Long waiting times for elective surgery are common to many publicly funded health systems. Inefficiencies in referral systems in high-income countries are more pronounced than lower and middle-income countries. Primary care practitioners play a major role in determining which patients are referred to surgeon and might represent an opportunity to improve this situation. With conventional methods of referrals, surgery clinics are often overcrowded with non-surgical referrals and surgical patients experience longer waiting times as a consequence. Improving the quality of referral communications should lead to more timely access and better cost-effectiveness for elective surgical care. This review summarises the research evidence for effective interventions within the scope of primary-care referral methods in the surgical care pathway that might shorten waiting time for elective surgeries.We searched PubMed, EMBASE, SCOPUS, Web of Science and Cochrane Library databases in December-2019 to January-2020, for articles published after 2013. Eligibility criteria included major elective surgery lists of adult patients, excluding cancer related surgeries. Both randomised and non-randomised controlled studies were eligible. The quality of evidence was assessed using ROBINS-I, AMSTAR 2 and CASP, as appropriate to the study method used. The review presentation was limited to a narrative synthesis because of heterogeneity. The PROSPERO registration number is CRD42019158455.The electronic search yielded 7543 records. Finally, nine articles were considered as eligible after deduplication and full article screening. The eligible research varied widely in design, scope, reported outcomes and overall quality, with one randomised trial, two quasi-experimental studies, two longitudinal follow up studies, three systematic reviews and one observational study. All the six original articles were based on referral methods in high-income countries. The included research showed that patient triage and prioritisation at the referral stage improved timely access and increased the number of consultations of surgical patients in clinics.The available studies included a variety of interventions and were of medium to high quality researches. Managing patient referrals with proper triaging and prioritisation using structured referral formats is likely to be effective in health systems to shorten the waiting times for elective surgeries, specifically in high-income countries.The online version contains supplementary material available at 10.1186/s12913-021-06140-w. Awareness of growing waiting times in health care is not a recent phenomenon and the challenges of waiting times and waiting lists have been subject to a wide variety of health service research. Presentation with multiple comorbidities and complex clinical conditions are a direct result of the advancing biological age and increasing life expectancy of populations over recent decades , 2. As aReferral methods play a major role in providing appropriate care for patients in many health care systems. The quality and efficiency in referral systems in Lower and Middle Income Countries (LMIC) are often undermined due to system-related inefficiencies such as poor infrastructure, relevant materials/equipment and insufficient skilled human resource . DespiteWaiting times are a key performance indicator for many healthcare systems, used to encourage improved performance in healthcare institutions, with the aim of delivering high-quality care without unnecessary delay . Many paDespite increased funding in recent years, the demand for many elective surgeries exerts a substantial and growing challenge . FurtherWaiting lists are considered as a non-price rationing mechanism for coping with excess demand . A largeThis paper is based on one of the sub-reviews in a major systematic review which takes a holistic approach to summarise policies, strategies and interventions that might reduce long waiting times for elective surgeries. We have conducted and reported the review according to PRISMA statement . The PRIThe electronic databases of PubMed, EMBASE, SCOPUS, Web of Science and Cochrane Library were selected for the search. Search terms were decided using MeSH headings and keywords for the scope of the full review. Major types of elective surgeries were searched for broader inclusion. After performing pilot searches, a detailed search list was finalised. The search strategy combined with three sets of search terms. The searches were run from 14th December 2019 to 7th January 2020 to include relevant articles published from January 2014 to December 2019 without language restriction. The search strategy used for PUBMED is presented in Additional file Inclusion and exclusion criteria: All types of literature published as a full article were included if they reported eligible studies. This includes original research published in journals, reports, editorials and literature reviews from the healthcare sector, governments and other related sectors irrespective of the income level of the country of research. Where a study was based on outpatient departments, the referral intervention needed to be targeted at reducing the patient waiting time for elective surgery. Health system interventions are often tested in quasi-experimental studies, rather than randomised trials due to the complexity of the approaches being investigated and the diversity of outcome measurements. Therefore, we included a range of study designs, with a design-specific assessment of risk of bias . Our maiThe surgeries require penetration of a body cavity are considered as major surgeries and all surgeries of abdomen, chest or cranium are major surgeries. Minor surgeries are generally superficial and do not require the penetration of a body cavities . EligiblArticle selection and data extraction: As the first step, the title and the abstract of the retrieved citations were checked by one reviewer (DR) to select relevant articles. Articles that were potentially eligible based on their title or abstract were retrieved in full and assessed for eligibility and relevance. Each potentially eligible article was discussed with the second reviewer (MC) and an agreement was reached on inclusion or exclusion.Quality assessment: The validity of the results of any systematic review of a health systems intervention depends on the methods used in the included studies (which may have used different designs), rather than universal experimental criteria . ConsideThe article screening process is shown in the PRISMA flow diagram Fig.  and the Summary of included studies: Nine studies were included in this review. The characteristics of the included studies are summarised below and further details are given in Tables Starting with the original studies;Damani et al.  reportedGabbay et al.  reportedCoyle et al.  reportedLogvinov et al.  reportedDo et al.  reportedDiamant et al.  reportedBachelet et al.  reportedDamani et al.  reportedFinally, Ballini et al.  reportedRisk of bias in included studies: All included studies complied with the eligibility criteria for this review. The quality of evidence in the included articles was measured using the most appropriate of three tools: ROBINS-I Cochrane risk of bias tool (5 studies); AMSTAR 2, a critical appraisal tool for systematic reviews that include randomised or non-randomised studies of healthcare interventions, or both (3 systematic reviews) and CASP Critical appraisal skill programme for cohort studies (1 study).ROBINS-I: Of the five studies, a lower risk of bias was found for the randomised trial . The othOne study used randomisation of participants . The remOne of the three included reviews was rated as high-quality overall  and the A single study was assessed using the CASP checklist and the authors had reported important confounding factors for the study .Synthesis of results: Meta-analysis was not applicable for this review, because of the heterogeneity in study designs and variability in how the outcome of interest was measured. Instead, we did a meta-synthesis with narrative analysis. The Table Surgical referrals are considered as the interface between the referral provider (primary care practitioner) and the specialist (surgeon). The intended pathway for the patient to be referred to and what information needs to be recorded depends on the goal of the referral . CertainReferral guidelines should be sufficiently standardised to allow those involved in making decisions about the referred patient to judge the appropriateness of the referral and to undertake a more detailed objective analysis of their needs. Referral letters that were unclear or not sufficiently informative led to dissatisfaction at surgery clinics , 42. TraThe proper reporting and adequate clinical assessment of the patient at their last consultation prior to surgery is important for informing the decision about whether to perform or cancel the elective surgery  and can Patient prioritisation for referrals is primarily based on clinical parameters. On the other hand, including non-clinical factors in the patients’ decision regarding a particular surgeon/hospital or surgery schedule with an open referral system was more effective  than theWhere there are multiple queues for surgical lists at the same surgery clinic, targeted patient referrals have had variable impacts on waiting times across different specialists. Single Entry Model (SEM) indicates the consolidation of multiple queues into a single queue. The application SEM while pooling referrals enabled patients to see the next-available surgeon for their procedure and improved timeliness and efficient patient flow . The sysStreamlining the triage of patients during referrals with standardized referral templates and resources enhances timely access to surgical services . Good coAlthough the studies included in this review showed variable interventions with medium-high quality evidence, our findings have confirmed a consistently positive outcome with improvements to the timeliness of referrals at the primary care level in health systems of high-income countries.This review is focused solely on the effects of referral methods that were intended to reduce waiting times for elective surgeries and which were reported in research studies published since 2013. It does not provide insights into the effects on waiting times of other forms of referral management that might be implemented for other purposes or on the research literature from before 2014. However, most literature suggests that the primary intent of referral management should be on prioritising care for patients most in need. Our limiting of the search to articles published in 2014–2019 may mean that we have failed to include some studies that would provide useful information. However, the use of systematic reviews published in this time window provides some insight into the older literature and our focus on recent evidence should increase the applicability of the results for contemporary practice in health systems. Our inclusion of observational studies helps to triangulate the constructs of interventional studies but brings with it concerns about their higher risk of bias. Finally, we have not been able to present meta-analyses of the effects of the interventions or determine if publication bias has affected our conclusions.On the basis of the available evidence, managing referrals by using triage and prioritisation of surgical patients is likely to reduce the waiting times for elective surgeries, by avoiding the overcrowding of surgical clinics with non-surgical referrals. Explicit and standard referral guidelines are more likely to be effective in selecting potential surgical patients if a structured referral formats are being used. In addition, using non-clinical information on a patient’s preferences for scheduling and switching their surgeon should also increase the timeliness of elective surgical care, and timeliness of patient flow may be increased with single entry models (SEM) for elective surgical services. Implementation of these interventions should mean that primary care practitioners and surgeons experience a more streamlined approach to elective care referrals.In summary, this review has identified some interventions that could be implemented in the referral process for an adult elective surgery that might shorten waiting times and reduce the length of waiting lists specifically for high-income countries. However, more research is needed, ideally in the form of randomised trials to determine the effects of these interventions more precisely and to inform decisions around their cost-effectiveness. This may be especially important given the impact of COVID-19 on elective surgery waiting lists in many countries, and the depletion of resources for routine health care.Additional file 1.Additional file 2.Additional file 3.


