Microbiological protection textile materials played an important role in the battle against the epidemic. However, the traditional active antimicrobial treatment of textiles suffers from narrow textile applicability, low chemical stability, and poor washability. Here, a high-strength adhesive nanosilver glue was synthesized by introducing nontoxic water-soluble polyurethane glue as a protectant. The as-prepared nanosilver glue could adhere firmly to the fiber surfaces by forming a flexible polymer film and could encapsulate nanosilver inside the glue. The as-prepared nanosilver had a torispherical structure with diameter of ~22 nm, zeta potential of −42.7 mV, and good dispersibility in water, and it could be stored for one year. Further studies indicated that the nanosilver glue had wide applicability to the main fabric species, such as cotton and polyester fabric, surgical mask, latex paint, and wood paint. The antimicrobial cotton and polyester fabrics were prepared by a simple impregnation–padding–baking process. The corresponding antimicrobial activity was positively correlated with nanosilver content. The treated fabrics (500 mg/kg) exhibited ultrahigh washing resistance  and wear resistance , equivalent breathability to untreated fabric, improved mechanical properties, and good flexibility, demonstrating a potential in cleanable and reusable microbiological protection textiles.  Public health and medical services have faced enormous challenges in recent years. Bioprotective fibers and related textile materials played a key role in the battle against the epidemic but also reflected their shortcomings . TraditiHowever, the application of nanosilver in antimicrobial textiles should meet the following requirements, such as high water solubility to achieve functional finishing of textiles by traditional dyeing equipment, wide applicability to mainstream textile fibers, no obvious decrease in the functional treatment on the wearing comfortableness of the fabric, and good oxidation and washing resistance to meet the cleanable and reusable needs. Such properties depend on the physical and chemical properties of the nanosilver colloidal solution, which are mainly determined by their capping molecules . NanosilIn the case of nanosilver functionalized fabrics, the capping molecules should possess good protection ability of nanosilver against chemical reagents and high combining capacity of fabrics to achieve good durability without damaging the wearability of the fabric. In our previous studies, we designed hyperbranched polymer-modified silver nanoparticles that can spontaneously adsorb and adhere to the fiber surface through electrostatic adsorption and hydrogen bonding interactions . However3) was purchased from Shanghai Institute of Fine Chemical Materials . Water-soluble polyurethane  was obtained from Qiancheng Plastic Chemical Materials Co., Ltd. . Artificial sweat was provided by Dongguan ChangFeng Automation Technology Co., Ltd. . Sodium borohydride (NaBH4), nitric acid (HNO3), and ammonia were purchased from Sinopharm . Gram-negative Escherichia coli  and Gram-positive Staphylococcus aureus  were obtained from Shanghai Luwei Technology Co., Ltd. . Nutrient agar and nutrient broth were purchased from Shanghai Zhongke Insect Biotechnology Development Co., Ltd. . Phosphorus-free ECE detergent was purchased from Xiangda Instrument Co., Ltd. .Cotton and polyester fabrics were acquired from Zhangjiagang Nellnano Technology Co., Ltd. . Silver nitrate  was slowly added to the above mixture through stirring until the color of the solution stopped changing. The resultant nanosilver glue was further added with water to achieve 50 mL solution, stored in a brown colored bottle, and kept in a dark place. Ammonia was gradually titrated into 10 mL of 7.87 g/L AgNO3 ammonia complex solution and then reduced by NaBH4 as described above. The mass ratio of polyurethane and silver was set as the optimal value determined by the above study. The resultant nanosilver glue was further added with water to achieve the set volume (50 mL) solution, stored in a brown colored bottle, and kept in a dark place. An aqueous polyurethane solution was mixed with 10–40 mL of 7.87 g/L AgNOThe as-prepared nanosilver glue was applicable to most fiber surfaces. In this study, cotton and polyester fabrics were chosen as the typical models of natural fiber and chemical fiber fabrics, respectively, and the typical coating process of nanosilver glue to ultradurable antimicrobial fabrics was described as follows: Cotton and polyester fabrics were washed with deionized water and ethanol several times to remove the impurities and dried at 60 °C for 1 h. The test fabric was then impregnated with nanosilver glue for 30 min with silver concentration ranging from 10 mg/L to 4000 mg/L and a bath ratio of 1:50. The test fabric was padded with a small laboratory padder  and baked at 60 °C for 60 min and at 120 °C for 10 min to induce the cross-linking reaction.−1 range and resolution of 2 cm−1 and X-ray photoelectron spectroscopy . The X-ray diffraction (XRD) patterns of samples were obtained by using an X-ray diffractometer . The mechanical property test was conducted on an electronic fabric tensile tester , where the fabrics  were stretched at a constant rate of 10 cm/min, and an intelligent fabric crease elasticity meter (YG(B)541E, Wenzhou Darong Textile instrument Co., Ltd., Wenzhou, China) in accordance with the standard GB/T 3819 vertical method. The breathability of fabrics was tested by using a digital fabric breathability tester (YG(B)416E, Yakai Instruments, Cangzhou, China). Bending rigidity was analyzed by using a fabric assurance by simple testing . The silver concentration in aqueous solution was detected through inductively coupled plasma mass spectrometry .The morphological structure and distribution of the nanosilver glue were observed by using a transmission electron microscope  at an acceleration voltage of 120 kV. The surface morphologies and chemical structure of samples were observed by using a field emission scanning electron microscope  and a SEM  equipped with an energy dispersive spectroscopy detector . UltraSTEM200X. The absorbance and concentration of the nanosilver glue were recorded by using an ultraviolet (UV) spectrophotometer . The zeta potential and size distribution of the nanosilver glue were measured by using a Zetasizer . The chemical structure of samples was analyzed through Fourier transform infrared  spectroscopy at the 4000–400 cmE. coli (1–5 × 106 cfu/mL) and S. aureus (1–5 × 106 cfu/mL) was evenly placed on the two agar dishes. Three Oxford cups were placed on each culture dish, and 2 mL of deionized water, polyurethane solution, and nanosilver glue solution were added dropwise. The plates were placed in a biochemical incubator for 24 h at 37 °C. The diameter of the suppression ring was measured and recorded. The absorbance of the bacteria at 546 nm was tested with a UV–vis spectrophotometer to study the antimicrobial kinetics of nanosilver glue against E. coli and S. aureus after 0–24 h of culture. The effect of the silver concentration in glue on antimicrobial activity was studied by testing the absorbance of the bacteria at 546 nm after 6 h of incubation. The antimicrobial activities of nanosilver glue were tested by using the Oxford cup inhibition zone method . The steE. coli (ATCC 25922) and Gram-positive S. aureus (CMCC 26003) were selected as the test strains. The modified oscillation method  was used to test the antimicrobial properties of cotton and polyester fabrics, as described in our previous paper . The surThe time-dependent release of silver ions was measured as follows. An amount of 1 g nanoglue-coated fabric was impregnated with 50 mL artificial sweat, and then, 2 mL immersion liquid was collected at immersion times of 0.5, 1 h, 2 h, 4h, 6 h, 12 h, and 24 h. The concentration of silver ion in the impregnation liquid was measured using inductively coupled plasma mass spectrometry (ICP-MS).The washing fastness of fabrics was measured in accordance with the GB/T 12490-2014 standard test method No.AS1 (China). The fabrics (10 cm × 10 cm) were washed with 150 mL of detergent (4 g of ECE detergent dissolved in 1000 mL deionized water) in a programable washing fastness tester  at 40 °C for 30 min, then rinsed with deionized water, and dried at 60 °C. Laundering durability was evaluated by measuring the difference in the antimicrobial efficiencies of the fabric samples after repeated 50 and 100 times of washing cycles.The wear resistance test of antimicrobial fabrics was conducted by using a Martindale friction testing machine in accordance with the GB/T 21196.2 standard test method (China), and the antimicrobial test of fabric samples was evaluated after 8000 times of standard wearing. 3 and sodium borohydride in water with polyurethane glue as a protective agent  was equal to the standard value of Ag0 (6.0 eV) (The SEM images of nanosilver glue film are illustrated in 101 cm−1 c. The haate bond . For nane groups . This fi(6.0 eV) e, furtheE. coli and S. aureus were chosen as typical model strains to evaluate the antimicrobial activities of nanosilver glue in this experiment. Deionized water and polyurethane were used as controls. E. coli- and S. aureus-inoculated nutrient agar surfaces. No inhibition zone was found around the Oxford cups of deionized water and polyurethane, indicating the nonantimicrobial effects of polyurethane. By contrast, an E. coli and S. aureus inhibition zone with diameters of around 3 and 5 mm was measured around the Oxford cup of nanosilver glue. This finding proved the good antibacterial activity of nanosilver against E. coli and S. aureus. The antibacterial activity was proven to depend on the nanosilver concentration. As shown in E. coli and S. aureus at 546 nm after 6 h of incubation decreased significantly from 1.008 to 0.013 and from 0.877 to 0.027 when the concentration of nanosilver increased from 0.5 mg/L to 6 mg/L. No significant increase was observed in the antibacterial activity above 4 mg/L, implying the minimum inhibitory concentration. E. coli and S. aureus within the measurement time of 24 h. The absorbance of pure E. coli and S. aureus suspensions (controls) increased significantly at 1–3 h contact time and reached a peak at the 20 h contact time. However, the bacterial concentration of E. coli and S. aureus in the nutrient solution containing nanosilver glue (4 mg/L) remained mostly unchanged and even showed a slight decrease, suggesting certain bactericidal activities of nanosilver at an ultralow concentration. Nanosilver was believed to hide in two places. One was encapsulated inside the polyurethane film, and the other was embedded in the interface of fibers and polymer film due to the early physical adsorption. As shown in −1 for the coated cotton fabric can be attributed to the carboxyl and the amide groups of polyurethane  maintained over 99% antimicrobial activities for E. coli and S. aureus after 0, 50, and 100 times of washing, demonstrating the strong adhesion and stability of nanosilver coating. The coated cotton and polyester fabrics (500 mg/kg) worn up to 8000 times showed 99.999% antimicrobial activities for E. coli and S. aureus equal to those of untreated fabrics, suggesting the good wear resistance of nanosilver coating. The strong bonding interaction between nanosilver-embedded polyurethane coating and fabrics was responsible for the good washing resistance and wear resistance. TG test results showed that the thermal stability of both treated fabrics was slightly improved compared