics of seven Morpho species living either in the forest canopy or in the understory. We show that butterflies from canopy species display strikingly higher climbing speed and steeper ascent angle compared with understory species. Although climbing speed increased with wing speed and angle of attack, the higher climb angle observed in canopy species was best explained by their higher body pitch angle, resulting in more upward-directed aerodynamic thrust forces. Climb angle also scales positively with weight-normalized wing area, and this weight-normalized wing area was higher in canopy species. This shows that a combined divergence in flight behaviour and morphology contributes to the evolution of increased climbing flight abilities in canopy species.Habitat specialization can influence the evolution of animal movement in promoting divergent locomotor abilities adapted to contrasting environmental conditions, differences in vegetation clutter or predatory communities. While the effect of habitat on the evolution of locomotion and particularly escape performance has been well investigated in terrestrial animals, it remains understudied in flying animals. Here, we investigated whether specialization of  Summary: Quantification of climbing flight kinematics among closely related butterfly species living in different strata reveals contrasted climbing flight ability, probably resulting from divergent flight behaviour and morphology. The evolution of animal locomotion is mainly driven by selection on the ability to perform fitness-related tasks such as escaping from predators, finding mates or defending a territory . HabitatSturnus vulgaris, individuals foraging in environments with high vegetation escape predators by flying close to the ground, whereas those foraging in open grass swards show steep and upward escape trajectories , resulting in contrasted performance metrics . However, morphology also crucially determines the achievable flight performance range . MorpholWe specifically focused on upward-directed flight, a power-demanding flight manoeuvre crucial for evading predators . Using sMorpho butterflies in total, including 10 individuals of two canopy species and 16 individuals of five understory species  or near the village of San Antonio de Cumbasa . We studied 26  species A. All buThe climbing flight experiments were performed in a large outdoor insectary within a few hours of capture. The experimental setup consisted of an outdoor insectary (9×4×2.5 m) containing a 1 m long funnel-shaped tunnel, built from dark mesh B. Each b−1, spatial resolution of 848×480 pixels) close to the tunnel exit, allowing the upward flight manoeuvre to be filmed simultaneously with all three cameras  and 31 flights from the five understory species (cumulating in 77 wingbeats). Per individual, we recorded on average 2.7±1.9 flight sequences (mean±s.d.), including 2.5±1.3 wingbeats per flight sequence; see http://biomech.web.unc.edu/wing-image-analysis/) for subsequent analysis of the effects of morphology on climbing flight performance.After being filmed, each individual was killed and weighed with a resolution of 0.01 g, and its detached wings were photographed in standardized conditions. From the photographs, we measured the wing area using WingImageProcessor in MATLAB , the horizontal, vertical and total distance travelled during the wingbeat , the corresponding wingbeat-average flight speeds , and the mean climb angle during the wingbeat (γclimb=atan(Uver/Uhor)  D.Subsequently, we quantified the climbing flight kinematics per individual as the mean values of all wingbeats measured for that individual. The wingbeat-based data were used to analyse how climbing flight kinematics varies between canopy species and understory species; the individual-based data were used to perform subsequent analyses of the effect of individual morphology on climbing flight performance.Morpho cisseis and Morpho theseus) and two understory species (Morpho achilles and Morpho sulkowskyi), and digitized three wingbeats for each species, each taken from a different individual .We quantified the detailed body and wingbeat kinematics executed for a subset of the recorded climbing flights. Because acquiring these data is labour intensive, we limited this analysis to the climbing flights of two canopy species  of each wing , we extracted the blade-element average value at mid-stroke, where aerodynamic force production is close to maximum (Uwingbeat) as the mean translational wing speed for all blade elements of all four wings throughout the wingbeat. We averaged the wing speed across the four wings because this speed was almost identical between the wings. Finally, based on the body kinematics, we determined the mean body pitch angle as the average angle between the body axis and the horizontal during the wingbeat (βwingbeat)  were measured for each of the four wings independently (left and right forewing and hindwing) and then averaged between the two forewings and the two hindwings, separately C. We est maximum . We quaningbeat) A. These Utotal and climb angle (γclimb). We used these two parameters because combined they define the climb speed  The first (study 1) focused on testing how climbing flight dynamics and performance vary between canopy and understory species. This analysis was based on the body trajectory data of the complete dataset of all recorded flight sequences. (2) The