The differential sensitivity of neutrons towards hydrogen and deuterium makes them a unique probe to study the structure and behaviour of asymmetric membranes. Molecular-level understanding of the interactions between antimicrobials and the Gram-negative OM provides valuable insights that can aid drug development and broaden our knowledge of this critically important biological barrier.The use of neutrons as a scattering probe to investigate biological membranes has steadily grown in the past three decades, shedding light on the structure and behaviour of this ubiquitous and fundamental biological barrier. Meanwhile, the rise of antibiotic resistance has catalysed a renewed interest in understanding the mechanisms underlying the dynamics of antibiotics interaction with the bacterial cell envelope. It is widely recognised that the key reason behind the remarkable success of Gram-negative pathogens in developing antibiotic resistance lies in the effectiveness of their outer membrane (OM) in defending the cell from antibacterial compounds. Critical to its function, the highly asymmetric lipid distribution between the inner and outer bilayer leaflets of the OM, adds an extra level of complexity to the study of this crucial defence barrier. Here we review the opportunities offered by neutron scattering techniques, in particular reflectometry, to provide structural information on the interactions of antimicrobials with  Lipid asymmetry is preserved by a specialised ensemble of proteins which ensure that LPS molecules are delivered to, and maintained within the outer bilayer leaflet, whilst misplaced PL are segregated to the periplasmic side of the bilayer or returned to the cytoplasmic membrane [Within the list of the highest-priority drug resistant pathogens highlighted by a recent WHO report, Gram-negative bacteria outnumber Gram-positives seven to two, with the top three critical-priority organisms all belonging to the Gram-negative class . The keymembrane . The unimembrane . Whilst The properties of the LPS layer render the OM up to two orders of magnitude less permeable to hydrophobic molecules in comparison with canonical PL bilayers ,7. This mcr-1 plasmid which codes for LPS modifications that inhibit their activity [The increasingly difficult challenge of reaching targets hidden behind the OM permeability barrier has driven efforts in antibiotic research towards targeting the OM itself ,17. The activity .in vitro to create models that mimic the OM structure and composition. These enable structural analysis using techniques such as scattering, under controlled conditions [in vitro models of the OM has provided a new platform for the investigation of the properties and behaviour of this crucial biological barrier. In particular, the use of specular neutron reflectometry (NR) offered valuable insights into the structure–function relationship of different LPS chemotypes and the effects of antimicrobials interacting with OM models [in vitro models of the OM and their interaction with antimicrobial compounds. We outline the advantages of neutron scattering in the context of structural techniques amenable to study the interaction of antibiotics with the bacterial surface, together with the current limitations that this approach entails.A detailed understanding of the OM biophysical properties and its interactions with antibacterial molecules is therefore critical to devise novel entry routes across the first defence line of multiple life-threatening pathogens. However, obtaining high-resolution structural information about the OM directly from the surface of bacteria is a challenging task. An alternative approach to the problem consists of reconstituting OM components nditions . In the M models . Here weStructural investigations of the OM began with the introduction in the late 1950's of electron microscopy imaging of intact and sectioned bacteria which produced the first visual evidence of the double membrane envelope of Gram-negative organisms . Since tin vivo [The advent of atomic force microscopy (AFM) paved the way for the investigation of the properties and behaviour of bacterial surfaces under conditions that are much closer to those relevant to antibiotic action in vivo . AFM hasin vivo ,38 and cin vivo . Additioin vivo . To datein vivo ,41,42. AOften used in combination with AFM, fluorescence microscopy has been extensively used to investigate the bacterial surface and its interaction with antibiotic molecules . Fluoresin vitro model membranes reconstituted using synthetic or purified membrane components. The appeal and convenience of model membranes is not only limited to their reduced complexity but extends to their ease of preparation and versatility. Under the right conditions, lipids self-assemble into vesicles amenable for solution studies which can later be fused onto hydrophilic surfaces to form planar lipid bilayers suitable to be investigated by several biophysical approaches [in vitro OM model bilayers via vesicle fusion has been proposed by Hsia et al. [in vitro model of the OM [The technical challenges posed by the complexity of natural cell membranes are often overcome by accepting the compromise of performing biophysical experiments on proaches . Althougproaches , vesiclea et al.  who fusea et al. . This mef the OM   in the buffer bathing the bilayer. This enables measurement of multiple sets of reflectivity data at different aqueous isotopic compositions, which can later be simultaneously fit to a single model of the interface  LPS monolayers at the air/water interface, (ii) solid supported asymmetric bilayers and (iii) floating asymmetric bilayers (In the last ten years, the combination of NR and bilayers .As well as constituting the starting point for the assembly of asymmetric PL/LPS bilayers, Langmuir monolayers of LPS, spread at the air/water interface, provide the simplest mimic of the outer leaflet of the OM . NR has Sequential Langmuir–Blodgett and Schaefer depositions onto a solid substrate of monolayers of deuterium labelled PL and hydrogenous LPS yield isotopically asymmetric PL/LPS bilayers . Unlike in vitro OM models [in vivo bactericidal activity against PmB-resistant Gram-negative pathogens [Due to their unique interaction with the OM, antibiotics of the polymyxin family have been the subject of several studies on M models ,28,29. Hathogens . Using aCombining a functionalised gold substrate with the asymmetric model described above enables a third type of model with the PL/LPS bilayer floating on a ∼20 Å thick water cushion separating it from the solid interface . Buildinin vitro and, more recently, in vivo [in vitro OM models has shown a clear potential for the investigation of LPS and the asymmetric OM structure [mcr-1 plasmid, have recently been shown to render the OM more susceptible to hydrophobic antibiotics and osmotic shock [in vitro [Access to neutrons remains limited to large scale facilities, however, thanks to their distinctive properties, their use to study biological systems, provides unique access to structural information, both  in vivo ,75. Heretructure ,56,57,68tructure  and drugtructure . The majtructure . Work toic shock . Thus, fic shock  could heic shock . The intic shock ,78. Builin vitro ,73, the in vitro . Reconstin vitro . Finallyin vitro . Given iin vitro . These aNeutron reflection is a powerful tool to study the structure of lipid bilayers and unique in its ability to reveal the asymmetric distribution of molecules across membranes. Lipid asymmetry is a crucial feature of the Gram-negative bacterial outer membrane, an important biological barrier that plays a fundamental role in the development of antibiotic resistance by some of the most clinically relevant pathogenic bacteria.in vitro asymmetric bilayers, that mimic the structure of the outer membrane, is progressively gaining traction as a tool for the structural characterisation of antibiotic-membrane interactions.The combination of neutron reflectometry and The development of increasingly realistic outer membrane models, together with further advance and implementation of techniques complementary to NR, has the potential to probe aspects of membrane biology inaccessible by other methods.


