People living with dementia often exhibit behavioural and psychological symptoms of dementia that can put their and others’ safety at risk. Existing video surveillance systems in long-term care facilities can be used to monitor such behaviours of risk to alert the staff to prevent potential injuries or death in some cases. However, these behaviours of risk events are heterogeneous and infrequent in comparison to normal events. Moreover, analysing raw videos can also raise privacy concerns.  In this paper, we present two novel privacy-protecting video-based anomaly detection approaches to detect behaviours of risks in people with dementia.  We either extracted body pose information as skeletons or used semantic segmentation masks to replace multiple humans in the scene with their semantic boundaries. Our work differs from most existing approaches for video anomaly detection that focus on appearance-based features, which can put the privacy of a person at risk and is also susceptible to pixel-based noise, including illumination and viewing direction. We used anonymized videos of normal activities to train customized spatio-temporal convolutional autoencoders and identify behaviours of risk as anomalies.  We showed our results on a real-world study conducted in a dementia care unit with patients with dementia, containing approximately 21 h of normal activities data for training and 9 h of data containing normal and behaviours of risk events for testing. We compared our approaches with the original RGB videos and obtained a similar area under the receiver operating characteristic curve performance of 0.807 for the skeleton-based approach and 0.823 for the segmentation mask-based approach.  This is one of the first studies to incorporate privacy for the detection of behaviours of risks in people with dementia. Our research opens up new avenues to reduce injuries in long-term care homes, improve the quality of life of residents, and design privacy-aware approaches for people living in the community. Dementia is a syndrome that involves progressive impairment of cognitive functions such as memory and thinking and can impact the insight, impulse control and judgement of a person . It can The behaviours of risk exhibited by PwD are episodic and infrequently occur in comparison to normal activities . TherefoConsidering the privacy aspect of PwD and staff and the infrequent nature of behaviours of risk events, we present novel privacy-protecting anomaly detection approaches to detect these behaviours. This paper proposes two privacy-protecting approaches for detecting behaviours of risk events in PwD as anomalies using unsupervised convolutional autoencoders using real-world video surveillance data collected from a dementia care unit. The proposed privacy-protecting approaches are based on data preprocessing steps that either extract skeletons of the individuals using human pose estimation algorithms , 12 or uWe investigate the effectiveness of both the window and frame-level approaches corresponding to 3D and 2D convolution autoencoders, respectively, to detect the behaviour of risk events in PwD as anomalies.We propose two privacy-protecting approaches, namely, skeleton and semantic segmentation mask-based approaches, that enable the two types of convolutional autoencoders to model the behaviours of risk in PwD related to the posture and actions of the individuals and their interaction with each other, and the objects in the environment using video surveillance data collected from a dementia care unit.We show that the proposed approaches perform equivalent to the unsupervised deep models trained on RGB videos, while protecting the appearance-based information of the people.The focus of this paper is to demonstrate the effectiveness of the proposed privacy-protecting approaches as an alternative and replacement to traditional RGB videos for the detection of behaviours of risk in PwD.To the best of our knowledge, this is the first work that utilizes human skeletons to model human posture, human–human interaction and human–object interaction-based behaviours of risk in PwD in a privacy-protecting setting. The contributions of this paper are threefold: We now present a brief overview of the existing work in the field of automatic detection of behaviours of risk in PwD using data modalities that include video. This is followed by a brief overview of the video-based anomaly detection methods that use skeletons or semantic segmentation masks to incorporate privacy in their design.The existing work in the automatic detection of behaviours of risk, such as agitation and aggression, in PwD focuses on the use of three different sensing modalities: wearable, computer vision, and multimodal sensing. Multimodal sensing refers to a combination of wearable, and/or computer vision, and/or other ambient sensors to detect behaviours of risk in PwD. Actigraphy/accelerometer has been used previously to detect agitation and has shown correlation . Since tThe video-based methods operate on pixel-based appearance and motion features in videos and hence can be sensitive to noise resulting from the appearance of the individuals. Extracting information specific to the body pose of the people in the form of skeletons can help filter out the appearance-related noise for detecting abnormal events related to the posture and actions of the individuals. Human pose estimation algorithms can be used to extract body joints in the form of skeletons of the individuals in the scene , 20. ComThe skeletons are a good privacy-protecting source of information about human posture. However, the quality of skeleton approximation depends upon the resolution of video frames and the degree of occlusion due to objects or people in the scene . OccludiIn this section, we describe the dataset used in this paper, the data preprocessing steps involved and the details of the convolutional autoencoders used to detect behaviours of risk in PwD.There is a scarcity of video data to study behaviours of risk in PwD in a residential care setting. The few existing approaches either use simulated environment or feasibility studies , 29. In mentclass2pt{minimRGB frames: These were the RGB video frames