with untreated samples  had no obvious influence on the particle size and zeta potential. The as-prepared nanosilver glue showed a UV–vis adsorption peak at 421 nm, a spherical morphology with average particle sizes around 22 nm, a negative surface charge of around −42.7 mV, and a minimum inhibitory concentration of 4 mg/L. The nanosilver glue could form a tough and flexible film with nanosilver content up to 10% by using a simple drying process. The nanosilver in the formed film remained metallic and was endowed with good stretchability, toughness, and adhesion, which could be tailored to various shapes and stretched to ~50% of its original length. FESEM, FTIR, XPS, and TG tests showed that the nanosilver glue successfully adhered to cotton and polyester fiber surfaces without blocking the space between the yarns. Antibacterial tests suggested that 500 mg/kg of nanosilver-glue-coated cotton and polyester fabrics showed 99.999% antibacterial activities against E. coli and S. aureus and maintained over 99% even after 100 times of standard washing. Further studies exhibited that the air permeability remained unchanged for 500 mg/kg of cotton and polyester fabrics, and the tensile strength, tear strength, CRA, and hydrophobicity were improved by increasing the silver content. The bending rigidity of cotton showed a slight increase for cotton and an obvious increase for polyester fabric. However, the coated polyester fabric still maintained good flexibility. Our nanosilver glue showed high adhesion, wide applicability to various materials, and high washing resistance and wear resistance, showing its high potential in microorganism protection textiles.In this study, we successfully fabricated a type of nanosilver glue with high adhesion to various materials, such as fabrics and surgical masks, good compatibility with latex paint and varnish that can be used for coating walls and wood, and excellent antimicrobial properties for


No correlations were found between VOC emissions and ice surface algal counts, but a weak positive correlation  between VOC emission rates from bare ice surfaces and incoming shortwave radiation was found. We propose that this may be due to the stress that high solar irradiance causes in bare ice microbial communities. Acetophenone, benzaldehyde, and phenylmaleic anhydride, all of which have reported antifungal activity, accounted for 51.1 ± 11.7% of emissions from bare ice surfaces, indicating a potential defense strategy against fungal infections. Greenland ice sheet microbial habitats are, hence, potential sources of VOCs that may play a role in supraglacial microbial interactions, as well as local atmospheric chemistry, and merit future research efforts.Volatile organic compounds (VOCs) are emitted by organisms for a range of physiological and ecological reasons. They play an important role in biosphere–atmosphere interactions and contribute to the formation of atmospheric secondary aerosols. The Greenland ice sheet is home to a variety of microbial communities, including highly abundant glacier ice algae, yet nothing is known about the VOCs emitted by glacial communities. For the first time, we present VOC emissions from supraglacial habitats colonized by active microbial communities on the southern Greenland ice sheet during July 2020. Emissions of C5–C30 compounds from bare ice, cryoconite holes, and red snow were collected using a push–pull chamber active sampling system. A total of 92 compounds were detected, yielding mean total VOC emission rates of 3.97 ± 0.70 μg m Volatile organic compounds (VOCs) are emitted by plants and microorganisms  for a vaArctic and subarctic vegetation in non-glaciated terrain has been shown to emit VOCs, and their emission rates are affected by climate warming . In addiAncylonema alaskanum  and left to melt into the ice for approximately half an hour. In the case of cryoconite holes, larger holes were selected so that the chamber bases could be placed within the water column to avoid collection of emissions from the surrounding bare ice surface. The grooves in the chamber bases were filled with water before placing the chamber enclosures over the bases, creating an airtight seal. Chambers were flushed at a flow rate of 1,000 mL min–1 for 10 min using battery-operated pumps connected via Teflon tubing. Incoming air was purified during flow through a PTFE filter and an activated charcoal filter to remove particles and VOCs, as well as by flow through a potassium iodide ozone scrubber (–1 for the duration of the sample collection (between 0.5 and 5.5 h), while the inflow rate was set to 210 mL min–1 to maintain a slight overpressure in the chamber and prevent ambient air from entering the chamber. The chambers were equipped with a fan to ensure a well-mixed headspace. Temperature and relative humidity in the chamber headspace were logged once per minute using a shaded iButton sensor .VOC emissions were collected using a push–pull enclosure setup  with trascrubber . FollowiAfter sample collection, cartridges were sealed with Teflon-coated brass caps and stored refrigerated, when possible, until analysis in Denmark. The chambers were removed, and, for bare ice samples, the top 2 cm of the ice within the chamber base was scraped off with a preconditioned ice axe and collected in sterile Whirl-Pak bags. Once melted, samples were homogenized and 2 mL of each sample was transferred to a 2-mL Eppendorf tube. The samples were fixed with glutaraldehyde  and stored in the dark at 4 °C until counting. Microscope counts of glacier ice algae and snow algae were performed on a Fuchs-Rosenthal hemocytometer  using an inverted light microscope (Olympus CK2). A minimum of 100 cells were counted where possible, but counts of the lowest cell concentrations contained fewer cells.n = 6), 2 (n = 2), and 4 (n = 2) h to account for potential background compounds, as well as compounds that were released continuously over time, using VOC-free air (Parker ChromGas Zero Air Generator) for inflow. The limitations of the sampling setup used in this study, such as the lack of steady state in the beginning of the sampling period and the possibility of compounds adhering to the chamber walls, have previously been discussed by Procedural blank VOC collections were performed in the laboratory in Denmark to account for VOCs released from the sampling materials and analytical systems. The sampling setup  was placed on top of a VOC-free polyethylene terephthalate film (prebaked at 120 °C for 1 h) in a refrigerated chamber (0 °C) that was equipped with adjustable lights  to mimic natural sunlight. Blanks were collected for 1 (–1). During analysis, the column was kept at 40 °C for 3 min, then raised to 210 °C at a rate of 5 °C min–1, and finally to 250 °C at a rate of 20 °C min–1. Duplicate standard solutions  . Sorbent tubes were thermally desorbed  at 250 °C for 10 min, cryo-focused at −5 °C, and injected onto an HP-5 capillary column  with helium as carrier gas  software . A total–1 air. Average concentrations in the procedural blanks were subtracted from those detected in the samples, taking into account the sampling time for compounds that increased over time in the blank measurements. Compounds for which sample concentrations were less than two times higher than concentrations detected in the blanks were excluded from the dataset. After blank corrections, only the compounds with concentrations above zero in at least 50% of the samples for each habitat were included in the final analysis. The VOC concentrations were then divided by the sampling mid-time, multiplied by the total chamber volume, and normalized to the plot surface area to obtain emissions rates in μg m–2 h–1. All analyses were performed in R version 3.6.3 (x + 1) transformed and unit variance scaled prior to principal component analysis (PCA).Emission rates were calculated according to on 3.6.3 . Correlar = −0.15, p = 0.50, n = 23), but correlated positively with the average incoming shortwave radiation at the time of sample collection .Weather conditions during the sampling campaign were highly variable, with bad weather conditions making sample collection impossible on 7 out of 16 possible sampling days. This was because of the risk of water damage to the electronics in the sampling system during periods of heavy rain, or persistent cloud cover causing a lack of charge on the solar panels that supplied the camp, making it impossible to charge the battery-operated pumps. Samples were collected whenever possible between 10 AM and 8.30 PM, covering most of the natural variation in the environmental conditions at the sampling site . The temr = 0.43, p = 0.015, n = 31) with the incoming shortwave radiation averaged over the sample collection period for each sample .Total VOC emission rates from bare ice had a low but statistically significant positive correlation , 1.63 ± 0.13 μg m–2 h–1 from cryoconite holes (n = 4), and 0.92 ± 0.08 μg m–2 h–1 from red snow (n = 2), indicating that bare ice surface VOC emission rates were on average 2.4-fold and 4.3-fold higher than cryoconite hole and red snow emission rates, respectively. Oxygenated benzenoids accounted for just over half of the total emissions from bare ice surfaces (Mean VOC emission rates [± standard error (SE)] were 3.97 ± 0.70 μg msurfaces , with thsurfaces , while tsurfaces  and alkasurfaces .–1) of glacier ice algae   or red snow algae . The limited number of samples collected from red snow and cryoconite holes prevented a similar analysis for these habitats.VOC emissions from bare ice surfaces did not correlate with the number . Our results show that Greenland ice sheet surfaces colonized by microbial communities are a source of VOCs. Bare ice surface VOC emission rates were approximately one-third of those detected from tundra heath , measured using a push–pull system similar to that used in this study  presented different VOC profiles, with bare ice surfaces yielding the highest total emission rates , and the potential effect of these two parameters could not be separated in this study.The average chamber headspace temperature and relative humidity did not correlate with either air temperature or air relative humidity. However, the range of relative humidity in the chambers was similar to the range of relative humidity in the ambient air, as recorded at the PROMICE weather station QAS_M, giving confidence that representative relative humidity was prevalent during sample collection. In some cases, chamber temperatures were up to five times higher than air temperatures during the time of sampling, likely due to the greenhouse effect inside the chambers. This effect cannot be mitigated in the current experimental setup. We note that the parameters recorded at the PROMICE station, namely, air temperature and air relative humidity, were measured at approximately 2.6 m above the bare ice surface. This may explain some of the temperature variation, as near-surface conditions are highly variable and are affected by near-surface meteorology and heat exchange with the glacier surface . ChamberMore than 60% of the tentatively identified compounds detected in samples collected for bare ice surfaces have been reported as microbial volatiles in the literature –9. This in situ measurements of VOC emission rates for supraglacial microbial habitats have been reported. –1 in melted snow samples but found no link with bacterial or fungal counts from those samples. The majority of the most abundant VOCs in our study have previously been reported as microbial VOCs, but we cannot disregard the possibility that snow or