For example, a role of metabolism in reactive oxygen species production in response to pathogens has been long known. Furthermore, macrophage transition from an anti to a pro-inflammatory phenotype seems to be at least partially mediated metabolic reprogramming from fatty acid oxidation to glycolysis. Recent studies have also shown metabolic plasticity during long-term responses to vaccine formulations of individual pathogen-associated molecular patterns. The capacity of macrophages to be trained, an emerging concept describing the “learning” capacity of innate immune cells, seems to be also underpinned by metabolic switches. This Research Topic aims to shed light on our current understanding of the role of metabolism in macrophage biology and versatility focusing on the immune response to infectious agents.Macrophages are key components of the immune response to infectious organisms, with a dual role eliminating infectious agents by phagocytosis while mediating defensive and inflammatory pathways. On the other hand, macrophages are also involved in the late response to infections by heavily contributing to tissue debris clearance and “wound” repair. The importance of the metabolic status of these immune cells on their responses has been overlooked for decades. However, recent advances in metagenomics and high-throughput metabolic profiling have partially unveiled the role of metabolism in macrophage responses. New evidence suggests that the metabolic status and specific metabolic pathways are instrumental for the modulation of macrophage responses to the ever-changing environment that these cells face Ramond et al., discuss the ability of some bacterial pathogens to manipulate macrophage metabolic programs to survive intracellularly and how mitochondria could play a paramount role by regulating cellular metabolism and immune responses against these pathogens. Although studies addressing this interplay need further refinement to reflect in vivo conditions, they encourage the implementation of host-directed therapies for clinical infections where manipulation of metabolic responses through this organelle can help to resolve infections. More specifically, to clarify the observed susceptibility to infection of bone marrow macrophages (BMMs) from CBA mice by two species of Leishmania, Bezerra de Menezes et al., performed a differential proteomic analysis of infected macrophages and found that host responses related to oxidative stress and iron metabolism might dictate the intracellular fate of this pathogen.Two of the articles in the topic are specifically dedicated to the role of macrophage metabolism during infection. In a review, Diaz-Bulnes et al., the authors address the phenomenon of metabolic rewiring in macrophage activation, and how those metabolic modifications allow those cells to adapt to changes in the environment (such as hypoxia or oxidative stress) maintaining their pro-inflammatory and anti-microbial activities. This response is coordinated by different metabolic mechanisms. In this review, they illustrate the relation of the metabolic and phenotypic plasticity of macrophages with hypoxia and endoplasmic reticulum stress, and how they can relate to human pathology and chronic inflammation. Two different reviews in the topic summarized, using different point of views, the relevance of macrophage metabolic activation with the focus on metabolic disease. Daemen and Schilling, highlight the multiple and relevant connections between obesity and macrophage metabolic activation. The article initially covers macrophage dynamics and diversity in liver and adipose tissue, both crucial areas for the study of metabolic diseases. The review also summarizes the available knowledge regarding the role of macrophage metabolic regulation for metabolic diseases and, in particular, obesity, also addressing the importance of niche determinants for this metabolic tuning and for the overall function of macrophages. Furthermore, Daemen and Schilling describe future challenges in the topic and suggest new therapeutic possibilities based on macrophage targeting in order to treat metabolic disorders. Meanwhile, the role of lipids in the metabolic reprogramming of macrophages is reviewed by Batista-Gonzalez et al.. They summarize the available knowledge regarding the role of lipids in macrophage plasticity from the importance of the differential fatty acid metabolism in the dichotomic, in vitro endorsed, M1 (pro-inflammatory) to M2 (anti-inflammatory) transition, down to the role of lipids on macrophage activation. In this review, the authors summarize new research challenging the old consensus that defines M1 as aerobic glycolytic cells and fatty acid oxidation as characteristic of M2 macrophages suggesting a more dynamic metabolic reprogramming. Finally, they suggest possible therapeutic interventions targeting lipid metabolism in order to prevent the activation of macrophages. Also in relation to lipids, original research by Sohrabi et al., highlight the relationship of trained innate immune responses to the reprogramming of different cellular metabolic pathways like those related to the synthesis of cholesterol and fatty acids. It is specifically explored the role of Liver Receptor X (LXR), a key regulator of these pathways, whose activation promotes a proinflammatory trained immunity phenotype in human monocytes.Different articles in the topic are focused on the importance of metabolic adjustments for macrophage activation. In a review by Erkelens et al. experimentally confirmed previous observations of the effect of Vitamin A in the modulation of inflammation within the gut. In particular, they showed that the vitamin favors the anti-inflammatory state of intestinal tissue resident macrophages within the gut. Furthermore, the article shows that Vitamin A increases Dectin-A expression levels in macrophages under homeostatic conditions. The authors further show that Dectin-A induction provokes the release of pro-inflammatory cytokines by macrophages. Based on these results, Erkelens et al. suggest that Dectin-A-induced signaling might provide a switch from an anti-inflammatory to a pro-inflammatory state of the macrophage when needed. Finally, another original research article is presented by Yang and Ding, demonstrating that MEK1/2 inhibitors unravel the interferon response via interferon regulatory factor 1 (Irf1) signaling in macrophages. They pinpoint a deficiency in Irf1 that abolishes the interferon response and prevents macrophages from reprogramming into an immunostimulatory phenotype. A MEK1/2 inhibitor enables IRF1-mediated interferon signature response in macrophages, while a combination of inhibitor and agonist of Irf1 have potential as therapy.In an original research article focused on the interface of nutrition and macrophage activation, The manuscripts included in this Special Issue highlight and expand our understanding of the adaptative capacity of innate immune cells in response to inflammatory signals and pinpoint the critical importance that metabolic rearrangements play in the response kinetics of these first responders of the immune system. They also highlight the variability and intricacies of these metabolic adaptations, shedding light into novel targets of intervention in order to fine tune the response of macrophages to different pathological conditions.HR, RP-R, JL, MM, and JA edited the topic. HR, RP-R, and JL wrote the manuscript. JA and MM reviewed and corrected the manuscript.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.


Rationale: High-intensity focused ultrasound (HIFU) therapy represents a noninvasive surgical approach to treat uterine fibroids. The operation of HIFU therapy relies on the information provided by medical images. In current HIFU therapy, all operations such as positioning of the lesion in magnetic resonance (MR) and ultrasound (US) images are manually performed by specifically trained doctors. Manual processing is an important limitation of the efficiency of HIFU therapy. In this paper, we aim to provide an automatic and accurate image guidance system, intelligent diagnosis, and treatment strategy for HIFU therapy by combining multimodality information.Methods: In intelligent HIFU therapy, medical information and treatment strategy are automatically processed and generated by a real-time image guidance system. The system comprises a novel multistage deep convolutional neural network for preoperative diagnosis and a nonrigid US lesion tracking procedure for HIFU intraoperative image-assisted treatment. In the process of intelligent therapy, the treatment area is determined from the autogenerated lesion area. Based on the autodetected treatment area, the HIFU foci are distributed automatically according to the treatment strategy. Moreover, an image-based unexpected movement warning and other physiological monitoring are used during the intelligent treatment procedure for safety assurance.Results: In the experiment, we integrated the intelligent treatment system on a commercial HIFU treatment device, and eight clinical experiments were performed. In the clinical validation, eight randomly selected clinical cases were used to verify the feasibility of the system. The results of the quantitative experiment indicated that our intelligent system met the HIFU clinical tracking accuracy and speed requirements. Moreover, the results of simulated repeated experiments confirmed that the autodistributed HIFU focus reached the level of intermediate clinical doctors. Operations performed by junior- or middle-level operators with the assistance of the proposed system can reach the level of operation performed by senior doctors. Various experiments prove that our proposed intelligent HIFU therapy process is feasible for treating common uterine fibroid cases.Conclusion: We propose an intelligent HIFU therapy for uterine fibroid which integrates multiple medical information processing procedures. The experiment results demonstrated that the proposed procedures and methods can achieve monitored and automatic HIFU diagnosis and treatment. This research provides a possibility for intelligent and automatic noninvasive therapy for uterine fibroid. Uterine leiomyomas (fibroids or myomas), the most common tumor in the female reproductive system, have a high prevalence (77%) and can cause reproductive dysfunction et al. proposed an integrated diagnosis and therapeutic system using intraoperative 5-aminolevulinic acid-induced fluorescence-guided robotic laser ablation for precision neurosurgery Some studies of image-assisted surgery have focused on intraoperative image-guided therapy  et al. presented a visual tracking method for coagulation lesions in a moving target. They applied coagulation lesion tracking to a special US-guided robotic HIFU system, and motion compensation was investigated using a moving kidney phantom based on respiratory motion data. Their study focused on the use of HIFU systems for lesion identification and included model experiments et al. used a triangulation method and three echoing signals  et al. applied a US sensor to estimate the motion of a moving organ  et al. used the above method for fibroid segmentation with a sensitivity of 84% et al. used the fuzzy c-means algorithm and morphological analyses to segment MR images of uterine fibroids in two steps and obtained an average Dice similarity coefficient (DSC) of 79.9% Noninvasive HIFU therapy has been widely studied for different lesion treatments. SeoIn this research, we propose a multi-information fused automatic real-time image-guided diagnosis and treatment system for HIFU therapy. The system framework includes automatic uterine fibroid diagnostics; intraoperative US lesion tracking and intelligent HIFU focus distribution strategies. All image information involved in HIFU treatment is automatically processed and fused. The strategy of operation is generated based on the fused image information.The proposed intelligent theranostics system is illustrated in Figure Our strategy for intelligent noninvasive HIFU therapy is shown in Figure. 