second approach (study 2) focused on how variation in flight and wingbeat kinematics impacts climbing flight performance. This analysis relied on both the whole dataset of flight kinematics and the supplementary dataset where the full body and wingbeat kinematics was tracked.We studied the climbing flight in Analysis of the climbing flight dynamics and performance consisted of two parts (study 1a and 1b). First, we studied how climbing flight performance differed between butterflies from the two microhabitats by analysing the dynamics of the climbing performance metrics flight speed and climb angle (study 1a) . Second,To analyse the dynamics of the climbing performance (study 1a), we first determined the temporal dynamics of flight speed and climb angle throughout the climbing flight manoeuvres of understory and canopy butterflies, separately. For this, we aligned the consecutive wingbeats of all recorded climbing flights, and then determined the average temporal dynamics of flight speed and climb angle for each microhabitat B–E.N=106 wingbeats for the canopy species, and N=77 wingbeats for the understory species. Phylogenetic ANOVA were used to test for differences between microhabitats while controlling for the phylogenetic distance between species (N=2 canopy species and N=5 understory species.To further compare climbing flight performance among microhabitats (study 1b), we tested how the eight flight kinematics parameters differed between understory and canopy butterflies, using a set of ANOVA, and a MANOVA on all parameters combined. Regular ANOVA were used to test the effect of microhabitat while controlling for species, individual, flight sequence and wingbeat number within the flight sequence . The dataset used was composed of  species , based o species A. The daAnalysis of how flight and wingbeat kinematics impact climbing flight performance also consisted of two parts (study 2a and 2b). First, we studied how the wingbeat kinematics varied between the understory and canopy species (study 2a). Second, we studied how variation in either flight or wingbeat kinematics on the one hand and morphology on the other hand affected climbing flight performance, based on an aerodynamic model of a flying butterfly (study 2b).Utotal) and ascent angle (γclimb). A flying animal can increase its climbing flight speed by enhancing the aerodynamic thrust magnitude, and it can maximize the climb angle by directing the aerodynamic thrust vector more upwards , we applied an aerodynamic thrust force model to a climbing butterfly, as described in T/mg is the weight-normalized aerodynamic thrust force produced during a wingbeat. This normalized force varies quadratically with the speed of the beating wing (Uwing), and linearly with air density (ρ), the weight-normalized wing area (S/mg), the wing angle of attack (α) and the angle of attack-specific thrust coefficient (TCα). Note that the weight-normalized wing area equals the inverse of wing loading. Here, we normalized the aerodynamic thrust with weight (mg) because flight accelerations scale with this ratio, and thus climbing speed should increase with this weight-normalized aerodynamic thrust force. This weight equals the product of body mass m and the gravitational acceleration scalar g.To study how Utotal) with the three kinematics and morphology metrics in Eqn 1 .Thus, to test how much kinematics control climbing flight speed, we used a set of linear regressions to correlate the climbing speed (Morpho butterflies control their ascent angle (γclimb) during climbing flight, we took the orientation of the aerodynamic thrust force vector into consideration (wingbeat) and in the body pitch angle (βwingbeat) (To study how deration D. Flyingingbeat) D.Behavioural adjustments in angle of attack will result in different lift and drag forces produced by the wings, causing an adjustment of the aerodynamic force vector relative to the body orientation. In contrast, a change in the body pitch angle reorients the thrust vector relative to the world reference frame, without changing the force vector relative to body orientation. This last mechanism is commonly used by insects as a way to control their forward flight speed  and to pA flying butterfly needs to adjust its body pitch angle via changes in its wingbeat pattern, and so the body pitch angle itself is not directly a flight control variable. Therefore, here we used body pitch angle as a flight kinematics parameter that quantifies the functional mechanism used by butterflies for reorienting of the thrust vector, and not a control variable itself.Morpho butterflies controlled their climb angle using a set of linear regressions, where we correlated the ascent angle (γclimb) with the two relevant kinematics variables, the angle of attack of the beating wing (αwingbeat) and the body pitch angle (βwingbeat). Using the same approach, we also tested how the corresponding morphology metric weight-normalized wing area (S/mg) affected climb angle.Based on this thrust force vector model, we tested how Fig. S1). There was a high variability in most of the measured parameters . During these climbing phases, canopy butterflies flew 18.2% faster than understory butterflies . Noticeably, for the same climb angle of 50 deg, canopy butterflies flew on average 25% faster than understory butterflies . The distribution of climb angles and climbing flight speeds  (Focusing on