ice melt may provide non-microbial sources of volatiles, hence further confounding simple correlations of cell counts with VOC emissions. Volatile release from melting ice needs to be assessed to better quantify potential microbial contributions to bare ice surface VOC emissions.Irradiance is one of the main drivers of ice sheet surface melt, and so any volatiles originating from the ice, as a result of melting out of recent and/or past deposition, will likely peak on sampling days with high irradiance. However, microbial contributions to the total VOC emissions from bare ice surfaces are also expected to peak at high irradiances for several reasons. First, VOC production is a common defense strategy in plants , as wellin situ carbon source.Cryoconite holes are considered hotspots for biogeochemical cycling in supraglacial systems . Thus, iC. reinhardtii . One-third of compounds detected in red snow VOC emissions have been reported as microbial volatiles  comparable to roughly one-third of those detected from tundra heath. This is especially interesting given that Greenland ice sheet bare ice surfaces present a source of VOCs in an area that has few other local VOC sources and that the Greenland bare ice extent is increasing in our planet’s warming climate. No direct correlation with ice surface algal counts was found, but the data presented here provide strong evidence that at least some of the detected VOCs are of microbial origin. A weak positive correlation between incoming shortwave radiation and VOC emission rates from bare ice surfaces was found, possibly indicating increased VOC emissions by supraglacial microorganisms in response to stress induced by high solar irradiance. Given that incoming shortwave radiation is correlated with chamber temperature, a microbial response to temperature increase, or increased volatilization from the liquid phase due to a local rise in temperature, may also contribute to higher VOC emission rates. Some of the main volatiles detected in bare ice surface emissions are known to have antifungal activity, pointing toward VOC emissions as a possible defense strategy against fungal infections in bare ice microbial communities. First estimates of VOC emissions from cryoconite holes (1.63 ± 0.13 μg m–2 h–1) and red snow (0.92 ± 0.08 μg m–2 h–1) are presented, suggesting that these habitats are also contributors to Greenland ice sheet VOC emissions. Further work is needed to identify constraints on and sources of supraglacial VOC emissions, focusing on both measurements in controlled and isolated systems, such as pure cultures or incubation experiments, and field measurements at different sites, under more controlled conditions and throughout the ablation season.This study is the first to report VOC profiles and emission rates from Greenland ice sheet surfaces colonized by active microbial snow, ice, and cryoconite hole communities. Bare ice surfaces present the largest source of VOCs, with emission rates (3.97 ± 0.70 μg mThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.All authors contributed with ideas to the experimental design. ELD collected all samples, carried out data analyses, prepared all figures, and wrote the first manuscript draft. CLD-M performed all GC–MS and PARADISe analyses. All authors contributed to the writing phase with discussion and revisions.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Osteoporosis is a common disease in elderly populations and is a major public health problem worldwide. It is not uncommon for spine surgeons to perform spinal instrumented fusion surgeries for osteoporotic patients. However, in patients with severe osteoporosis, instrumented fusion may result in screw loosening, implant failure or nonunion because of a poor bone quality and decreased pedicle screw stability as well as increased graft subsidence risk. In addition, revision surgeries to correct failed instrumentation are becoming increasingly common in patients with osteoporosis. Therefore, techniques to enhance the fixation of pedicle screws are required in spinal surgeries for osteoporotic patients. To date, various instrumentation methods, such as a supplemental hook, sublaminar taping and sacral alar iliac screws, and modified screwing techniques have been available for reinforcing pedicle screw fixation. In addition, several materials, including polymethylmethacrylate and hydroxyapatite stick/granules, for insertion into prepared screw holes, can be used to enhance screw fixation. Many biomechanical tests support the effectiveness of these augmentation methods. We herein review the current therapeutic strategies for screw fixation and augmentation methods in the surgical treatment of patients with an osteoporotic spine. With an aging population and longer life span, osteoporosis is becoming increasingly common ,2. In paTo date, various options of surgical instruments, such as a spinal hook ,9,10, suWe herein review currently available therapeutic strategies for screw fixation and augmentation methods in the surgical treatment of patients with an osteoporotic spine.Appropriately sized and positioned pedicle screws are important for obtaining optimal fixation in the osteoporotic spine . Larger Taken together, meticulous preoperative planning is essential for successful surgical outcomes. Surgeons should select the appropriate size of pedicle screws and carefully consider the number of screw placements and the number of fusion levels in order to obtain sufficient spinal stability and rigid screw fixation. Surgeons can use other surgical techniques to reinforce screw fixation, as described below .There is increasing evidence that pelvic fixation may become the standard method of various instrumented spine surgeries . Pelvic Long constructs extending from the thoracic spine to the distal lumbar spine and/or sacrum result in large lever arms and cantilever forces, causing substantial stress at the base of the construct. Furthermore, pedicle screws placed at S1 have a significant risk of screw loosening, especially in elderly patients with osteoporosis, as the S1 pedicles in the sacrum largely include cancellous bone ,24. TherAlthough pelvic fixation can be performed using various surgical procedures, the currently available iliac screw fixation and S2AI screw fixation techniques are considered the most common methods. A previous study suggested that S2AI screw fixation in adults may have a lower risk of mechanical failure and complication than iliac screw fixation . Indeed,While pelvic fixation using S2AI screws provides strong stability at the base of the construct , the impThe connection of an intra-sacral rod to pedicle screws placed at the lumbar spine is a technique for lumbosacral fixation that was reported by Jackson and McManus . ISBSs cThe cortical bone trajectory (CBT) screw technique is another pedicle screw insertion method described in 2009 by Santoni et al. . The CBTTraditional pedicle screw fixation requires extensive tissue dissection to expose the entry points and allow for lateral-to-medial screw trajectory insertion. In contrast, CBT screw insertion through a caudomedial starting point can reduce the length of incisions of the superior facet joints and paraspinal muscles ,40. ThusPrevious studies have suggested that modified pedicle screw insertion techniques penetrating the superior endplate of the vertebral body can be useful for enhancing the screw fixation strength ,16,17,41Minamide et al., compared fixation with an L-5 pedicle screw and an S-1 transdiscal screw connected by a rod to fixation with the standard L5–S1 construct . They foTaken together, these findings suggest that the screw penetrating the endplate may be a useful procedure for maximizing the thread contact with the dense bone purchase and achieving solid spine fusion . Indeed,Previous studies suggested that the groove entry technique and hooking screw technique for thoracic percutaneous pedicle screw (PPS) fixation may provide greater screw stability than conventional pedicle screw fixation ,44. In tIt has been reported that the placement of substances into the tapped screw hole increases the bone–metal interface friction force and enhances the mechanical strength of screw fixation ,46,47,48Several previous studies have suggested that an HA stick or HA granules inserted into the tapped screw hole may help enhance the initial strength of pedicle screw fixation ,22,23. FRecently, we reported a new augmentation technique for PPS fixation using HA granules . We devePMMA bone cements appear promising for the augmentation of pedicle screw fixation biomechanically in both osteoporosis and revision spine surgery models . Burval However, there is a risk of cement extravasation leading to potentially neurological or cardiovascular complications with cement use ,50. In aSeveral biomechanical studies demonstrated that expandable pedicle screws improved screw pullout stability compared with standard pedicle screws in osteoporotic spines ,58. It hHowever, expandable pedicle screws carry a risk of vertebral bone destruction during screw removal, which may cause nerve root or dural injury . NeverthPrevious studies have suggested that a sublaminar band can be useful for augmenting screw fixation ,12. A biThe polyester sublaminar band is soft and flexible, and the anterior–posterior spinal canal space occupied by the band is less than that taken by a sublaminar wire steel cable, thus avoiding direct spinal cord trauma during sublaminar passage. The flat configuration of the cable distributes the load over a larger contact area under the lamina than metal wires without producing imaging artefacts on postoperative magnetic resonance imaging (MRI) . In contPrevious biomechanical studies demonstrated that the augmentation of the pedicle screw with a laminar hook can improve stability of the spinal instrumentation ,9,10. ThSeveral studies have reported that a central hook–rod construct is effective for spinal osteotomy site closure ,63. In aOsteoporosis is a common disease in elderly populations and is a major public health problem worldwide . It is nAs described above, various screwing techniques to enhance fixation stability have been available for spinal instrumentation surgeries . In addiTo achieve success in surgical treatment for osteoporotic spines, meticulous preoperative planning is necessary. It is important to carefully consider the number of pedicle screw placements and the number of fusion levels in order to obtain sufficient spinal stability and rigid internal fixation. It is also essential to preoperatively assess not only the bone quality but also the bone morphology, including fusion/ankylosis of the spine in adjacent levels and spinal deformity and global alignment. The extent of fixation, including the level at which the construct ends, is critical to the prevention of junctional failure. The increased junctional strain at the termination of the construct may lead to accelerated deformity and kyphotic collapse, especially in osteoporotic patients ,65. The Anterior column support increases load-sharing and decreases strain on the posterior screw fixation construct . CombineDrug administration to improve osteoporosis can reportedly enhance pedicle screw stability and spinal fusion after surgery ,68,69,70More rigid screw fixation with effective augmentation may reduce the risks of screw loosening and implant failure and provide better postoperative clinical outcomes. Further translational studies should thus be performed to provide firm evidence concerning the optimal surgical treatment of the osteoporotic spine.