2. Images information is extracted by the intelligent diagnostics and planning processing from the involved medical devices, including MR, US and HIFU devices. All this information is combined into a global information set and used to determine the treatment strategy. The HIFU probe treats the lesion area based on this information. The treatment strategy is monitored by medical images and physiological indicators. Additionally, the doctor further confirms and monitors all information during treatment to ensure safety.To initialize the contour of the intraoperative lesion, accurate preoperative image segmentation at both the structural and sequence scales is needed. Intraoperative initialization requires both the structural information from the image and accurate spatial position correspondence. Therefore, we fuse and transfer the structure and position information from the preoperative image to the intraoperative stage. In the intraoperative stage, lesion information obtained from image processing is used as the primary treatment basis. The automatically identified lesion area is treated with the confirmation of the doctor. Warning and handling of accidents are also necessary. However, movement of the patient when she feels uncomfortable can lead to unexpected focus shifts, and the treatment process needs to be stopped when such events occur. Warnings about large movements are also sent. Finally, our intelligent HIFU therapy mode combines automatic image information processing confirmed by the doctor.Joint preoperative and intraoperative information are combined with general lesion information for treatment. During the processing of medical images in HIFU therapy, the MR and US images are processed separately and combined with the results of the position information acquired from the HIFU device. The proposed procedure has four associated steps:Step I: Preoperative diagnosis and planning. A novel multistage convolutional neural network is proposed to segment the uterine fibroids in the MR image for preoperative diagnosis. Three different parts of the network are connected to extract image features from the sequence scale, structure scale and pixel scale. Different stages of the network reduce the possibility of fibroid misjudgment by extracting effective lesion information.Step II: Contour initialization of the intraoperative lesion region. The rigid registration is completed based on the spatial information of the HIFU device and the MR device. Then, the segmentation results obtained from preoperative MR images are used as the initial contour for subsequent tracking in the intraoperative US image.Step III: Intraoperative lesion tracking. An improved morphological active contour without edges (MACWE) Step IV: Based on the auto-outlined lesion area, the HIFU foci are automatically distributed in the autoidentified lesion area. An image-based auto unexpected movement warning and human monitoring are used to guarantee safety.To achieve automatic preoperative image diagnosis, we propose a multistage convolutional neural network for MR image segmentation and a rigid MR-US registration procedure.The proposed segmentation network consists of three components as illustrated in Figure After the uterine fibroids are recognized and segmented from the MR image, the contour result is combined with the MR device's spatial information and the HIFU device's position information. The fused information, which consists of preoperative lesion contour and position information, is used to initialize the US video.P p U, from the positioning system. Similarly, the position information of the human body in the preoperative MR image, P p M, can be obtained. The MR segmentation results can be set as the initial contour for the US lesion tracking relay on P p U and P p M. Thus, accurate MR segmentation is important.The initial contour of the intraoperative lesion tracking is obtained from preoperative MR segmentation results, spatial information of the HIFU device and intraoperative US video. The position of the patient during the operation is fixed. Thus, we can obtain the corresponding spatial information of the patient's body in the US image, defined as In the semantic segmentation task, the pixels are not equal in complexity, and the background or simple part accounts for most of the image In classification tasks, an excessively deep classification network consumes more computer memory and results in degradation 1 is from the weighted binary cross-entropy for the classification network The model contains three convolution blocks and two fully connected blocks. Each convolution block consists of convolution, batch normalization (BN) and max-pooling layers. The sigmoid classifier, as the final output layer, produces the probability distribution of tumor judgment and maps the real number from 0 to 1. To train the classification network, we use an objective function derived from binary cross-entropy, which is suitable for binary classification with the use of the sigmoid. Because the ratio of the positive class and the negative class is imbalanced by 1:2, we add a weight to each class on the loss function. The loss function LIn clinical applications, false-negative errors of tumor judgment are inevitable and more severe than false-positive errors for diagnosis. To avoid the occurrence of false-negative errors in actual fibroid sections, the simple fibroid judgment is replaced by fibroid section judgment. The image before the first image with fibroids is defined as the beginning of the section, and the image after the last image with fibroids is defined at the end of the section. This procedure ensures that the true region of the lesion is covered as much as possible. The procedure of fibroid section judgment is shown in Figure The HIFU clinical dataset used in the paper was provided by the National Engineering Research Center of Ultrasound Medicine. The MR dataset was collected from six different hospitals and five different MR scanners . The multisource causes variation in the image morphology. Occasionally, some regions that are extremely similar to uterine fibroids in grayscale and shape appear in the MR image. Such interference regions lead to misjudgment in the segmentation result. Conventional mathematical methods have some limitations for processing images with large morphological differences, such as a lack of generality. To solve the problems of data difference and image internal interference, we draw on the idea of domain adaptation and instance normalization The proposed physiological structure processing network aims to eliminate the disadvantages and reduce the adverse effects of image diversity, as illustrated in Figure x scale. Different from the previous two convolution layers, the third convolution layer extracts the grayscale value by the dot product value of the convolution kernel and the image matrix. Each value expresses the image grayscale feature on the x scale and constitutes the initial image in the gray feature tensor. To prevent calculation errors caused by a zero denominator in the dividing operation, we add a unit vector to the feature tensor and produce the gray feature tensor. The last step of the preprocessing network is to divide the image by the gray feature tensor and obtain the preprocessed image.As shown in Figure The architecture of the segmentation baseline consists of two parts, the encoder and decoder paths. Each path contains five blocks . To train the fibroid segmentation CNN model, we use the sigmoid classifier to semantically segment the image and determine the probability of whether the pixel belongs to the uterine fibroids.2 for the segmentation network is defined asThe loss function we use is the DSC, which is optimized using the adaptive-moment-estimation (Adam) method n∈Y and the ground truth binary region xn∈X. The DSC considers only the contribution of the shared target region of individual similarity and ignores the shared context. The DSC loss function has the advantage of minor calculations and is suitable for uterine fibroid segmentation where the sums run over the N voxels of the predicted binary segmentation region yThe networks of stage II and stage III are trained together, and multiple stages are optimized together end-to-end. With forward propagation, the weight in both network parts can be adjusted simultaneously. The MR images in the experiments were resized to 256 × 256 in two dimensions. Each selected case included the following characteristics: 1) preoperative MR images of signal and multiple cases, 2) MR series collected by different MRI machines, 3) manual delineation of the fibroid contour by two experienced obstetricians on the source images. Finally, 320 cases were gathered with permission, and each case contained 25 images. To train the CNN model, 320 cases were randomly divided into three sets comprising 250 cases as the training dataset, 30 cases as the validation dataset, and 40 cases as the testing dataset. The model parameters were saved automatically when the highest validation accuracy was obtained. All CNN models were trained and tested on a workstation outfitted with an NVIDIA GeForce 1080 Ti Graphics processing Unit (GPU). We used an adaptive-moment estimation with a batch size of 4 and epochs of 200. During training, the learning rate was set as damped with an initial value of 0.01 and an attenuation rate of 0.9 in 1,000 global steps.The demands of image-guided HIFU therapy are real-time processing and accurate lesion contour. The deformation of the contour is completed by MACWE. The entire framework of intraoperative US processing is illustrated in Figure Before the lesion tracking task, the original US image needs to be preprocessed. The focused ultrasound probe of the HIFU device is mounted in a sink that causes an echo in the US image. Unlike conventional ultrasound examination with the ultrasonic couplant, the HIFU ultrasound probe cannot be close to the patient's skin. The gap between the probe and the skin introduces noise in the ultrasound image. Thus, denoising and contrast enhancement of the image are required. In contrast to normal image noise, the echo speckle in the US image generally has a shape and a larger structure. We use a morphologically similar operation instead of ordinary image filtering.In u isAn interfering structure that can easily be identified in HIFU ultrasound images is the bladder. The morphology of the bladder is similar to that of fibroids in US images due to the internal liquid. To improve image contrast and reduce disturbance, especially in the lesion, we use an efficient linear conversion to highlight the lesion and remove the bladder. The entire preprocessing function of an input US frame is the initial contour from MR segmentation results.where κ is the kernel of similar operation andIn many clinical cases, the lesion area in the intraoperative ultrasound is incomplete due to continuous tissue deformation, resulting in significant error that we call 'overflow' in the contours produced by the MACWE. To solve this problem, we added an incomplete area processing procedure in the level set generation step. The procedure detects the proportion of the level set. If most values are positive (inclined gradients) in the level set, the contours are essentially aligned with the real lesion area. The incomplete area processing is defined asBased on the morphological active contours without edges (MACWE) model, we propose an improved method to specifically target the lesion area in HIFU ultrasound images. The basis points for the clinician to judge the lesion area in the ultrasound image are relative position, regional grayscale, and regional shape. Generally, the model-based image segmentation method requires accurate initialization of the contour, which is solved by the previous rigid MR-US registration. Thus, mainly the shape and grayscale characteristics of the lesion are considered.C and image I, the function In 1 and λ2 are the parameters of each term, and and  are the mean values inside and outside the contour of Iwhere μ, v, λI n(p) u, the current hypersurface in I n(p) u is defined as the level set 1/2 of a binary embedding function un: Z → {0, 1} based on the initialized contour C n init The traditional single iterative morphological ACWE algorithm can be represented asFor a given preprocessed US frame dD and dE are the erosion and dilation operations, respectively. SI◦IS is the compound morphological operation.where 1 and λ2 as the unit values. Finally, the algorithm is represented asTo automate the process, we set image feature parameters μ, v, λSI◦IS consist of four discrete segments 𝒦3= {k 1 3, k2 3, k3 3, k4 3}. The operators are the length of three pixels and involve all shapes of the edge. This form of the operator has a good effect on natural image segmentation. The targets in the natural image usually have a clear relationship between structure and semantics. However, the uterine fibroids in the HIFU US image generally have internal structures. This kind of character causes grayscale differences inside the lesion. During the operation, the clinician mainly focuses on the outer contour of the uterine fibroids rather than the internal structure. Although existing discrete operators have better sensitivity to subtle structures, the small discrete operators obtain undesirable contours and make some mistakes, especially