the climb performance metrics (study 1a) climb angle and climbing speed, we found that butterflies from canopy species tended to climb both steeper and at higher speed as compared with understory butterflies A–D. Our t speeds F shows tP=0.006) F.The temporal dynamics of the climb angle shows that climb angle increased throughout the consecutive wingbeats within each climbing manoeuvre B. While In contrast, the temporal dynamics of flight speed was quite different between canopy and understory butterflies C. For boUver=1.07±0.46 m s−1; understory species: Uver=0.72±0.35 m s−1, P<0.001), whereas the horizontal speed did not differ significantly between understory and canopy species , confirming the greater climbing performance observed in canopy species as compared with understory ones. See also Fig. S3A for a principal component analysis performed on all the climbing parameters. These differences were no longer significant when controlling for phylogenetic distance (Table S1). The effect of selection across microhabitats could thus not be distinguished from the phylogenetic divergence, probably as a result of the low number of species studied, limiting the statistical power.The multivariate statistical analysis applied to all climbing flight parameters (study 1b) shows a significant effect of microhabitat on climbing parameters . For each of these four species, we digitized three wingbeats, each taken from a different individual. Based on these data, we reconstructed the average wingbeat kinematics for the understory butterflies and canopy butterflies separately .Fig. S2). Most apparently, the deviation angle of the hindwings differed between groups: canopy butterflies deviated their hindwings further from the stroke plane than understory butterflies during both the upstroke and the downstroke . Also, the forewings of understory butterflies seem to operate at slightly higher wing stroke amplitudes and wing rotation angles, but these differences were not significant .This detailed study 2a on wingbeat kinematics revealed several differences between canopy and understory species (Morpho butterflies (study 2b), using our aerodynamic model for climbing flight performance (Eqn 1). For this, we quantified the body and wing morphology of all individuals used in the flight datasets  (Morpho cisseis and Morpho theseus) and two understory species (Morpho achilles and Morpho sulkowskyi). The resulting wingbeat kinematics dataset consisted of 12 digitized wingbeats, each from a different individual, and three individuals per species (We consequently studied how variation in wingbeat kinematics and morphology affects climbing performance in species) , and the species  and 3F.S/gm, as a metric of butterfly morphology, because of its high relevance in climbing flight performance (Eqn 1) , whereas butterfly weight did not differ between canopy and understory species (P=0.53).We focused on the weight-normalized wing area  Eqn 1) . This mo . This mFig. S3). This showed similar trends in the two datasets , whereby the canopy species flew faster, climbed steeper, travelled further per wingbeat and had a lower wingbeat frequency than the understory species. All these differences were significant for the complete dataset (N=183 wingbeats), but for the reduced subset (N=12 wingbeats), only climb angle was significantly higher in the canopy species than in the understory species.Our body and wingbeat kinematics analysis was based on a subset of 12 wingbeats from 12 different individuals (highlighted in N=12 wingbeats), we were unable to use the wingbeat kinematics dataset to quantitatively compare the wingbeat kinematics between canopy and understory species (study 2a). Instead, we used the wingbeat kinematics dataset primarily to study the functional effect of wingbeat kinematics on climbing flight performance in Morpho butterflies in general (study 2b). This analysis was based on the aerodynamic thrust force model for flapping flight (Eqn 1), and should allow us to determine how Morpho butterflies adjust their body and wingbeat kinematics to vary their climbing flight performance.Thus, because of its reduced sample size  and forewing (but not hindwing) angle of attack . Flight speed also decreased linearly with weight-normalized wing area (S/gm) , suggesting that canopy butterflies (showing higher S/gm) may have lower flight speed, contradicting the previously found higher climb speed in canopy species. A negative correlation between S/gm and flight speed (S/gm), and understory butterflies tend to have higher horizontal flight speed .Second, we tested how climb angle varied with the functionally relevant kinematics and morphology metrics angle of attack, body pitch angle and weight-normalized wing surface area . This shR²=0.72, F10,9.88=25.55, P<0.001). In addition, the wingbeat-average body pitch angle was a markedly 50% higher in canopy species than in understory species , (Wingbeat-average climb angle was positively correlated with body pitch angle (=0.001), D. The inThese combined results thus show that the higher climb performance observed in canopy species is due to differences in both flight behaviour and morphology, as the higher climb angles exhibited in canopy butterflies are jointly enhanced by their higher body pitch angle and weight-normalized wing area.Morpho butterflies. We showed that in canopy species, climbing speed and ascent angle are on average