extracted from the sampled videos, without further processing.Openpose skeleton frames without background: Openpose  was usedOpenpose skeleton frames with background: Openpose  was usedDetectron skeleton frames without background: Detectron2  was usedDetectron skeleton frames with background: Detectron2  was usedSegmentation mask frames without background: Semantic segmentation masks  depictinSegmentation mask frames with background: Semantic segmentation masks  was apprThe frames were converted to grayscale, normalized to the range   and resized to The original videos had a frame rate of 30 frames per second. However, to ensure efficient use of computational resources, the frames were sampled at 15 frames per second for analysis, retaining only half the frames. Oftentimes, there were presence of multiple individuals and occluding objects  in the common areas of the unit. This made it difficult for the pose estimation algorithms to approximate the skeletons. Hence, we used two different pose estimation algorithms, namely, Openpose  and Deteus paper .Convolutional autoencoders (CAEs) learn to reconstruct the input image(s) at output by minimizing the reconstruction error during training. In general, CAEs follow an unsupervised learning approach and are trained using only normal behaviour samples. The intuition behind use of CAEs is that as they learn to reconstruct only samples representing normal behaviour during training, they are expected to give high reconstruction error for anomalous samples at test time. In existing literature, CAEs have been observed to perform well for single-scene video anomaly detection  and exteThe CAE-3DConv model was adapted from the previous work by Khan et al. , and conThe CAE-2DConv model consisted of an encoder–decoder architecture, which forced the model to learn only the key spatial features in the input window. Compared to CAE-3DConv. here the encoder consisted of 2D convolution and max-pooling blocks to encode the input. The 2D convolution blocks were responsible for 2D convolution operation, followed by batch normalization and ReLU operation. A convolution kernel of size , and Both CAE-3DConv and CAE-2DConv models were trained using input windows containing only the normal activities to minimize the following reconstruction error:We performed experiments to investigate the effectiveness of the proposed privacy-protecting approaches in detecting behaviours of risk in PwD in comparison to RGB video inputs. We trained the CAE-3DConv and CAE-2DConv models on RGB video and different privacy-protecting inputs using the same experimental setup. Both the CAE-3DConv and CAE-2DConv models were trained for 70 epochs and used Adam optimizer with a learning rate of 0.001. The models were implemented in pytorch v1.7.1 and pytorch lightning v1.5.2  and run Table The performance of the privacy-protecting without background approaches was lower in comparison to with background and the RGB video input. This can be attributed to the lack of information related to the objects in the environment. The behaviours of risk in PwD are a combination of different types of anomalous behaviours, including, human posture, human–human interaction and human–object interaction-based anomalies. The privacy-protecting approaches without background fail to model the human–object interaction-based anomalies, leading to poor performance. Particularly, the segmentation mask without background input contains only semantic boundaries of the individuals in the scene leading to the absence of sufficient information regarding the posture and interaction of the individuals with each other and the environment.The spatial information-based CAE-2DConv model performed slightly better than the spatio-temporal CAE-3DConv model, except for Openpose skeleton without background. The video surveillance data used in this research were taken from the common area of a dementia care unit. As such, there is frequent movement of a number of people within the video scene, leading to crowded scenes of people and objects moving at different paces. This makes it difficult for the methods to model the temporal information within the scenes, leading to lower performance when the temporal information within the window is leveraged.The baseline value for the PR curve, as can be seen in Figs. From the above observations, it can be concluded that the privacy-protecting with background approaches that involve extracting only the skeleton information or masking the body region of the individuals in the video frames can both protect sensitive information and achieve an equivalent performance in comparison to RGB input. These results pave the way for furthering biomedical research in care and community settings to utilize videos without breaching the privacy of individuals in the form of their identifiable information. Further, the analysts can still infer the activities in the scene from the segmentation masks/skeletons.Our approaches allow leveraging the important contextual information in the video frames while protecting the privacy of the individuals by not considering the identifiable appearance-based features. The contextual information refers to features related to the background and the interaction of the individuals with each other and the objects in the environment. The use of skeletons and segmentation masks can help to develop privacy-protecting solutions for private or community dwellings, crowded/public areas, medical settings, rehabilitation centres and long-term care homes to detect the behaviour of risk events in PwD. Cameras, such as ‘Sentinare 2’ from Altumview , can dirProviding care for PwD in care settings is challenging due to the increasing number of patients and understaffing issues. Untoward incidents may happen in these facilities that can put the health and safety of patients, staff, and caregivers at risk. Utilizing existing video infrastructure can lead to the development of novel deep learning approaches to detect these behaviours or risk events, prevent injuries and improve patient care. However, RGB videos contain identifiable information, and their use is not straightforward in a healthcare setting. In this work, we proposed two