in HIFU ultrasound images with low signal noise ratio (SNR). To obtain the external contour of the lesion, we replace the 𝒦3 morphological operator by a single large segment 𝒦15= {k15} with a length of fifteen pixels. For larger-sized operators, smaller interference structures in the image can be removed during morphological operations. Additionally, the computational time of a single operator is less than four operators. The operator 𝒦15 and the effect of 𝒦15 on fine structure removal are shown in Figure In the previous algorithm, the morphological operations in The entire algorithm flow, including morphological operations, template matching, and mutual information, involves matrix operations that can be calculated by the GPU efficiently. We implement all matrix operations in the GPU by processing each pixel in the US image with one corresponding GPU thread. The parallel computing speed achieves triple or quadruple the improvement compared with central processing unit (CPU) processing and satisfies the real-time guidance requirement in the clinic.In HIFU therapy, the patient is requested to lie prone on the HIFU device and avoid body movement. However, displacement during surgery is inevitable. Therefore, monitoring of large movement and timely interruption of treatment are necessary for intelligent HIFU treatment. To detect large movement in the current frame, mutual information (MI) detection is performed every 10 frames in the US video. For ultrasound video with 30 FPS, every-ten-frame detection implies a one-third second monitoring period to meet clinical monitoring requirements. The formula I and template u as random variables with samples i, li, iu = a…b are defined as follows:When a large movement is detected, the alarm message is sent and forces the treatment to stop until the image is stable. To maintain the tracking accuracy of lesions during large movement, we apply a template-matching method of normalized cross-correlation (NCC) var(x) and E(x) represent the empirical mean and variance for vectors x∈Rb-a. After P is confirmed, the vector from P to the centroid of contour Cn is considered a moving force m with a parameter to α next frames.where Finally, the last step of tracking is defined asIn HIFU therapy, the focused ultrasound probe applies energy to the lesion area. To accurately automate the focus distribution based on the processed image, the ultrasound probe images the layered lesion area under the control of a controller. The ultrasound probes move at the same interval in the same direction, which spatially produces a layered ultrasound image and the entire abdomen. Finally, the 2D distribution in each layer is combined into a 3D focus distribution. Our method tracks the lesion area in the ultrasound image at the current location. Therefore, the proposed intelligent treatment plan is implemented separately in each layer of the ultrasound video.In the proposed intelligent HIFU treatment process, we deploy the focus distribution based on the real-time lesion contour. According to clinical experience, we choose one quarter of the center of the lesion contour as the intelligent treatment area. In this treatment area, the focus of the focused ultrasound is evenly distributed at 0.5 cm. The final focus is three-dimensionally distributed in the lesion based on the automatically identified lesion contour. All movements and focus distribution are intelligently completed based on fused information. The proposed image-based HIFU focus distributing strategy is shown in Figure The experiment includes three parts: preoperative MR segmentation accuracy evaluation, intraoperative US lesion area tracking accuracy evaluation, and systematic clinical experiments. For the evaluation of intraoperative ultrasound lesion tracking, we first evaluated the accuracy and processing time of the lesion tracking method. Afterward, we evaluated the improvements in our proposed methods and procedures. Finally, we integrated our system with the clinical system to provide image-guided information to doctors in 8 clinical HIFU surgeries. To verify the feasibility of the intelligent HIFU therapy, we additionally evaluated our image-guided system to help improve the HIFU surgical learning curve.The preoperative diagnostic information includes preoperative MR image segmentation and MR-US rigid registration. Because the accuracy of MR-US rigid registration is based on the accuracy of the HIFU and MR devices, we mainly evaluated the performance of the multistage MR segmentation network.We present a series of experiments to analyze the impact of each of the main contributions and to justify the choices made in designing the proposed end-to-end segmentation network of uterine fibroids and the US tracking procedures. To quantitatively evaluate the performance of the present method, the DCS and Jaccard index (JI) We calculated the average values for DCS and JI in the testing dataset with the baseline (segNet) and the proposed structure processing network (Str- segNet) and entire network (Seq-Str-segNet) to validate the improvement of the structure scale processing stage and sequence scale processing. The quantitative comparison of the evaluation metric for each method is reported in Table Figure In the real-time US lesion tracking experiment, the accuracy of real-time lesion tracking procedures and the calculating efficiency for clinical HIFU guidance were evaluated. The computing platform included a CPU (Intel (R) Core (TM) i7-4790K) and a GPU (NVIDIA GeForce 1080 Ti). The corresponding US videos with HIFU device information were provided by the National Engineering Research Center of Ultrasound Medicine . We randomly selected 10 typical and clinical cases as the experimental US image data.15 for MACWE instead of the traditional discrete morphological operator 𝒦3. To verify the rationality of the change, we compare the accuracy and efficiency of 𝒦15 and 𝒦3. The tracking results for two examples of 𝒦15 and 𝒦3 are shown in Figure 15, and the third line is the tracking result of 𝒦3. The results show that the contour of 𝒦15 is more accurate than that of 𝒦3 and has better robustness. To quantitatively evaluate the results of two kernels, we used DSC, the Hausdorff distance (HD in mm) 15 provides better results comprehensively. In addition, we tested the computational efficiency of the two kernels on the same computing platform, as shown in Table 15 is twice as fast as 𝒦3 on the GPU and more than three times faster on the CPU.We used two 20-second US videos manually labeled by the doctor (as the gold standard) to quantitatively evaluate the improved MACWE. The content was used to assess the accuracy of lesion tracking and the calculation efficiency. In the previous description, we proposed a single morphological operator 𝒦We also evaluated the proposed large displacement detection and incomplete area processing method. First, we verified the performance of large displacement detection as shown in Figure To assess the feasibility of the proposed intelligent treatment strategy, we integrated the system with a commercial HIFU system  and performed clinical surgical validation. In the clinical trial, the operations were performed under the joint guidance and assistance of preoperative MR segmentation and intraoperative real-time ultrasound lesion contours. In the experiment, the HIFU focus was distributed according to the proposed intelligent treatment procedure. One doctor monitored and confirmed the position of the focus through the real-time US image. Additionally, another doctor simulated a traditional HIFU treatment procedure on the same patient's data, and the traditional HIFU focus distribution was used as the gold standard. The surgical side of the surgical scene is shown in Figure A total of 8 clinical experiments were performed, and clinical information of the surgical procedure was recorded. Each case was randomly selected and included preoperative and intraoperative images, spatial coordinates and metrology of the intraoperative focused ultrasound focus, postoperative contrast-enhanced ultrasound, and MR images. An image-guided treatment process is shown in Movie 2 . During Furthermore, we compared the focus distributions simulated generated by different operators. We used the focus performed by the senior doctor  as the gold standard, including the distribution and number of foci. Foci automatically distributed by the proposed intelligent strategy were recorded (Auto). The procedure was then simulated by both an intermediate operator  and by a junior operator  with image guidance and without image guidance. Two examples of the experimental results are shown in Figure As shown in Figure We also evaluated the learning curves of two trainees with similar levels of understanding of HIFU and repeated eight cases in the same order with or without image guidance. Figure In HIFU therapy, automatic, efficient, and precise processing of different medical information is vital. In this paper, we propose an intelligent and real-time HIFU theranostics strategy for uterine fibroids and applied this strategy in clinical treatment. Different medical images were processed and integrated by several novel methods and procedures. The treatment strategy generated valid HIFU focus distributions based on auto-obtained joint medical information. For preoperative diagnosis, we propose a novel multistage neural network for MR image segmentation. Different scale information of the MR image is screened by the network. The segmentation results of the MR images are transmitted to the treatment processes as the diagnostic information. In the intraoperative part, we integrate the spatial positioning information of the HIFU device with real-time processing of intraoperative ultrasound to provide image guidance for the doctor. In addition, we propose an improved MACWE method and multiple procedures to address clinical image problems. An image-based unexpected movement warning and other physiological monitoring options are used during the intelligent treatment procedure for safety assurance. Finally, the treatment strategy is generated from image-based intraoperative information.To evaluate the efficiency of our image guidance system, we evaluated preoperative MR image segmentation accuracy and intraoperative ultrasound lesion tracking accuracy and performed clinical trials. The experiments confirmed that the entire pipeline could address complex data from clinical HIFU environments. In the evaluation of US lesion tracking methods, we compared the automatically obtained lesion contour with the manually labeled contour and showed that the proposed method obtained an accuracy of 90.67%. The evaluation experiments showed that the proposed framework not only had better robustness for HIFU ultrasound images but also had a faster calculation speed than the conventional method. The entire image calculating speed could reach 30 FPS. In clinical experiments, we integrated the image guidance system with the commercial HIFU system and performed 8 procedures. Clinical experiments on volunteers verified that the intelligent treatment strategy could achieve the basic distribution of HIFU foci. Moreover, the image guidance system effectively reduced the difficulty of HIFU operation based on evaluations of the learning curve.The proposed intelligent HIFU theranostics strategy proved feasible in quantitative and clinical trials. However, at the current stage, the doctor still needs to confirm and monitor the operation and communicate with patients to ensure efficacy and safety during intelligent HIFU operation. In future work, more physiological signals will be integrated into the system for more comprehensive information acquisition. Moreover, the dosage of high-intensity ultrasound will be considered in the intelligent treatment strategy. To develop future intelligent HIFU theranostics, we will further focus on automatic surgical planning and the combination of decision- making and information. More intraoperative information will be included in subsequent studies, especially intraoperative feedback.In this work, we introduced an intelligent HIFU theranostics system. The system includes auto lesion detection in preoperative MRI and intraoperative real-time US images, an intelligent HIFU focus distribution strategy image-based unexpected movement monitoring. Several new proposed methods are used to ensure the accuracy and efficiency of the automatic theranostics procedure. Quantitative and qualitative experiments show that our system can accurately obtain lesion information and provide intelligent therapy in clinical cases.Supplementary movie 1.Click here for additional data file.Supplementary movie 2.Click here for additional data file.