higher than those of understory species. The dynamics of climbing flight manoeuvres was markedly different between canopy and understory butterflies: while overall velocity was similar in the two groups, canopy butterflies maintained and built on this velocity through subsequent wing strokes, resulting in a fast and steep climbing trajectory. In contrast, butterflies from understory species lost velocity as they climbed, and had trajectories with less steep slopes. This suggests a higher climbing performance in canopy species, which could allow these canopy butterflies to perform better upward escape flight manoeuvres.In this study, we investigated the influence of microhabitat specialization on the evolution of climbing flight performance in Morpho butterflies, and tested how those kinematics affected ascent angle and climbing flight speed. Using an aerodynamic model-based approach (Eqn 1), we showed that flight speed scaled positively with both the speed of the flapping wings and the mean wingbeat angle of attack of the forewings, but not of the hindwings. Wing speed and angle of attack were however comparable among butterflies from canopy and understory species. The higher climbing flight speeds measured in canopy species thus cannot be attributed to a difference in wingbeat kinematics. Wing movements should be measured on a larger sample of butterflies to ascertain the role of wingbeat kinematics in the contrasted climbing performance of canopy and understory species. Interestingly, the positive effect of wing angle of attack on climbing speed was found for the forewings only, supporting a prominent role of forewing movement during butterfly flight  from those of performance capacity .Morpho species may thus result from and/or be enabled by divergent morphology.Selective pressures promoting specific escape abilities in different habitats may ultimately result in morphological changes, altering the limit imposed by morphology on performance . The conS/gm), which is the inverse of wing loading W/S and a parameter strongly constraining flight ability and particularly manoeuvring capacity  limits forward flight speed but in turn enables the execution of sharper turns  is generally associated with higher horizontal flight speeds required for weight support  impeding the execution of upward escape flight, compared with butterflies mostly flying in the open environment of the canopy. Fast and steep ascending flight, which is among the most efficient aerial escape tactics (Morpho butterflies (Although the difference in climbing performance was striking among microhabitats, the small number of species studied here limits our capacity to disentangle the relative influence of phylogenetic distance and microhabitat selection on the evolution of this trait. Contrasted selective pressures exerted by microhabitats might nevertheless influence the evolution of climbing flight performance in  tactics , may thu tactics . Climbin tactics , which aterflies .Morpho species are known to feed on decaying fruits falling on the forest floor (Morpho butterflies are critical to assess how variation in ecological traits, such as escape tactics or foraging behaviours, may influence the evolution of their flight behaviour and performance.Finally, the evolution of climbing flight ability is also probably influenced by ecological factors other than escape from predators. All st floor . Food acst floor , especiast floor . In contst floor . Further


To date, atypical periprosthetic femoral fractures (APFFs) are not well-understood due to a relatively small number of studies on the topic. Moreover, there is a paucity of guidance regarding the appropriate treatment strategy. We would therefore like to present and analyze a series of five APFF cases (one incomplete APFF and four complete APFFs) that were treated in two university hospitals; a review of the literature is also provided. The results provide additional and useful information regarding the treatment strategy for APFFs. Bisphosphonates (BPs) are one of the most commonly used osteoporotic drugs because of their proven efficacy to reduce the incidence of fragility fractures . HoweverTheoretically, APFFs seem to share some patient characteristics, risk factors, and difficulties in treatment with AFFs and PFFs. However, the treatment for APFFs can be more complex, because they combine the challenging poor healing potential of AFFs and the fixation difficulty due to the presence of femoral stem prosthesis in PFFs. To achieve fracture healing in AFFs, the recommendations require not only using intramedullary nail (IMN), which provides a mechanical advantage and is less disturbed from osteoclast inactivation, but also considering biological augmentations in the selected patient . PlatingThis study aims to (1) report on the patient characteristics, treatment methods, and outcomes of the selected APFF cases; (2) analyze them in terms of potential causes for success or failure; and (3) review the literature related to APFF treatment outcomes and develop an up-to-date treatment strategy.This study was approved by the Institutional Review Board of the Faculty of Medicine, Ramathibodi Hospital, Mahidol University (COA. MURA 2019/760). We retrospectively reviewed the patients with PFFs operated on in 2006-2021 at Ramathibodi Hospital and Phramongkutklao Hospital. Inclusion criteria were the patients who (1) were diagnosed