Electroconvulsive therapy (ECT) is proposed to exert an effect on white matter (WM) microstructure, but the limited power of previous studies made it difficult to highlight consistent patterns of change in diffusion metrics.We initiated a multi-site mega-analysis and sought to address whether changes in WM microstructure occur following ECT.To this end, diffusion tensor imaging (DTI) data (n=58) from 4 different sites were harmonized before pooling them by using ComBat, a batch-effect correction tool that removes inter-site technical variability, preserves inter-site biological variability and maximizes statistical power. Downstream statistical analyses aimed to quantify changes in Fractional anisotropy (FA), Mean Diffusivity (MD), Radial Diffusivity (RD) and Axial Diffusivity (AD), by employing whole-brain, tract-based spatial statistics (TBSS).ECT increases FA in the right splenium of the corpus callosum and the left cortico-spinal tract. Both the left superior longitudinal fasciculus and the right inferior fronto-occipital fasciculus showed increases in AD. Increases in MD and RD could be observed in overlapping white matter structures of both hemispheres. Finally, responders showed significantly smaller FA values in the left forceps major and smaller AD values in the right uncinate fasciculus compared with non-responders.This is the first and largest multi-site mega-analysis to demonstrate that ECT normalizes altered WM microstructure in important brain circuits that are implicated in the pathophysiology of depression. Furthermore, responders appear to present a more decreased WM integrity at baseline, which if replicated could serve as a biomarker for ECT response.No significant relationships.


Identification of lymph nodes (LNs) draining a specific site or in obese macaques can be challenging.Indocyanine Green (ICG) was administered intradermal (ID), intramuscular, in the oral mucosa, or subserosal in the colon followed by Near Infrared (NIR) imaging.After optimization to maximize LN identification, intradermal ICG was successful in identifying 50–100% of the axillary/inguinal LN at a site. Using NIR, collection of peripheral and mesenteric LNs in obese macaques was 100% successful after traditional methods failed. Additionally, guided collection of LNs draining the site of intraepithelial or intramuscular immunization demonstrated significantly increased numbers of T follicular helper (Tfh) cells in germinal centers of draining compared to nondraining LNs.These imaging techniques optimize our ability to evaluate immune changes within LNs over time, even in obese macaques. This approach allows for targeted serial biopsies that permit confidence that draining LNs are being harvested throughout the study. To this end, we had previously developed minimally invasive (MIS) laparoscopic techniques to permit sampling of key immunologic sites,Sampling of peripheral lymph nodes in macaque models can also present challenges, with the need to sample sites repeatedly combined with factors such as obesity; LNs can be hard to find resulting in missed time points. As macaque studies are typically only powered sufficiently to detect differences when all samples are evaluated, missed time points can impact the ability to achieve meaningful and statistically significant results. Additionally, for events that are localized in nature, such as immunization at a specific site, it can be important to ensure that the LNs collected are actually draining the site of interest. Drainage patterns can vary by individualTo address issues associated with accuracy of lymph node collections, we have optimized a dye‐guided lymph node (DGLN) collection technique for use in the macaque model. This technique utilizes the dye indocyanine green (ICG) which is visualized with near infrared (NIR) imaging to allow for 1. Targeted collection of LNs from specific sites, and 2. Identification of LNs in obese animals where they were not identifiably using standard methods. Here, we present the DGLN technique for identification of MLN in obese animals and the identification of LNs draining the site of immunization in both the oral mucosa  and bicep muscle (IM inoculation). Draining LNs were compared to nondraining LNs from these sites  following immunization and significant differences in replication and levels of key immune cells within germinal centers were identified.22.1All animals were housed, cared for, and used in accordance with the policies of the Institutional Animal Care and Use Committee (IACUC) at the ONPRC, an AAALAC‐accredited institution, which abides by the USDA Animal Welfare Regulations, the Public Health Service Policy on Humane Care and Use of Laboratory Animals and the Guide for the Care and Use of Laboratory Animals. All enrolled macaques were SPF  prior to the start of the study and were captive‐born at ONPRC. All animals were singly or socially housed in accordance with their IACUC protocol. Throughout the study, all animals were uniformly fed Purina LabDiet 5000 , daily nutritional enrichment items  and had ad libitum access to water. In addition to social housing and nutritional enrichment, animals received environmental enrichment in the form of toys, music/radio, and television daily. All animals were observed for health or behavioral concerns at minimum twice daily by research, veterinary, and/or husbandry staff. Animals requiring euthanasia for study or clinical endpoint purposes were sedated with an intramuscular injection of 20 mg/kg ketamine HCl  followed by an overdose of intravenous pentobarbitol solution, in accordance with the AVMA Guidelines for the Euthanasia of Animals.Nineteen rhesus macaques , ranging from 3 to 16 years of age and 5.6–12.1 kg were used to optimize ICG administration for fluobeam PLN detection. Dose volume, number of injection sites, and duration post‐administration were analyzed to optimize LN signal identification for inguinal and axillary LN sites. Collection of MLN and PLN using ICG guidance in obese rhesus macaques was performed as part of their primary study once standard methods had failed to identify an MLN and/or PLN at that timepoint. Thirteen obese rhesus macaques  ranging from 7 to 17 years old and 7.5–16.1 kg were evaluated; 7/13 were used for PLN, and 9/13 were used for MLN detection. A total of 2 rhesus  macaques (n = 1/group) were injected with ICG labeled 10 014_F8.V3.N7.SOSIP (natively HIV envelope folded trimer) either by IM injection in the biceps brachii or IEp injection in the oral mucosa. A boost injection at the same site was performed 4 weeks after the initial immunization, and the draining LNs  were collected 2 weeks after the boost. Unbound ICG was also injected at the site of immunization prior to surgery to ensure that sufficient signal was present in the draining LNs for identification by NIR. Nondraining  LNs from the same site  were collected at the same time for comparison to the draining LNs.2.2LLC) was reconstituted with sterile water to 5 mg/mL and then diluted with sterile saline to 0.625 mg/ml concentration prior to use. Macaques were sedated with an intramuscular injection of 10–12 mg/kg ketamine HCl  and 0.015 mg/kg dexmedetomidine hydrochloride . ICG was administered through a 25‐27G needle at volumes of 0.1–0.4 ml per site intradermal (ID), IEp in the oral mucosa, IM in the biceps brachii, or subserosal in the colon to identify draining LNs using NIR imaging. Real‐time NIR imaging was performed using a Fluobeam800 for PLN and a Stryker 1688 for laparoscopic procedures.ICG (Diagnostic Greenrsb.info.nih.gov/ij) to determine the pixel area of fluorescent signal to determine optimization for the technique.For the inguinal LN site, the lower caudal abdomen and proximal medial thigh were sites of ICG administration; for the axillary LN site, the proximal medial brachium and anterolateral aspect of chest were sites of ICG administration. ICG injection sites were selected approximately 5 cm from the expected location of axillary/inguinal LNs to avoid overlap of the NIR signal from the injection site with the expected location of the LNs. The Fluobeam800 was positioned 20 cm from the patient, and 0.1–0.2 ml of ICG (0.625 mg/ml) ID was administered at 1 or 2 sites. The Fluoptics Fluorescence Imaging for Surgery (Fluobeam Software v. 4.0) was utilized  to capture 5, 10, and 15 minute time points. Image fluorescence were quantified by pixel threshold analysis using ImageJ software  that developed broadly neutralizing antibodies approximately 3 years post‐infection (cite 25 122 781). For conjugation, protein was mixed with NH‐Reactive ICG and Reaction Buffer and incubated for 10 min at 37°C. The reaction was stopped with WS buffer and filtered, and the labeled protein was diluted into phosphate‐buffered saline (PBS). Labeling efficiency was determined by assessing absorbance at 280 nm and 800 nm and calculating the absorption coefficient. ICG‐labeled Env vaccine formulation was prepared with the following adjuvants so that each injection contained 100ug Env trimer, 100ug R848 (Invivogen), 10ug MPLA (Invivogen), and 1% Alhydrogel (Invivogen). The labeled SOSIP trimer was injected in a volume of 0.1 ml/site into the oral mucosa of both the left and right cheek pouch. For IM, a volume of 0.25 ml was injected into the right and left biceps brachii.10 014_F8.V3.N7.SOSIP Envelope protein was expressed in HEK293 cells and purified by GNA lecting chromatography as previously described.2.4Biopsies were performed as described previously.2.5PLN and MLN were obtained as described above and were processed then stained for flow cytometric analysis. Lymph nodes were collected in R10 media  at biopsy and were then processed using a pestle and screen for CD‐1  in a tissue culture dish. Processed tissues were strained through a 40 μm filter into a 50 ml conical, centrifuged at 700 rcf for 10 minutes, and resuspended in R10 before obtaining the cell yield using a Horiba ABX Pentra 60 C+ hematology analyzer.2.6Formalin‐fixed, paraffin embedded 5 μm sections were deparaffinized and decloaked in 1% citrate buffer at pH 6.0 (Vector Laboratories). Sections were blocked in 1%BSA with goat serum prior to staining for CD3 , PD‐1 , and CD20  overnight. Following primary staining, tissues were washed and incubated with goat anti‐mouse Alexa Fluor 488  and goat anti‐rabbit Alexa Fluor 594  for 1 hour at room temperature. Background signal was quenched by 1‐minute incubation using Vector TRUEView Autofluorescence Quenching Kit (Vector Laboratories). Coverslips were mounted, and nuclei stained using Vectashield Hardset Antifade Mounting Medium with DAPI (Vector Laboratories). Composite tissue images were acquired on a Keyence BZ‐X710 fluorescence microscope at 20× magnification. Identification of active germinal centers and T‐cell quantification were performed using open‐source packages available in FiJI.2.7p value was less than .05.All data are shown as standard deviation (SD) of the mean. Optimization results were analyzed using paired t‐tests for comparison of signal intensity at different times within an animal and unpaired t‐tests for comparison of volume and number of sites using GraphPad Prism  version 9.0 for Windows. Results were considered statistically significant when the 33.1p = .032), 3.3‐fold greater at 10 min (p = .007), and 2.6‐fold greater at 15 min (p = .013) when one