Bladder cancer (BC) is the most common cancer of the urinary tract in the United States. Imaging plays a significant role in the management of patients with BC, including the locoregional staging and evaluation for distant metastatic disease, which cannot be assessed at the time of cystoscopy and biopsy/resection. We aim to review the current role of cross-sectional and molecular imaging modalities for the staging and restaging of BC and the potential advantages and limitations of each imaging modality. CT is the most widely available and frequently utilized imaging modality for BC and demonstrates good performance for the detection of nodal and visceral metastatic disease. MRI offers potential value for the locoregional staging and evaluation of muscular invasion of BC, which is critically important for prognostication and treatment decision-making. FDG-PET/MRI is a novel hybrid imaging modality combining the advantages of both MRI and FDG-PET/CT in a single-setting comprehensive staging examination and may represent the future of BC imaging evaluation. Other PET tracers have also been investigated for potentially improved diagnostic capability due to altered urinary excretion profiles. We aim to review these imaging modalities used in the staging and restaging of BC and the potential advantages and limitations of each imaging modality.Bladder cancer (BC) is the most common cancer of the urinary tract in the United States, accounting for 80,470 new cases and 17,670 deaths in 2019 fluorodeoxyglucose (FDG), which is a radiolabeled glucose analog that is taken up by the GLUT2 transporter that is overexpressed in many cancer cells. Typically, FDG-PET/CT is performed for the whole-body staging of cancers. The radiotracer is injected approximately 60 min prior to the scan, and the patients are instructed to follow a specific diet and activity regimen prior to the exam to avoid muscle activity and alterations in radiotracer biodistribution due to endogenous insulin release. A review of the Surveillance, Epidemiology, and End Results (SEER) database from 2004 to 2011 demonstrated that a total of 36,855 patients underwent a PET/CT scan for bladder cancer of any stage and that the number of scans markedly increased throughout the time period, resulting in excess national spending of $11.6 million for this imaging modality choline and [11C]acetate. Choline is a naturally occurring small molecule incorporated into tumor cells via phosphorylation by choline kinase. Acetate is a naturally occurring metabolic substrate that enters the fatty acid metabolic pathway. Both of these pathways/processes are upregulated in a number of cancers. However, a limitation of these radiotracers is the short half-life of 11C (20 min), which requires an on-site cyclotron for production and significantly limits geographic distribution. A systematic review and meta-analysis of the diagnostic accuracy of [11C]choline and [11C]acetate for lymph node staging in patients with bladder cancer found a pooled sensitivity of 66% and a pooled specificity of 89% [While FDG remains the most commonly utilized radiotracer in oncologic imaging, its pitfalls have led to research on other radiotracers. The two most studied alterative radiotracers used in the imaging of bladder cancer are [y of 89% .PET/MRI scanners are being increasingly installed and utilized throughout the world, leading to increased interest in appropriate applications of these scanners in clinical use. In particular, pelvic imaging is well suited for PET/MRI owing to the superior soft-tissue characterization gained by PET/MRI compared with PET/CT. For bladder cancer, the information gained from both PET and MRI is complementary and can lead to increased confidence in reporting findings, increased sensitivity of detection, and overall improvement in comprehensive staging in a single imaging examination. Given the novelty of this new hybrid imaging modality, there is limited research in the field of bladder cancer with PET/MRI. A pilot study of 21 patients with MIBC who underwent preoperative FDG-PET/MRI found that FDG-PET/MRI had a performance similar to that of CT for the detection of the primary tumor, but the detection of lymph node metastases were limited by the lower number of patients with true pathological lymph node involvement . A separBladder cancer imaging through multiple modalities provides important information in the staging and restaging of patients with BC. CT is the most widely available and frequently utilized imaging modality for BC and demonstrates good performance for the detection of nodal and visceral metastatic disease. MRI offers potential value for the locoregional staging and evaluation of muscular invasion of BC, which is critically important for prognostication and treatment decision-making. FDG-PET/CT is somewhat limited in the evaluation of the primary BC due to urinary excretion of the radiotracer, but shows good performance for the detection of nodal and visceral metastatic disease and offers the potential to detect metastatic disease in lymph nodes that are not anatomically enlarged. FDG-PET/MRI is a novel hybrid imaging modality with potential advantages of both MRI and FDG-PET/CT in a single-setting comprehensive staging examination, but scanner availability and limited research on the topic limit its widespread use.


The role of small game in prehistoric hunter-gatherer economy is a highly debated topic. Despite the general assumption that this practice was uneconomic, several studies have underlined the relevance of the circumstance of capture – in terms of hunting strategies and technology – in the evaluation of the actual role of small mammals in human foraging efficiency. Since very few studies have focused on the recognition of bone hunting lesions, in a previous work we explored the potential of 3D microscopy in distinguishing projectile impact marks from other taphonomic marks, developing a widely-applicable diagnostic framework based on experimental data and focused on Late Epigravettian projectiles. Even though we confirmed the validity of the method on zooarchaeological remains of large-sized mammals, the reliability of the experimental record in relation to smaller animals needed more testing and verification. In this report we thus present the data acquired through a new ballistic experiment on small mammals and compare the results to those previously obtained on medium-sized animals, in order to bolster the diagnostic criteria useful in bone lesion identification with specific reference to small game. We also present the application of this renewed methodology to an archaeological context dated to the Late Glacial and located in the eastern Italian Alps. A more general trend towards a broad-based economy, consisting of increased diversification of animal resources and greater inclusion of small game in the diet , is attested however only at the end of the Pleistocene during the Late Glacial. This process, documented in most of Mediterranean Europe and southern Levant, has been extensively debated and studied at different scales, searching for consistent chronology and causes that could clarify such a synchronous and widespread change in human behaviour13. Contrary to expectations, as studies in the fields increased, an underestimated complexity of this phenomenon has been revealed. Several factors have been called into question - including demographic pressure, social stress, climatic shifts, changes in subsistence patterns, technological innovations and resource depression - and their relevance has been tested case by case29.The role of small game in prehistoric hunter-gatherer economy is a highly debated topic. Although the exploitation of small prey was not uniform across time and space, their inclusion in the hominin diet is well documented in the Mediterranean area since the Middle Pleistocene31, the practice of small mammal hunting appears uneconomic regardless of taxa differentiation. Since prey items are ranked according to their energy return balance , small-bodied and difficult-to-catch mammals are usually categorised as low-ranking resources32. Going beyond this generalisation, several studies underlined other variables, such as circumstance of capture, hunting technology and game catchability, could be relevant in the evaluation of the actual role of small mammals in human foraging efficiency36. Moreover, mass collecting appears to be the most economically profitable method of capture for this kind of resource40. Determining the modality of capture, in terms of hunting strategies and technology, is thus of critical importance for understanding changes in human diets and shifts in game exploitation during Prehistory.Investigating shifts in game exploitation from the perspective of evolutionary ecology and, specifically, through the application of the prey choice model57. Conversely, very few studies have focused on the recognition of projectile impact marks , although they represent the only taphonomic evidence directly connected to hunting technologies employed by humans. Works documenting a few or single archaeological evidence of impact, such as stone fragments embedded in bone or healed wounds, are more numerous79. Despite some experimental works gave new insights towards the development of a methodology in PIMs recognition88, the recent application of high-resolution quantitative methods in bone taphonomy have mostly addressed cut mark, carnivore tooth marks and trampling marks characterisation106.Zooarchaeological and taphonomic studies have made important contributions toward the reconstruction of the dynamic interactions between humans and their prey, primarily defining the criteria useful in the identification of the accumulation agents of a faunal assemblage 107. Even though we confirmed the validity of the method on zooarchaeological remains of large-sized mammals108 the reliability of the experimental record in relation to smaller animals needed more testing and verification. Bone dimension and thickness could indeed affect their resistance to projectile impacts, influencing the morphometry of hunting injuries and the representativeness of PIM classes.As such, in previous work we used 3D microscopy for the development of a diagnostic method aimed at the distinction of bone hunting injuries from other taphonomic marks: the experimental PIMs taken as reference were produced exclusively by Late Epigravettian projectiles, shaping a widely-applicable framework useful for the identification of hunting bone lesions in other Late Glacial cultural complexesMyocastor coypus) and their comparison to results previously obtained on medium-sized animals, in order to adjust the diagnostic criteria useful in PIMs identification with specific regard to small game. We also present the application of this renewed methodology to an archaeological context dated to the Late Glacial and located in the eastern Italian Alps. This site, named Riparo I of Grotte Verdi di Pradis (hereafter Pradis Cave), represents an optimal case-study, being interpreted as a marmot specialised hunting camp, occupied seasonally by Late Epigravettian hunter-gatherers for the exploitation of this animal resource.In this report we thus present the data acquired through a new ballistic experimentation on small mammals truncated bladelets. Each lesion was classified by following the terminology proposed by O’Driscoll and Thompson58 and analysed, when possible, through 3D digital microscopy, acquiring multiple morphometrical parameters useful for statistical processing and comparison107.The previous ballistic experimentation conducted by the authors was aimed at the distinction of hunting lesions caused by Late Epigravettian projectiles on medium-sized ungulates117.The main methodological result consists of the recognition of drag and puncture marks as the only diagnostic categories in PIMs identification. Critical for their interpretation are the following elements: a) the location of the mark; b) the presence of flaking and/or cracking as secondary feature of both categories; c) the occurrence of specific morphometrical values, different for drags and punctures (detailed below); d) the presence of embedded stone characterised by diagnostic impact fractures107. The distinction of drags from carnivore tooth marks and cut marks (produced both by unmodified flakes and retouched implements) was tested through a 3D microanalysis, considering the parameters listed in the analytic method section .Since drag marks were the most frequent experimental bone injury (50%), they are assumed to be the most likely preserved on zooarchaeological material. This category is mainly located on ribs (40%), vertebrae (34.3%) and long bones, such as the radius and ulna (14.3%). The drag profile is extremely sharp and clean, especially on the most resistant bones, with frequent unilateral flaking (48.6%) and cracking (34.3%)107. Puncture marks appeared significantly deeper than carnivore tooth pits and were defined by a statistically lower RAV107. The distinction of impact punctures from corrosion cavities was easily conducted considering some morphological features such as the contour of the opening area and the shape of the slopes and floor. This experiment showed that some peculiar features of both drags  and punctures  were strictly connected to Late Epigravettian projectiles, confirming the hypothesis of a significant relationship between tip design and PIMs morphometrical features107.Puncture marks, recorded on the