privacy-protecting approaches for detecting the behaviours of risks in PwD, an application, where safeguarding the privacy of the individuals is a major concern. The proposed approaches are based on either extracting body postures in the form of skeletons for the people or using semantic segmentation to mask the body areas of the people in the video scenes. The privacy-protecting inputs were passed as image input to two types of convolutional autoencoders that learned the characteristics of normal video scenes and identified behaviours of risk scenes as anomalies. We investigated both window and frame-level approaches for behaviours of risk detection as anomalies using convolutional autoencoders with 3D and 2D convolutions, respectively. We demonstrated that the privacy-protecting approaches based on skeletons (AUC(ROC) = 0.812) and semantic segmentation (AUC(ROC) = 0.823) with background information are able to detect behaviours of risk in PwD as anomalies with a similar performance in comparison to the RGB video input (AUC(ROC) = 0.822). Hence, the skeletons and semantic masks may be viable substitutes for the appearance-based information of the people in the scene and can help preserve their privacy.From a clinical perspective, this work is an important step towards developing video-based privacy-protecting behaviours of risk detection system in long-term care, residential care and mental health inpatient settings. An anomaly detection framework is helpful in this regard as the behaviours of risk encompass a wide range of actions, such as falls, hitting, banging on the door or throwing furniture. In addition, it does not need the appearance characteristics of the individuals. However, the challenges in this approach are that any unusual or infrequent event, such as large moving objects or crowded scenes, could be flagged as events of interest, leading to increased false positives. A clinical monitoring system based on this technology will need to have methods in place to avoid disruptions due to these false positive alarms. Our future work includes investigating active learning approaches to reduce false positives while training the autoencoders. Further, a multimodal approach will be investigated that uses privacy-protecting input modalities like skeletons, optical flow maps or semantic masks.


In this study we address the automatic segmentation of selected muscles of the thigh and leg through a supervised deep learning approach.The application of quantitative imaging in neuromuscular diseases requires the availability of regions of interest (ROI) drawn on muscles to extract quantitative parameters. Up to now, manual drawing of ROIs has been considered the gold standard in clinical studies, with no clear and universally accepted standardized procedure for segmentation. Several automatic methods, based mainly on machine learning and deep learning algorithms, have recently been proposed to discriminate between skeletal muscle, bone, subcutaneous and intermuscular adipose tissue. We develop a supervised deep learning approach based on a unified framework for ROI segmentation.The proposed network generates segmentation maps with high accuracy, consisting in Dice Scores ranging from 0.89 to 0.95, with respect to “ground truth” manually segmented labelled images, also showing high average performance in both mild and severe cases of disease involvement (i.e. entity of fatty replacement).The presented results are promising and potentially translatable to different skeletal muscle groups and other MRI sequences with different contrast and resolution. Recent technical advances of muscle MRI imaging have led to an evolution from traditional qualitative evaluation into what is currently known as quantitative imaging (qMRI), in which a large amount of diagnostically relevant information (such as fat substitution and edema) can be quantified and extracted from muscles of subjects affected by neuromuscular diseases , 23, 28.Up to now, manual drawing of ROIs has been considered the gold standard for the extraction of quantitative data from muscles in clinical studies , 24. It All these aforementioned methods provided a high level of accuracy of the generated segmentation maps with respect to ground truth labelled images, ranging from 0.8 to 0.97 values of Dice Similarity Coefficient  for the different tissues, with the deep learning-based methods performing better in the cases of severe fat substitution , 10. IndAs for the automatic segmentation of individual muscle regions, atlas-based approaches have been proposed in  for the Moreover, in  a cascadIn the present work, as a further step towards the automatization of muscle ROI drawing, we aimed to develop an automatic segmentation tool based on deep learning techniques to create single-muscle segmentation maps at thigh and leg level, starting from manually segmented multi-contrast quantitative muscle MRI scans of both healthy subjects and patients affected by two different neuromuscular diseases. In the interest of reproducibility and of benefiting the community, we are sharing the resulting automatic segmentation tool as an open-source repository, available at .For this project, we included 54 subjects   using integrated spine and body surface coils. The patient was lying supine in the scanner with 18–channel phased–array coils positioned either on the thighs and the legs during acquisition, with simultaneous acquisition of both sides . The MRI protocol included a 3D six-point multi-echo gradient echo (GRE) sequence with interleaved echo sampling  of the MESE acquisition, equidistant from the femur head and the tip of the patella, and were subsequently registered to the multi-echo GRE dataset with the creation of new corresponding ROIs, which were manually adjusted by the same operator. Two additional ROIs were drawn in the GRE space in the neighboring slices to the medial registered slice, ending in a final number of 7 slices per thigh segmented.For what concerns the leg, segmentation was performed in the third slice of the MESE acquisition and then subsequently registered to the multi-echo GRE dataset where it was manually adjusted. Two additional ROIs were drawn in the multi-echo GRE dataset on the neighboring slices, ending in a final number of 3 slices per leg segmented.The slices to be segmented were chosen as the most representative of the upper, middle and lower thigh, and, for simplicity, only for the middle portion of the lower leg .We separated the available dataset of scanned subjects into training and validation subsets, for the DNN learning process, and a test subset for its testing. 