site on the limb (arm or leg) and one site on the chest/abdomen were used for axillary and inguinal, respectively, at a 0.2 ml volume of ICG . We then assessed the signal intensity in the nodes at different times post 2 site ID administration of 0.2 ml of ICG and determined that 5 min was optimal through visual assessment.ICG was injected at a concentration of 0.625 mg/ml at various locations in the axillary and inguinal areas of macaques as either a single site or at two sites. This concentration was sufficient to allow for identification of multiple LNs down the lymphatic draining chain of nodes and had a mild quenching effect at the center of the injection bleb when imaged  of the nodes present at the site demonstrating detectable fluorescent signal using the Fluobeam800, and all animals had at least 2 nodes that were positive at each site. Thus, optimization of our dye‐guided lymph node (DGLN) collection technique resulted in 0.2 ml/site x 2 site ICG injections with imaging performed at least 5 minutes post ID administration for PLN detection.3.26/animal (0.8–120 × 106). For MLN, the same concentration of ICG was used for DGLN but was injected at ~0.4 ml/site at 2–3 sites subserosally in the colon .ICG as optimized for PLN DGLN collection was used in a total of 7 obese rhesus macaques after traditional methods for identification of axillary/inguinal LNs had failed. PLN were identified and collected in 100% (7/7) using DGLN resulting in an average lymphocyte yield of 31.1 × 103.3p < .001) and submandibular (p < .01) locations. The elevated size and number of GCs, as well as increased Tfh cell levels in ICG+ lymph nodes, strongly suggest vaccine‐specific responses are occurring at tissue‐draining lymph nodes and can be identified by the ICG infrared imaging.NIR imaging was utilized to target LNs draining the site of an HIV Env SOSIP immunization in two rhesus macaques, one immunized via IM injections and a second via an IEP injections to the oral mucosa. Two doses of vaccine were administered 8 weeks apart. Two weeks after the second vaccination, either axillary (in IM vaccinated) or submandibular  lymph nodes were obtained. NIR DGLN imaging was utilized to identify both nondraining Figure  and drai4We demonstrated the utility of our optimized DGLN technique using ICG, an FDA‐approved fluorescent dye that has both an extremely good safety profile and minimal potential to impact immune responses, for identification of both PLN and MLN in rhesus macaques. This resulted in a 100% rate of identification of LNs that could not be identified by traditional methods. Avoiding missed timepoints is key not only for ensuring that missed samples from unique animals do not skew results, but also to ensure that sufficient numbers are present to achieve statistical significance. Most macaque studies are only powered sufficiently to allow for statistical comparisons when the data set is complete, and missing samples that require elimination of an animal from a timepoint or from the study can result in a loss of the statistical power required to adequately answer an experimental question. We have demonstrated that these fluorescence DGLN techniques can be combined with other techniques to permit a more comprehensive evaluation of changes in the immune system over time in macaque models. It is not uncommon for us to perform comprehensive evaluations inclusive of PLN, MLN, spleen, liver, bone marrow, bronchoalveolar lavage, and mucosal biopsies  collected during the same anesthetic event at up to 8 timepoints during the course of a study. This is performed using minimally invasive techniques that optimize animal welfare, minimize impacts to the animals and associated experimental results, and allow for longitudinal comparisons within an animal avoiding the inter‐animal variability seen in animal intense serial sacrifice studies.Additionally, we demonstrated the ability to use ICG DGLN to identify draining versus nondraining LNs present at axillary and submandibular sites. These initial analyses utilizing one macaque vaccinated via the oral IEp route, and one vaccinated via the IM route were able to provide evidence that ICG‐positive LNs draining the site of immunization were distinct from those that were physically adjacent but ICG‐negative. Indeed, both greater numbers/size of B cell follicles were observed in the ICG‐positive LNs, as well as higher levels of Tfh positive cells in the germinal centers of these follicles. The levels of Tfh likely represent the proliferation of these cells following exposure to the SOSIP trimer vaccine, Tfh cells then stimulate B cells to expand and eventually produce antibodies. Importantly, the identification of nodes with extensive germinal center formation provides confidence that we were able to correctly identify the active draining lymph nodes responding to vaccination, allowing us to selectively harvest the sites of immunologic importance for downstream analyses.These DGLN techniques can be employed either using a labeled target The use of NIR imaging is ideal for DGLN identification given the long wave length which leads to greater tissue penetration when compared to other fluorescence imaging techniques. ICG is ideally suited as a DGLN contrast agent for use in NIR detection of draining LN given the long (>800 nm) wavelength of emission and the corresponding depth (~1 cm) of tissue fluorescence signal penetration.In conclusion, we have developed NIR DGLN techniques for identification of PLN and MLN, providing a powerful tool that can be used to study immunity in macaques with a high degree of accuracy and specificity. Applications that analyze lymph node‐specific responses are heavily dependent on accurate recovery of immunologically active lymphoid tissue and thus will greatly benefit from the incorporation of NIR‐guided LN identification. More broadly, because ICG can be coupled to many types of molecules, this technique is compatible with virtually any approach aimed at identifying trafficking of biologics or compounds to the lymph nodes, creating the opportunity to selectively and accurately analyze outcomes of LN‐specific interventions.The authors do not have any conflicts of interest in regards to this publication.This study was performed in accordance with the ethical policies of the journal, and in compliance with Guide for the Care and Use of Laboratory Animals, the USDA Animal Welfare Regulations and the Public Health Service Policy on Humane Care and Use of Laboratory Animals.


Fractures of the superior pubic ramus can be treated with screw insertion into the osseous fixation pathway (OFP) of the anterior column (AC). The entry point determines whether the screw exits the OFP prematurely. This can be harmful when it enters the hip joint or damages soft tissues inside the lesser pelvis. The exact entry point varies between patients and can be difficult to ascertain on fluoroscopy during surgery. The aim of this study was to determine variation in the location of the entry point. A retrospective single center study was performed at a level 1 trauma center in the Netherlands. Nineteen adult patients were included with an undisplaced fracture of the superior pubic ramus on computer tomography (CT)-scan. Virtual three-dimensional (3D) models of the pelvises were created. Multiple screws were placed per AC and the models were superimposed. A total of 157 screws were placed, of which 109 did not exit the OFP prematurely. A universally reproducible entry point could not be identified. A typical crescent shaped region of entry points did exist and was located more laterally in females when compared to males. Three-dimensional virtual surgery planning can be helpful to identify the ideal entry points in each case. Pelvic and acetabular fractures are severe musculoskeletal injuries, which are mostly caused by high-energy traumas. Minimal invasive fixation techniques can avoid the morbidity of open surgery in these injuries . These tOpportunities for minimal invasive fixation of the pelvis and acetabulum are conceptualized as osseous fixation pathways (OFPs). OFPs are geometrically complex corticated bony cylinders of different dimensions and orientations that can accommodate intraosseous implants . The antThe location of the entry point varies between patients. Typically, intraoperative fluoroscopy is required to determine its exact location . This caOnly two studies have investigated the localisation of entry points into the OFP of the AC. Their approach involved spatial measurements that were reported as average positions on the pelvis. Such results are less practical during minimally invasive techniques where the soft tissue envelope remains intact. In addition, the theoretical average position of an entry point will often vary between actual patients.We hypothesise that there is an entry zone, a collection of multiple entry points, from which screws can safely be implanted into the AC of adult patients. The aim of this study was to investigate the extent of variation in entry points. To this end we performed a three-dimensional (3D) analysis of the anatomy of AC of the pelvis and safe screw positions in the OFP.Ethical review and approval were waived for this study by the local medical research ethics committee, due to the fact that all included computer tomography (CT)-scans were acquired during standard treatment, anonymisation and retrospective nature of the study. The requirement for patient consent was waived as all image data were anonymized and de-identified prior to analysis. This study followed a guideline named the strengthening the reporting of observational studies in epidemiology statement (STROBE) .This is a retrospective single-center study performed at the Radboud University Medical Center (RUMC), a level 1 trauma center in Nijmegen, the Netherlands. Trauma patients were considered eligible for inclusion if they were treated at RUMC between 2017 and 2021. They were included when 16 years or older and sustained a fracture in the AC of the pelvis. Exclusion criteria were preoperative CT scans of the pelvis with a slice thickness of more than 1.0 mm and fracture dislocation where percutaneous screw placement would technically not be possible. All image data were pseudonymized and de-identified prior to analysis. The keys to patient-specific data were kept separate from the study data and were only accessible to the principal investigators. Patient-specific characteristics as sex, age and length were derived from the electronic medical records and collected in an Excel database .The primary outcome was to determine the extent of variation in entry points for screw insertion into the OFP of the AC of the pelvis. Secondary outcome measures were the diameter and length of the screws. Differences between men and women were also studied, with both anatomical variations and variations in entry point.Digital Imaging and Communications in Medicine (DICOM) files corresponding to the selected CT scans were retrieved from the Picture Archiving and Communication System (PACS). The DICOM files were used to create virtual models of the pelvis using Mimics . The threshold values were set to identify cortical bonee, i.e., from 226 to 1504 Hounsfield units (HU). The femora and the lumbar vertebra upward of L5 were removed from the models. The models were subsequently smoothened (Smooth Mask function). The models were exported as Stereolithography (STL) files.