entire skeleton, were the second most frequent experimental trauma (24.3%). When the projectile caused the complete perforation of the bone, an imprint characterised by a more or less regular polygonal outline was generated. Cracking occurred in 48.1% of the punctures. Punctures were compared to carnivore pits and corrosion cavities considering the following parameters acquired through the 3D microanalysis: opening area (A), volume (V), depth of cavity (D) and the ratio between the opening area and volume of pits (RAV)To sum up, new clues in PIMs identification and distinction from other taphonomic marks were inferred through the application of 3D digital analysis, qualifying drags and punctures as reliable archaeological indicators of prey capture modalities.A total of 90 arrows armed with Late Epigravettian lithic backed points and bladelets were shot against 8 coypu fresh carcasses . It is also noteworthy that in 19.1% of cases (n = 4) small lithic fragments remained embedded in the groove. Lastly, drags documented on coypu incisors , the morphometric analysis was focused only on drags. The profiles and morphometric parameters of experimental drags were significantly different from the features of experimental cut marks produced on coypus as a reference , representing about 99% of the whole faunistic assemblage. The number of identified marmot specimens amounts to 11,285 fragments, with the minimum number of individuals being 637122 , located on all skeletal areas . Flaking is, however, visible on 24% of the cases (n = 6), whereas cracking and feathering are absent. As already suggested in a previous work108, this evidence confirms that a moderate post-depositional alteration of bone surfaces - in the form of roots etching, corrosion, manganese staining, trampling marks, corrosion and weathering - does not compromise the feasibility of the diagnostic method.The analysis of bone surface modifications led us to the identification of several possible PIMs, morphologically comparable to drag and puncture marks obtained through the ballistic experimentation on coypu carcasses Fig. . Drags w3D measurements, acquired on 21 of 25 specimens because of integrity issues, state that several drag morphometric parameters - such as depth of cut, breadth at the top, breadth at the floor of the cut and RTF index - are consistent with the features of experimental drags and significantly different from that of cut marks, both archaeological and experimental Fig. .Figure 4Lastly, a Principal Component Analysis carried out on experimental and archaeological items considering all the metric parameters, confirmed the interpretation of 21 marks as impact drags with only two uncertain specimens Fig. . PC1, wh107.Three archaeological punctures have been morphologically recognised on the base of the polygonal outline, even if two of them are associated with a fracture. They are located on a coxal and two femurs Fig. . Because124. Lastly, archaeological drags on ribs were proven as unreliable because of the possible overlapping of their morphometric features with cut marks, making them hard to distinguish from one another.This study yielded three main methodological outcomes. Firstly, relatively few puncture marks were produced compared to a very high number of impact fractures and drags. This confirms that a bone dimension and thickness affects its resistance to projectile impacts, which in turn influences the representativeness of PIM classes. Impact fractures were indistinguishable from other kinds of fractures, making drag marks the most relevant category for the identification of PIMs on small game zooarchaeological remains. Secondly, the consistency of drags morphometric data coming from the two experiments revealed that drag features are not influenced by bone size and thickness, confirming the validity and feasibility of the method. A similar result has been achieved by other studies which tested the influence exerted by prey size and bone dimensions in the morphology of cut marks108 where we supported O’Driscoll and Thompson’s hypothesis58 that only a considerable amount of force could plunge lithic fragments into bones. Although we did not find any small lithic fragments embedded in Pradis archaeological drags, supporting evidence comes from a similar context, the Madgalenian site of La Colomb . This cave, interpreted as a camp specialised in marmot hunting, yielded a marmot shoulder blade with lithic fragments fixed in two possible hunting lesions121.Discussing elements useful in drag recognition, several features were confirmed as relevant for a first qualitative assessment: the location of the mark; the presence of flaking and/or cracking as secondary features; and the occurrence of lithic fragments embedded into the groove. The new experimental data confirmed the critical importance of the occurrence of lithic fragments embedded into the groove, as already assumed in a previous publicationOur study also confirms the effectiveness of 3D digital microanalysis in PIMs recognition. The collection of reproducible morphometrical parameters that can be processed through statistics, is indeed critical for the validation of the above qualitative assumptions. Even if some features - such as the depth of cut, breadth at the top, breadth at the floor and RTF index - emerged as more relevant than others in the application of this method on small mammal faunistic assemblages, the overall reliability of the diagnostic framework has been successfully tested. Moreover, we verified that a moderate post-depositional alteration of bone surfaces does not compromise the feasibility of the method on archaeological specimens.128. Stone-tipped projectiles seem to have been used almost exclusively on “large” game, with a few exceptions in North America and Alaska130. In light of this varied scenario, different authors questioned which capture technology could have been employed in order to make small game’s exploitation economically profitable, especially if compared to ungulates hunting39. As for Pradis Cave, our analysis connected marmot hunting to the use of the bow-arrow delivery system and lithic tipped projectiles, excluding the development of sophisticated acquisition techniques formerly unknown. The hypothesis that the bow and arrow was widespread in Europe during the Upper Palaeolithic has been proposed by many scholars for different chrono-cultural contexts137. Moreover, the existence of Late Epigravettian composite lithic projectiles - with a backed point hafted as piercing element and backed (bi)truncated bladelets functioning as lateral cutting elements - has been frequently proposed on the basis of backed tool’s design, use-wear analysis and the characteristics of lithic fragments found embedded in archaeological PIMs139. The Pradis data is further supported by a marmot predation using lithic tipped arrows uncovered at the La Colomb Cave site. The hunting scenario reconstructed by Tomé and Chaix121, wherein the animal was hit while sitting on two legs, strongly resembles the Pradis data which suggest a prevalence in predation of marmots in alert position, on the basis of experimental and archaeological PIMs location  have been classified as younger than 4 months, we could infer a predation between August and October, just before marmot winter hibernation . Although this occupational pattern needs more verification , it suggests a predation during the most economically profitable period. A similar assumption has been proposed for Clusantin Cave nearby Pradis Cave120 and other marmot hunting sites dated to Late Glacial and located in the western Alpine region121. Several sites between Vercors mountains and southern Jura showed a strong connection between their altitudinal location and paleoenvironmental data, suggesting the existence of a mobility strategy specifically linked to the exploitation of high altitude resources during the season which provides the maximum energetic return121.Zooarchaeological and taphonomical analyses demonstrated that the marmot assemblage from Pradis Cave was primarily accumulated by Late Glacial hunter-gatherers. The dominance of such small prey is uncommon for Late Glacial sites of north-eastern Italy and is probably connected to the location and function of the rock shelter. The presence of at least 637 marmot individuals suggests a well-developed capture strategy and efficient hunting gear. The use of several techniques to hunt large numbers of small prey - including bows and arrows, throwing weapons, nets, snares or traps - has been reported by ethnographic and historical sources141 in an open forest environment characterised by an ecotonal zone between the woods and the alpine prairie141. We can thus suppose that such ecological conditions sustained large marmot colonies, well adapted to the rocky habitat along the abrupt slopes surrounding the Pradis plateau120. Therefore, the recurrent human visits in that area focusing on marmot mass exploitation could be the result of multiple factors integrating both prey availability and human choice, in the framework of the progressive human colonisation of mid-altitude territories that had been previously abandoned during the Late Glacial Maximum143. The specialisation of these sites in marmot hunting fits well in the logistical mobility system assumed for Late Epigravettian hunter-gatherers of north-eastern Italy, structured on sites that were functionally complementary to each other and located at different altitudes144. The continual return of humans to that area was certainly based on the local availability of this specific resource and we could not exclude that hunting with bow and arrow led humans to carry out a sort of resource management, selecting mostly adult individuals and avoiding juvenile preys, in order to allow marmot colonies to reproduce and thrive over time.During the first part of the Late Glacial interstadial when Late Epigravettian human visits occurred, Pradis Cave and the close Clusantin Cave were both located below the upper limit of the tree lineThis work presents new data obtained through a ballistic experiment on small-sized mammals. It also compares the results to those previously obtained on medium-sized animals, in order to bolster the diagnostic criteria useful in PIMs identification with specific reference to small game. As main methodological outcome, drag marks were recognised as the most relevant category for the identification of bone hunting lesions on small game zooarchaeological remains. Moreover, the consistency of drags morphometric data coming from the two experiments revealed that drag features are not influenced by bone size and thickness, confirming the validity and feasibility of this diagnostic method on small-sized animals.The results presented in this work provide not only specific evidence for the identification of hunting traumas caused by Late Epigravettian projectiles, but also comparative data useful for taphonomic analyses of bones from other Late Glacial cultural contexts, especially those characterised by backed tools as projectile implements.As already discussed, the reconstruction of hunting strategies and technology is critical to evaluate the contribution of different animal resources in human foraging efficiency. This is particularly significant in reference to small-sized mammals, whose economic role in human foraging strategies is generally underestimated. Being hunting injuries reliable indicators of the agents of bone accumulation and prey capture modalities, we thus propose to make the identification of PIMs a standard practice in zooarchaeological studies.Myocastor coypus) is a small-sized mammal very similar to alpine marmot (Marmota marmota) in weight and size (about 3–10 kg and 40–70 cm in head-and-body length). As such, the experiment involved 8 fresh coypu carcasses shot by 90 Late Epigravettian arrows equipped with backed points and bladelets made of flint  in order to collect experimental cut marks on small mammal bones.Coypu . The identification of the surface alterations and the distinction of different taphonomic agents were carried out with reference to well-established literature57. The descriptive criteria and the terminology used for PIMs, follows O’Driscoll and Thompson58 as already discussed in Duches et al.107.Taxonomic and skeletal identifications were carried out on the basis of the complete alpine fauna reference collection of MUSE-Science Museum of Trento. Marmot NMI was calculated on the number of left emimandible while the evaluation of age classes was based on dental use-wear and eruption stages from Couturier collection of the Natural History Museum of Grenoble, already analysed by Fournier149. Each drag was analysed through the acquisition of different metrical parameters, according to criteria already published  and using one cross-section per mark in its median part . Regarding the tooth marks compared to experimental drags, they consisted of 58 present-day scores found on ungulate carcasses collected in the field and were attributable to small wild carnivores and dogs107. All the measurements considered in this paper have been directly acquired by two of the authors (FB and JC), using the software directly installed in the digital microscope. They also processed the data in the various statistical tests using the PAST software150.Moreover, the values were compared to 123 experimental cut marks produced with unretouched flakes: 22 were inflicted on cattle autopodialsSupplementary Information.