with PFFs around the femoral stem and (2) had at least four of the five major features listed in the ASBMR 2014 revised case definition [A total of five cases, one incomplete and four complete fractures, were included in this study. All cases involved female patients with an average age of 66 years. Four (80%) had underlying autoimmune diseases and received prednisolone and immunosuppressive medications. All fractures occurred at or around the mid or tip of stem (Vancouver B1), and all prostheses had a well-fitted cemented femoral stem. The average duration of bisphosphonates was 10.6 years (range 3-14 years). The mean duration of prodromal symptoms was 3.4 months (range 1-8 months). One incomplete APFF, with a previous unipolar hip prosthesis, was treated with revision hip arthroplasty due to failed conservative treatment. In the four complete APFFs, one was treated with revision long femoral stem with plate augmentation, and the other three were treated with open reduction with internal fixation (ORIF) with plate and screws using additional augmentation techniques. Three of five fractures (one incomplete and two complete) achieved union with a mean union time of six months after the first operation. The residual two fractures were re-operated with fixation and finally achieved fracture union. The data from each case are presented separately and then summarized in Table Case 1A 78-year-old female presented with left thigh pain for three months in February 2016. She underwent left cemented unipolar hemiarthroplasty for 10.3 years due to a femoral neck fracture Figure . Her undCase 2A 48-year-old female presented with left PFF after slipping and falling on the floor in January 2017. Previously, she had a history of bilateral osteonecrosis of the femoral heads (ON) and underwent bilateral cemented total hip arthroplasty 15 years prior Figure . Her undCase 3A 53-year-old female presented with right thigh pain while walking and a fall in January 2010. Previously, she had bilateral ON and underwent bilateral cemented total hip arthroplasty 18 years prior, followed by revision cemented THR with an anti-protrusion cage due to aseptic loosening Figure . Her undCase 4A 77-year-old female presented with left thigh pain after having pre-syncope and falling on the floor in April 2019. Previously, she underwent left cemented bipolar hemiarthroplasty six years prior due to femoral neck fracture and right total hip arthroplasty 11 years prior due to hip osteoarthritis (OA). Her underlying issues were an old cerebrovascular accident with full recovery (old CVA) and SLE for 16 years. Her medications are shown in Table Case 5A 77-year-old female presented with left thigh pain after slipping and falling on the floor in March 2020. Previously, she underwent cemented total hip arthroplasty 10 years prior due to hip OA. Her underlying diseases were dyslipidemia and osteoporosis. Her medications are shown in Table APFFs are fractures characterized by atypical features of AFFs and occur around femoral stem prostheses. Although both APFFs and AFFs would have a theoretically similar pathogenesis, only a few studies have addressed APFF patient characteristics , and preRegarding patient characteristics, the findings from this study were mostly comparable to the previous studies that have larger sample sizes -8 Table. Our datConcerning the treatment of incomplete APFFs, our findings demonstrated the high failure rate of conservative treatment, which is comparable to the results from a systematic review by De Cicco et al. . In the The result of complete APFFs from the present study showed that there was an implant failure rate as high as 50% Cases 2 and 4) requiring revision surgery before achieving bone union. Our findings were comparable with those in previous studies since the prevalence of fracture complications requiring reoperation in APFFs varied between 38% and 57% ,8. In te and 4 reThis study also has some limitations. First, due to the rarity of APFFs and the nature of a retrospective case series, our sample sizes were small, and some risk factors might not be recorded. There was also no comparative group or risk factor analysis. Second, our literature review included only articles from an English-based database and might therefore be inadequate. APFFs are uncommon periprosthetic fractures that share some unique features with AFFs, including patients’ characteristics and fracture-related complications. Early diagnosis and prompt, appropriate treatment are the keys to successful treatment. This study showed that the common patient characteristics in APFFs are being female, using long-term bisphosphonates, and having a Vancouver B1 fracture pattern. Moreover, our results demonstrated a high proportion of patients of younger age having autoimmune disease receiving glucocorticoids and immunosuppressive medications. Therefore, APFF screening should be performed in these high-risk patients, especially on those who had abnormal hip and thigh pain.Regarding the treatment strategy, APFFs show a poor fracture healing potential and require special attention. Incomplete fracture has a poor-to-fair success rate and mostly requires prophylactic fixation with plate and screws or revision arthroplasty in some cases. Complete fractures require optimization of the fixation strategy and using, if possible, local and systemic biological enhancement for improving treatment outcomes.