® . 3DMedX® was used to perform the virtual screw placement in the AC. The transparency of the pelvic models was downgraded to make the intra-osseous course of the screw visible. Inlet and obturator-outlet view were used to assess screw placement inside or outside the OFP. The models of the screws had a diameter of 3.5 mm or 6.5 mm, corresponding to the diameter of commonly used screws. As many well-positioned screws as possible were placed per case. All digitally placed screws were supervised by senior authors with extensive experience in treating pelvic fractures (E. Hermans and V.M.A. Stirler). Files were uploaded in 3DMedx. The CT slices were superimposed on the 3D model. Random reslicing was carried out by scrolling through the central axis of the screw, to verify whether the screws were positioned correctly  in order to obtain only right-sided segmentations. The clinically most optimal screw position was retained per case. Three different methods were used for superimposing in 3DmedX®. In the first method, the Iterative Closest Point (ICP) algorithm was used to match segmentations based on the anatomy. The anterior column was selected as the focus for ICP matching. In the second method, a Procrustes analysis was used for matching. Four landmarks were used for the Procrustes analysis: the entry- and exit point of the screw, the pubic tubercle and a point at the pubic crest closest to the pubic symphysis. The final method consisted of superimposing the models manually.The variation in entry points can be compared between cases by superimposing the 3D segmentations of the different pelvic models. The models were mirrored using Meshmixer®. The distance from the lower limit of the entry points to the border of the inferior ramus was measured as well. The circumference of the entry zone was measured using Circumference Calculation in 3DMedX®. The region of entry points was selected, and the size of the selected area was assessed using the function called Area Calculation Selection.Male and female cases were analyzed separately, and the entry points were compared with each other. Additionally, the distance between the upper limit of the entry points and the pubic tubercle was measured using a digital ruler in 3DMedXA total of nineteen patients were included in this study. Patient characteristics were presented in A total of 157 virtual screws were placed. A total of 109 (69%) screws did not exit the anterior column prematurely and had entry points that were accessible through a retrograde approach. A total of 13 (8%) screws were placed in the OFP of the AC but their entry points were deemed surgically dangerous. Another 35 (22%) screws did exit the OFP prematurely .Three different methods were used to determine the variation in the entry points. Superimposing the pelvic models resulted in inadequate matching when based on anatomy or on landmarks on the OFP. Matching of the pubic bones seemed adequate, but the OFPs diverted .The best results were gained after manually matching the models, especially when males and females were analysed separately .Subsequently, the entry points’ zone wase determined using the superimposed 3D models of males and females. A crescent-shaped zone of entry points was identified on the pubic bone. The entry zone was located more laterally in females when compared to males . A univeA quantitative analysis of the entry zones was performed . In mostKnowledge of appropriate entry points can be helpful in the expedient and safe placement of screws into OFPs. In this study, we used a 3D analyses to investigate the variation in entry points for screw placement inside the OFP of the AC of the pelvis. A universally reproducible entry point could not be determined. Rather, crescent-shaped entry zones were identified, from which screws may be safely placed into the OFP of the AC.Few studies have investigated screw placement into the AC column. Only two studies investigated the location of entry points. Ochs et al.  investigDifferences between males and females were assessed in both the present study and previous studies. According to Ochs et al., the entry point was located significantly more laterally from the symphysis in females (34.4 ± 5.5 mm versus 26.7 ± 3.8 mm) and closer to the cranial margin of ramus superior ossis pubis (9.9 ± 1.4 mm versus 12.2 ± 1.4 mm). These findings are consistent with the results of our study. The screw diameter was also investigated. In 90% of the males, the anterior column accommodated a 6.5 mm screw. A 6.5 mm screw did not fit one third of the females. These findings are in line with the results of previous studies, which say that the maximum diameter of a screw is statistically significantly larger in males compared to females [In previous studies on this topic, only one optimal screw was placed per patient. An ideal screw position was defined when the screw was entirely placed within the periacetabular corridor with a central position at the narrowest points of the corridor . This deIn this study, multiple screws were positioned per patient. More than 150 screws were analysed. This illustrated that there are different options for correct screw placement per patient, which allows for greater flexibility during surgery. All possible options for screw insertion are included in the results.Several limitations require mentioning. Greater accuracy may have been achieved if a larger population was studied. Screws were mostly placed in intact AC. We assume that similar results are achieved in undisplaced fractures. Manually matching 3D models was superior to automated methods but not perfect. Other methods, such as Statistical Shape Modelling (SSM), may arguably result in more accurate matching. However, we expect the difference to be minimal.Virtual 3D reconstruction models provide practical insight into patient-specific anatomy and can also be useful in preoperative virtual surgery planning ,9. PreopIn conclusion, variation exists in the entry point for screw insertion into the anterior column of the pelvis. A universally reproducible entry point does not seem to exist. Interestingly, the location of the entry point is more lateral in females. A patient-specific surgical planning based on a virtual 3D segmentation can be used to provide personalized medicine.


Background: Atopic dermatitis constitutes one of the most common inflammatory skin manifestations of the pediatric population. The onset of many inborn errors occurs early in life with an AD–like picture associated with a deregulated IgE response. The availability of proteomic tests for the simultaneous evaluation of hundreds of molecules allows for more precise diagnosis in these cases. (2) Methods: Comparative genomic hybridization microarray (Array–CGH) analysis and specific IgE evaluation by using allergenic microarray (ISAC) and microarray (ALEX2) systems were performed. (3) Results: Proteomic investigations that use multiplex methods have proven to be extremely useful to diagnose the sensitization profile in inborn errors with deregulated IgE synthesis. Four patients with rare diseases, such as recessive X–linked ichthyosis , Comel–Netherton syndrome , monosomy 1p36 syndrome (OMIM: 607872), and a microduplication of Xp11.4 associated with extremely high levels of IgE: 7.710 kU/L, 5.300 kU/L, 1.826 kU/L, and 10.430 kU/L, respectively, were evaluated by micro– and macroarray multiplex methods. Polyreactivity to both environmental and food allergens was observed in all cases, including the first described case of association of X–chromosome microduplication and HIE. (4) Conclusions: Extensive use of proteomic diagnostics should be included among the procedures to be implemented in inborn errors with hyper–IgE.(1)  S. aureus and other microbial antigens have been identified [Atopic dermatitis (AD) constitutes one of the most common inflammatory skin manifestations, affecting up to 20% of the pediatric population and 10% of the adult population, associated with a complex interplay between genetic and environmental causal factors not all fully understood to date . Intrinsentified .Genetic pieces of evidence depict a complex network involving epidermal barrier dysfunction and dysregulation of innate and adaptive immunity in AD patients. GWAS and Immunochip analysis have identified a total of 19 susceptibility loci for AD. Mutations in the human filaggrin gene (FLG) are the most significant and well–replicated genetic mutations associated with AD. However, several studies demonstrated that FLG knockout alone is not sufficient to cause an AD–like phenotype in mice, suggesting the intriguing hypothesis that additional atopy–promoting gene variants are required to cause atopy in the context of FLG deficiency . Indeed,STAT3) gene, which plays an essential role in signal transduction of several cytokines to their respective receptors . STAT3–HIE, also known as Job’s syndrome, is associated with recurrent staphylococcal skin abscesses (cold abscesses), chronic mucocutaneous candidiasis (CMC), cystic pneumonia, and skeletal abnormalities [On the other hand, some inborn errors often look similar to AD, both clinically and due to the finding of elevated levels of circulating IgE, which sometimes also leads to delays in proper diagnosis. This hyper–IgE (HIE) syndrome may also be associated with severe allergic phenomena, so it is essential to consider extensive in vitro allergy investigations in these cases, to unveil any hidden sensitizations that may, at various levels, impact the patient’s quality of life. HIE syndromes are a series of rare primary immunodeficiencies characterized by extremely high levels of IgE, AD–like eczema, and recurrent skin and/or lung infections (HIE triad), often associated with high mortality due to severe infections and high risk of malignancies. As mentioned, discriminating such forms from pictures of severe atopic dermatitis, especially in early childhood, can be very difficult. The most common form of HIE is caused by an autosomal dominant (AuD) negative mutation in the human signal transducer and activator of transcription 3  gene that encodes for lymphoepithelial kazal type–related inhibitor (LEKTI), whose deficiency is associated with elevated desmoglein 1 degradation and subsequent stratum corneum detachment , trichorrhexis invaginata, also known as “bamboo hair,” and HIE [FOXP3 gene codifies a transcription factor essential for the development of CD4+ T regulatory (Treg) cells. The absence of Treg cells prevents physiological regulation of the immune response, which will therefore be dysregulated on both the autoimmune and allergic response sides, thus causing the immunodysregulation polyendocrinopathy enteropathy X–linked (or IPEX) syndrome [An autosomal recessive (AR) form of HIE is caused by a deficiency of the dedicator of cytokinesis 8 (DOCK8). A DOCK8 deficit results in severely affected adaptive immunity and lymphocyte and dendritic migration; hence, the high susceptibility to viral skin and lung infections. T lymphocytes are Th2–imbalanced, with very high IgE levels and often severe food allergies, unlike what is observed in STAT3–HIE . Comel–N and HIE . The X–lsyndrome .It is widely known that the prototypical STAT3–loss–of–function (LOF)–associated HIE syndrome is not associated with “allergic” phenomena despite the very high levels of circulating IgE due to both the reduced mast cell degranulation capacity in these patients and a likely lack of IgE affinity maturation due to a functional deficiency of T follicular helper 13 cells . On the Over the past 15 years, approximately 63,000 individuals referred to our institute, an Italian reference centre for dermatology and allergology and member of the European Reference Network on Rare and Undiagnosed Skin Disorders (ERN–skin), have been evaluated by proteomic methods for specific IgE assay. Of these individuals, a few dozen were affected by rare forms of inborn errors  characterized by deregulated IgE synthesis, in the context of immunodeficiencies of the most disparate kinds. Previous studies have documented the sensitization proteomic profile in these patients ,14, but ® Laboratory Information System  and a customized electronic database. Sera from the participant were collected and stored at the time of the visit.The study participants  were enrBased on the presence of multiple congenital anomalies and dysmorphisms, a comparative genomic hybridization microarray (Array–CGH) analysis using the CytoChip oligo–array ISCA 4X44K resolution  was performed according to the recommendations of the manufacturer. Data were then analyzed by using the Blue–Fuse for microarrays software package (BlueGnome).Mutation screening was performed on blood genomic DNA by amplification and bidirectional Sanger sequencing of SPINK5 exons and flanking intronic borders (GenBank NM_006846.3), as described . LEKTI e® , and Allergy Explorer–ALEX2® .The blood samples were stored during the control visits from 2012 to 2022 for the evaluation of IgE reactivity by using ImmunoCAP ISACIn brief, ISAC slides were washed, rinsed, and dried at room temperature (RT). Undiluted serum (30 μL) from each patient was pipetted onto the slide and after 120 min incubation at RT in a humid chamber, slides were washed, rinsed, and dried. IgE binding was detected by the addition of an antihuman secondary antibody. Slides were then washed, rinsed, dried, and stored in the dark until scanning. Images were acquired by scanning allergen biochips with a CapitalBioLuxScan™ 10K microarray scanner. IgE values were expressed as ISU arbitrary units (ISAC Standardized Units) corresponding to IgE antibody levels in the ng/mL range (detection limit: 0.3 ISU–E).In the ALEX test, 300 allergens, including molecules and extracts, are spotted onto a nitrocellulose membrane in a cartridge chip, which is then incubated with 0.5 mL of a 1:5 dilution of serum, containing a CCD inhibitor under agitation. After incubation for two hours, the chips are washed three times and a pretitred dilution of antihuman IgE labelled with alkaline phosphatase is added and incubated for 30 min. After another cycle of extensive washing, the enzyme substrate is added, and after eight minutes, the reaction is stopped. The membranes are dried, and a charge–coupled device camera measures the intensity of the colour reaction for each allergen spot. The dedicated software digitalizes the images and prepares a report that lists the allergens and components and their score in kUA/mL. Finally, an arbitrary calibration curve is obtained by reacting four spots with decreasing concentrations of specific IgE corresponding to <0.35, 0.35–1, 1–5, 5–50 kUA/L and >50 kUA/L. We considered positive a concentration of ≥0.3 kUA/L.X–linked recessive ichthyosis (RXLI), the second–most common form of ichthyosis , is a benign form of ichthyosis that affects almost exclusively males and is inherited through carrier females. RXLI is an epidermal lipid metabolism defect due to inactivating mutations or deletions in the steroid sulfatase STS gene (Xp22.3), thus resulting in disorders of cornification, mainly localized on extensor and flexor areas of the extremities, chest, and neck (“dirty neck” appearance) .A 12–year–old male patient with RXLI already identified in another centre came to our observation as suffering from a severe form of atopic dermatitis accompanied by elevated levels of IgE (7710 kU/L). After evaluation with macroarray proteomics , the fol2 test, currently available in the market can eliminate 60–70% of cases or at least reduce the CCD signal, using a specific inhibitor.Interestingly, very high levels of IgE directed against cross–reactive carbohydrate determinants (CCD) were observed in this case (Hom s Lactoferrin | CCD marker ), undersComel–Netherton syndrome is characterized by an ichthyosiform AD–like pattern, a bamboo–like hair–shaft defect and multiple atypical manifestations, caused by SPINK5 mutations .A 52–year–old patient with NS, carrying the SPINK5 mutation c.238insG (ex4)/nonsense R217X (ex 8), associated with absent expression of LEKTI, came to our observation presenting a scaly erythroderma cutaneous form and complaining of violent postprandial abdominal pains. Elevated IgE levels (5300 kU/L) were observed.Proteomic diagnostics  demonstrThe patient was also allergic to several food allergens such as nsLTP , arginine kinase (thermolabile molecule of crustaceans), and parvalbumins from fish  see .A diet eliminating the foods that emerged from the microarray test had determined an immediate and marked improvement in the patient’s cutaneous picture, and gastrointestinal symptoms, with a great improvement in the perceived quality of life.The 1p36 deletion syndrome is characterized by multiple congenital abnormalities  and mental retardation . DeletioA 20–year–old patient affected by the 1p36 deletion syndrome caused by a cr.1 short–arm microdeletion in the p36.33p36.32 region, already identified at birth in another centre, was affected by a cognitive and stature developmental delay, ear asymmetries, sunken eyes, flat nasal bridge, and chronic inflammatory polyarthritis. Cognitive retardation was associated with communication and relationship difficulties, motor stereotypies, bilateral convergent strabismus, locomotion difficulties, ataxic gait, cingulate hypotonia, and easy fatigability. The girl communicated with gestures, complained continuously, especially at night, and often showed urticarial rash accompanied by severe itching, so she was placed on continuous antihistamine therapy.Routine blood tests showed elevated IgE levels (1826 kU/L), and therefore the patient was accompanied to our out clinics, where a proteomic test was performed , showingThe modification of the dietary regimen operated based on the molecular tests, with the exclusion of wheat and fruit, resulted in the total disappearance of the symptoms that had been afflicting the young patient for several years, showing the usefulness of a multiplex approach even in those genetic defects characterized by a neurodevelopmental disorder with or without anomalies of the brain, with poor or absent speech, and inability to collect a correct clinical history.A 7–year–old woman presented a clinical picture characterized by cognitive defect, overweight, cerebellar malformation, (cerebellar hypoplasia left hemisphere), bilateral clubfoot (undergone surgery), AD, marked keratosis pilaris, acanthosis nigricans, thick lips, hair anomalies , HIE syndrome, was the firstborn of a healthy, young and unrelated couple, and had a healthy sister two years younger. No family history of genetic diseases was reported. The patient was delivered at 39 weeks of gestation by caesarean section. Her birth weight was 3 kg and her length was 52 cm. She was breastfed for three months.The genomic analysis identified a microduplication of 535 kb on chromosome Xp11.4, bridging from 38,561,541 bp to 39,096,541 bp, according to the human genome NCBI Build 38 version . Karyoty® microarray containing an increasing number of allergens tested from 79 to 103, 112, and, more recently, 295 with the ALEX2® macroarray), showed an IgE polysensitization picture, including environmental, both indoor (house dust mites and moulds) and outdoor , and food allergen reactivity  and sensitization to various environmental allergens. Serial evaluations carried out with multiplex methods , reactivity to cross–reactive carbohydrate determinants was evidenced. This sensitization, devoid of any clinical impact, may result in false in vitro reactivities capable of, if not identified, fouling the diagnostic data and inducing erroneous behavioural interventions . This fiIn another case (microduplication of Xp11.4), reactivity to autoreactive (SOD) allergenic components was shown. Manganese superoxide dismutase (MnSOD), which may play a role as an autoantigen  in the context of an autoimmune reactivity, can elicit a specific IgE response in a subset of patients with atopic dermatitis . This paThe patient suffering from monosomy 1p36 Syndrome had no inborn error correlated with altered IgE response , so she pos skin dendritic cells (DCs) from atopic dermatitis patients is significantly higher when compared to skin DCs of not allergic healthy individuals [In this study, in addition to inborn errors already known, such as NS, 1p36 deletion syndrome, and RXLI, we reported a case of micro–duplication of Xp11.4 involving 2 genes, TSPAN7 and MID1IP1. TSPAN7  is a member of the tetraspanin superfamily, cell–surface proteins found in all multicellular eukaryotes. Genetic defects of the relative coding gene (TM4SF2) are associated with X–linked intellectual disability . Howeverividuals . On the ividuals . Lyon’s ividuals ,35. In tividuals . Even inWhen Xp duplication occurs, doubled genes can cause functional disomy of usually inactivated genes in normal females with XX karyotype. In the patient we describe, two genes were present in the duplicated 535 kb microsegment. One of them has already been deemed responsible for X–linked mental retardation (TSPAN7), but little is known about MID1IP1 in the literature, except for an association with rye allergy . MoreovePossible limitations to the use of proteomic methods for the multiplex evaluation of IgE reactivity are the high costs and the need for specific expertise for the correct interpretation of results, which must always be correlated with the clinical condition of the individual patient. Future aspects are represented by the continuous evolution of these approaches, which have seen a tripling of the components available for diagnosis in the space of a few years, making it possible to increasingly refine the diagnostic precision and impact in daily clinical practice, even in such rare diseases.In conclusion, the new methods currently available to study IgE sensitization in vitro through the use of molecular components organized in micro– or macroarrays represent an important innovation in the diagnosis of even the rarest diseases, such as inborn errors with HIE. They enable us to not only verify whether circulating IgE have matured their affinity and are responsible for allergic sensitization, but also, if this is the case, to establish precisely the sensitizations present, even in cases in which the patient’s cognitive defect does not allow for correct allergological classification. This approach improves both the management of skin pathology and the quality of life of individual patients. Finally, investigations on genetic variants of risk for AD in patients with inborn errors associated with HIE could help to understand their susceptibility to manifest AD–like clinical symptoms .