Non-missile transorbital penetrating head injuries are relatively rare, though potentially fatal injuries. Trajectory for intracranial entrance is typically via the orbital roof, the superior orbital fissure (SOF), or the optic canal. Non-metallic intracranial penetrating injuries are even scarcer and may pose unusual diagnostic and surgical challenges. Here we present and discuss a unique case of a penetrating injury by a wooden foreign body (FB) which entered and expanded the inter-dural space of the lateral cavernous sinus (CS) sinus wall without intracavernous or intradural involvement. The patient was a 71 year-old male who fell face-down and sustained a penetrating transorbital injury by a dry twig fragment, which passed through the SOF and into the interdural space of lateral wall of the ipsilateral CS. The patient was fully conscious (GCS15) at presentation but had severe ocular injury . The wooden FB was successfully removed via a minimally invasive subtemporal intradural approach with no apparent immediate or long-term complications. We emphasize the unusual diagnostic and surgical challenges related to this kind of rare injuries as reflected by the decision-making considerations taken in the presented case. Transorbital intracranial injuries are rare and potentially fatal , 2. ThisHere we present and discuss a unique case of a penetrating wooden FB injury located within the interdural space of the lateral CS wall without intracavernous or intradural involvement.Under the influence of alcohol intoxication, a 71 year-old male fell face-down at the edge of an agricultural field. After returning home, his children have noticed a prominent swelling around his left eye. He refused any medical attention for 2 days. finally, 3 days post-injury, he presented to the emergency department with fever and severe left periorbital edema accompanied with a purulent discharge. The patient was fully conscious, though uncooperative. A small superior eyelid puncture wound around the left medial cantus was observed. Neurological examination revealed complete left ophthalmoplegia and blindness (no light perception) of the left eye. Fundoscopy revealed a pale left optic nerve. No signs of meningeal irritation were observed. Sensory functions of the left trigeminal nerve were hard to evaluate (due to poor cooperation by the patient). Muscles of mastication functions were preserved.Non-contrast CT of the head was performed and revealed a linear shaped, segmented into 2 pieces  hypodense, air-mimicking FB, extending from the upper medial part of the left orbit , through the SOF (sparing the globe) and into the cranial space . Prompt No evidence of facial or cranial bones fractures.The FB is located lateral to the left carvernous sinus with no evidence of intra cavernous involvement. Extra-cavernous-Intra-dural location of the FB was assumed.No evidence of active bleeding, vascular injury, or CS thrombosis in the CTA and CTV studies .The patient was planned for a 2-staged surgery—first, maximal safe removal of the orbital part of the FB (by the ENT team), followed by a second intra-cranial procedure.The ENT surgeon opened and expanded the small palpebral entry wound and performed intra-orbital exploration. The superficial part and only the proximal end of the deep part of the FB (which was found out to be a dry twig) could be resected via this approach. Pus evacuation and meticulous intraorbital irrigation were performed. The impression of the ENT surgeon at that stage was that the residual intraorbital FB is stuck and further removal of it is highly dangerous. Therefore, head CT scan was performed  the patiThe patient was placed in the supine position with his head fixated by a Mayfield head clamp and rotated to the right. Small subtemporal craniotomy (2.5 × 2.5 cm) through a small linear scalp incision was performed. To minimize subsequent temporal lobe retraction, CSF drainage trough a small dural incision was performed at this early stage, followed by drilling of the temporobasal edge of the craniotomy window with high speed drill. The dura was incised in a “C-shaped” fashion. Gentle retraction of the temporal lobe and additional CSF drainage allowed for the exposure of the thickened hyperemic superficial layer of the lateral wall of the CS. In contrast to our expectations the FB was not visualized at this stage. Adjacent non-injured anatomical structures were recognized, namely, the mesial part of the temporal lobe, the tentorial edge, the left optic nerve, the left ICA and PCom, and the left oculomotor nerve . CarefulEnterobacter species growth in the microbiological assay of the pus sampled during intraorbital surgery). The patient was discharged home about 2 weeks after presentation. During the follow-up period no infectious-related complications (ocular or CNS) were observed. The patient did not have any suspected seizure events nor any apparent cognitive deterioration. Follow-up MRI scan a year after the injury did not demonstrate any pathological findings other than mild gliosis detected at the basal temporal lobe , therefore raising a radiological diagnostic challenge for intracranial wood injuries , 21. HelWood, with its porous consistency and organic nature, serves as a medium for microbial proliferation, and thus may increase the risks to complications such as panophthalmitis, abscess, and Fistula  . Careful inspection of the pre-operative CTA can yield important insights regarding the spatial relationship between the FB and the nearby arterial structures. For marginal cases, in which higher resolution demonstration of the vessels is required, the utilization of formal angiography and\or MRA\MRV is warranted. If vascular damage is suspected, the surgical strategy should consider early proximal and distal control of the main vessels, or alternatively, pre-surgical endovascular intervention with conventional angiogram for intravascular inflatable balloon insertion . In the The medial aspect of the temporal lobe and the lateral CS can be approached from different directions\craniotomies  , 25, 30.As mentioned above, if we were to know the correct location of the FB in the inter-dural space of the lateral wall of the CS pre-operatively, we would have consider utilizing an extradural approach as a reasonable alternative to the intradural approach that we chose. We consider the lack of direct intradural basal temporal lobe manipulation, which may result in the development of undesirable neurological sequalae  as It provides the mandatory spacious exposure required for the safe visualization and removal of the FB.It allows for a more perpendicular working angle with regard to the axis of the FB which in turn allows for pulling movements along this axis.It is less complex and less destructive The main pros and cons considerations for the utilization of each approach for the current case are summarized in the We believe that complete resolution of the infection by a relatively short course of targeted IV antibiotic treatment can be attributed mainly to the following factors:The infection seemed to be restricted to the extracranial\orbital space.The unusual location of the FB between the two layers sparing both the CS and intradural space.Specifically, it is likely to assume that the preserved superficial layer lowered dramatically the risk of local spreading of the infection into the adjacent CS sinus septic thrombophlebitisMeticulous orbital and intracranial irrigation.Watertight suturing of the dura propria at the end of the surgery.Targeted antibiotic treatment post-operatively.In conclusion, transorbital penetrating cranial injury by a wooden FB is rare and poses the risk for potentially life-threatening complications. Careful anamnesis, physical examination and inspection of pre-operative imaging accompanied by multidisciplinary approach  are of highly importance for optimal emergent surgical planning and execution leading to good outcome for the patients.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The Institutional Review Board of Soroka Medical Center waived the requirement for informed consent for the publication of this case report.EA, AS, and IM: study conception, data collection, data analysis, manuscript writing/editing, and final approval of manuscript. RS, AA, YS, TK-E, and GZ: data collection, data analysis, manuscript editing, and final approval of manuscript. All authors contributed to the article and approved the submitted version.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.