Nectarines are perishable fruits grown in Southern Europe, valued for their sensorial properties. Chilling is used in the supply chain for Northern European consumers, while Southern European consumers can access fresh, locally grown fruit or cold-stored supermarket fruit. Cold storage and fruit ripening affect texture and flavour. Here a consumer survey and hedonic testing compared the appreciation of nectarines (cv. Big Top) in Italy and at two UK sites (n = 359). Fruit was at the commercial harvest stage, or stored at 1 °C or 5 °C for seven days, then sampled after two days’  or four days’ (second UK site) ambient recovery. In the consumer survey, the most important factors involved in purchase decision were ripeness, texture, colour, taste and price. Named varieties were more important to Italian than UK respondents, whilst ripeness, price, taste, blemishes, aroma, and ‘best before date’ were more important in the UK. In sensory analyses, fruits at the commercial harvest stage were preferred to those stored at 1 °C. Preference for the 5 °C stored peaches depended on recovery time. Distinct clusters of peach sensorial attributes were positively or negatively linked to hedonic rating. Factors important in purchase decisions did not affect hedonic rating in the tasting. Prunus, which includes peaches and nectarines, are rich in phytochemicals [Fresh fruit and vegetables are increasingly recognised as key components of a healthy diet ,2 with phemicals  and henchemicals . Howeverhemicals . PeachesChilling injury manifests as internal reddening and changes in texture, with a loss of flavour even before the appearance of visible symptoms . This isQuality is a key driver for peach purchase and consumption , of whicRipening has a key role in improving peach organoleptic quality, including changes in pigmentation, increasing sugar to acid ratio, softening, and an increase in aroma . FlavourTrends in food waste are similar across developed countries , althougIn this study, we compared preferences of the same nectarine cultivar “Big Top” which had been stored at different temperatures, at two stages of post-chilling recovery, between participants in southern Italy and the UK. Fruit tasting was carried out in university canteens to provide a food consumption context, which has been shown to be important for this type of study . The aimPrunus persica (L.) Batsch) ‘Big Top’ was used in this study. All peaches were within the FFV-26 recommendation for peaches and nectarines  [The melting flesh, semi-free stone yellow peach cultivar (code 2A)  as stateSamples were prepared for all tasting sessions in a food industry standard preparation area on the day of each consumer test. In Italy (site IT) stored samples were removed from cold storage and stored at 22 °C for two days prior to the consumer tests. On arrival in Cardiff  the peaches were divided into two groups, for testing immediately at one site (Site CF1) and at the second site (Site CF2) after further room temperature (20 °C) storage for two days.For each test site, on the day of tasting each peach was sliced to provide six segments of approximately equal size, avoiding the central stone , by reseWhen all the samples for each day of consumer testing were ready, they were transported in insulated food-grade boxes to the test sites. Products were served at room temperature of approximately 20 °C. Fruit for the site CF2 consumer test were left whole and unboxed on the tables in the food preparation area, clearly labelled with their codes. Site CF2 peaches were served as for site CF1 and site IT outlined above.Ethical approval for the study was obtained from the appropriate committees (Sta-1353 and SREC 1906-02) before commencement of the consumer test. The test was set up in three locations over the course of two days. On day one of the study, testing was undertaken in the main catering facility of a university situated in southeast Wales, UK (site CF1) and in the main hall of a university campus canteen in Calabria, Italy (site IT). On day three it took place in the main foyer of a second university in southeast Wales, UK (site CF2).The consumer test followed BS EN ISO (2020)  guidanceThe test consisted of a printed questionnaire  of closeParticipants then tasted the samples provided, in a forced randomised order indicated on the questionnaire to reduce positional bias or order effect , and ratStatistical analysis was conducted in R v3.6.3  using RStudio and packages car, dplyr, dunn.test, emmeans, gridExtra, ggmosaic, ggplot2, igraph, mvabund, multcomp, MuMIn, ordinal, RVAideMemoire, tidyr, tools, and vegan ,44.p value adjustment. A separate model checked for any effect of participant gender. The relationships between hedonic responses (liking), site, treatment, and the site–treatment interaction were modelled using a cumulative link mixed model for ordinal response variables. Respondent ID was included as a random term to account for non-independence. Pairwise tests were conducted based on least-squared means with Tukey p value adjustment.Using a binomial multivariate generalized linear model , relationships were modelled between the characteristics influencing the decision to purchase peaches, frequency of purchasing peaches, and national effects. Univariate tests were conducted using analysis of deviance with a step-down resampling procedure for A model averaging approach was used to assess the influence of (a) peach characteristics and (b) respondent characteristics on the hedonic responses (liking). Cumulative link mixed models were used due to the ordinal response variable. Respondent ID was a random term in all models. Two groups of models were constructed: peach characteristics in all possible combinations and personal characteristics in all possible combinations. An Akaike information criterion test (AICc) was used to rank models within each group, from best (lowest AICc) to worst (highest AICc). Within each group, models with ΔAICc < 6  were considered good . For thep values were adjusted for multiple testing using the Benjamini–Hochberg procedure [p value adjustment.The co-occurrence of pairs of peach characteristics was tested using Fisher’s exact tests. rocedure . Signifip values were adjusted for multiple testing using the Benjamini–Hochberg procedure [Finally, for every sensorial characteristic, the mean hedonic rating was calculated per respondent for (a) samples where that characteristic was reported present, and (b) samples where it was reported absent. The difference in hedonic ratings between the two cases was regressed against the respondent’s gender, country, frequency of purchasing peaches, and whether they considered particular characteristics important when purchasing peaches. rocedure .The selected test methodology is suitable for studying the impact of sensory characteristics of a product on degree of liking, independently of the product’s extrinsic characteristics such as brand, and studying the effect of a commercial or presentation variable, in this case transport . The metTest Environment: It was not possible to control completely the absence of communication  between the consumers, which would guarantee independent responses. Researchers did ask the consumers not to discuss the results with each other and were nearby to remind consumers of this during the test. Minimal interaction was observed.Test timing: The time of a test can influence its results. Sensory guidelines indicate that 10 a.m.–lunch is a suitable time for tasting as it corresponds to usual consumption time for products such as fruit, e.g., as a part of lunch or a snack. In our study, the timing was over the lunch period. However, we did ask participants to consume the peaches before rather than after their lunch, to minimise the impact of sensory overload or taste masking   with lower experience and income levels but higher education levels (compared to the population). However, the experiment was conducted outside term time and thus mainly included University staff and postgraduate students. In addition, this applied to all three sites, making them comparable. However, the samples may still not be representative of the whole populations in the two countries in terms of age and educational level.A total of 359 participants returned completed assessments for analysis across the three sites, 103 participants (28.69%) from site CF1, 101 (28.13%) from site CF2 and 155 (43.18%) from site IT. The total sample consisted of 174 males (48.47%), 179 females (49.86%) and six who did not disclose their gender (1.67%). Of the 359 participants, 165 (45.96%) were aged 18–29 years old whilst only 18 (5.01%) were over the age of 60, with nearly equal distribution of the remainder between the 30–39 , 40–49 , and 50–59  age groups .The majority of participants described their ethnicity as Caucasian , with less than 5% in each category of black , Asian , Hispanic ; other was used as an ethnicity description by only 1.95% (n = 7) of the participants, and 1.11% (n = 4) did not answer the question . A total2 = 84.3, p < 0.001), and most (73%) of the Italian respondents were in the frequent buyer category (>1 per week), while over 50% of UK buyers bought peaches less than once per month . At site IT, scores among the three treatments were remarkably consistent . Likewise at site CF1, testers significantly preferred the commercial harvest peaches over those stored at 1 °C , and also preferred them over those stored at 5 °C . At site CF2, in contrast, 5 °C stored peaches were significantly preferred to 1 °C stored peaches , but commercial harvest peaches were not significantly preferred to either storage treatment.There was significant interaction between site and treatment in predicting hedonic rating scores , but only marginally significantly different for the commercial harvest peaches . Sites CF1 and CF2 differed significantly only for the 5 °C storage treatment .There was no significant difference in preference for the three treatments between sites IT and CF2, even though the site CF2 tasting took place two days later than those at sites IT and CF1. Hedonic ratings at sites IT and CF1 were significantly different for both storage treatments  in all possible combinations, and person characteristics  in all possible combinations. AICc was calculated for each model to rank models within each group from best (lowest AICc) to worst (highest AICc). The peach-based models had consistently lower AICc than the person-based models A, indicaWithin each group, models with a ΔAICc < 6  were considered good . The numFor the peach-based models, a further subset was taken where ΔAICc < 2, i.e., up to twice the AICc of the best model in the group, which was considered indistinguishable from the best model  Table S. CoefficOf the forty-five pairwise combinations of characteristics, twenty-seven were significantly non-randomly associated with each other . Figure 4,1068 = 184.7, p = 0.001). Taking the characteristics individually, acidity, aroma, crunchiness, firmness, juiciness, and sweetness were significantly affected by the interaction between site and treatment , suggesting that this does not explain the cross-cultural differences noted. However, we showed that there was a significant difference in the factors influencing purchasing decisions between respondents in the two countries, and that the country of the respondent was more important than frequency of buying in influencing the importance of different purchase characteristics.As recently noted, fresh goods buyers’ purchasing behaviour follows the same established patterns as observed in consumers buying packed goods , and lesOf the twelve characters relating to purchasing decisions included here, four can be considered extrinsic characteristics , five intrinsic  and three relate to experience  . The finOne key difference between UK and Italian respondents was that those in Italy were more likely to consider variety name as important when purchasing, suggesting that peach varieties are not broadly known by consumers in the UK. Few studies have considered variety name as a factor in fruit purchase decisions. Selection by variety is very important in apples, where it has been linked to high frequency purchasers , howeverThe presence of blemishes was more important for respondents in the UK than in Italy. The high proportion of buyers who place a high value on unblemished fruit has been noted in previous studies ,19,51. TTrained panels are frequently used in the sensory evaluation of fruit, including peaches e.g., ,57. HoweComparing the two sites where peaches of equivalent ripeness were tasted (sites IT and CF1), it was found that commercial harvest stage peach samples were preferred at both sites. This perhaps suggests that attributes reported as related to freshness, such as taste, crispiness, juiciness, and flavour , were deCold storage is known to affect hedonic qualities in peaches, although these are not always detectable after only one week in storage ,57. At aOverall, the characteristics of peaches were more important compared to person-related characteristics for determining consumer appreciation at tasting, indicating a general agreement across respondents on the preferred characteristics in peaches. Amongst the person-related characteristics, site was the most important factor, perhaps due to the differences in peach ripeness between sites CF1 and CF2. In agreement with previous studies ,26, sweeAn important finding was that respondents’ stated preference for particular characteristics when purchasing peaches had no significant effect on their liking for the samples once tasted. For example, it might have been predicted that respondents who listed aroma as an important purchase criterion would more heavily penalise peach samples that they did not perceive as aromatic. The absence of such an effect suggests that there may be a disconnect between what a consumer expects to like and what they actually enjoy. This has implications for the supply chain, as the criteria used by a consumer at the point of purchase need to be differentiated from their perceptions at the point of consumption.Storage of peaches at higher temperatures has been shown to improve consumer appreciation  but is k