44 subjects  at the different time points, for a total number of 110 scans, were included in the training and validation subsets, whereas remaining 10 patients at their initial scan time-point were included in the test subset.A total number of 770 thigh and 330 leg slices with corresponding manually-drawn ground truth segmentations were thus available as a working dataset for the DNN learning process and cross-validation.We processed the input volume with a slice-wise approach. Figure Each manual segmentation map was preprocessed through the application of consecutive area opening and closing filters, with an area threshold of 4 pixels, to eliminate small structures which resulted from noise in the registration operation of the MESE onto the GRE dataset.Horizontal and vertical translations: separate independent translations of the left and right thighs (or legs) per image in the horizontal and vertical directions, with bi-cubic spline interpolation. The amounts of each translation were uniformly sampled in an interval of values computed per image with the maximum value given by the shortest distance of the thighs (or legs) to the image borders. These transformations enhanced invariance with respect to the relative position between the left and right thighs (or legs) in the training process;Rotations: independent rotations of the left and right thighs (or legs) by amounts uniformly sampled per image in the interval Piecewise affine transformations: separate affine transformations applied on each neighborhood of points on a Elastic transformations: local transformations obtained in terms of displacement fields with Gaussian kernel smoothing, with strength uniformly sampled per image from the interval  and standard deviation of the kernel uniformly sampled per image from the interval . Bi-cubic spline interpolation was chosen to determine new pixel values for the transformations. These transformations enhanced elastic distortions-invariance, representing realistic tissue variations, in the training process.The aforementioned data augmentation was applied to the available dataset of 770 thigh and 330 leg slices to obtain 5000 annotated images for thigh and leg respectively. We randomly separated this augmented dataset into a training dataset of 4500 elements and a validation dataset of 500 elements, to perform a cross-validation analysis on the network performance.We applied data augmentation to the available annotated slices in the training and validation datasets to gain robustness in the network predictions on unseen cases and to make the network learn realistic deformations without these being represented in the available training data. In particular, on each training and validation image and on each corresponding manual segmentation we randomly applied elements in the following sequence of transformations :VNet 26]. DiffThe current study had some limitations. First, since ground truth segmentations were available only on selected slices of the MRI volume stack (see Sect. In this study, we explored the applicability of deep neural networks in ROI drawing of muscles of the lower limbs, with promising results in terms of accuracy compared to the standard manual reference currently in use. The application of neural networks to substitute or at least greatly accelerate the work of human operators in ROI drawing can be extremely helpful in clinical studies, where a large amount of data have to be analyzed. Once reliable dedicated datasets of muscle ROIs are collected, deep neural networks can be promisingly applied for segmentation of other sequences with different contrast and image resolution and also to different anatomic districts.


Emerging zoonotic infections present a serious global health threat . It is eIt is reasonable to anticipate that the number of outbreaks and pandemics could increase in the near future. This assumption is based on multiple factors such as increased frequency of travel, particularly to remote areas, climate change, intensified global trade, susceptibility to pathogen exposure and several more . In addiThe SARS-CoV-2 pandemic has emphasized global weaknesses in the detection and control strategies aimed to minimize the human health and socioeconomic impact of exposure to zoonotic pathogens . It is cA major obstacle in our progress towards developing novel therapeutics and vaccines is the limited knowledge of the reservoir host disease dynamics, virus-recipient host interactions and the detailed processes that permit the establishment of infection . This isIn this Special Issue, we presented manuscripts  providing an insight on the distribution, spread, and protein functions of SARS-CoV-2, Puumala orthohantavirus and Ebola viruses. In the original research manuscript, Au and colleagues addressed the important issue of detecting and analyzing SARS-CoV-2 virus transcription in infected cells . AlthougIn another original study by Davidyuk et al., the genetic diversity of Puumala orthohantavirus (PUUV) in the Republic of Tatarstan (RT) was analyzed . PUUV prThis Special Issue also features review manuscripts on Ebola virus protein function, structure, and immune properties. The manuscript by Jain et al. presents a detailed analysis and discussion of the multi-functional features of each Ebola virus (EBOV) protein . EBOV beIn the final review, Jain et al. present a concise description of the cell types targeted by EBOV . In addiCollectively, this Special Issue highlights the proficiency with which viruses are able to cause severe disease and the complexities of virus–host interactions that impact both disease pathology and outcome. The knowledge attained from the articles covered here could aid the development of more specific peptide-based antivirals, monoclonal antibodies, and novel vaccines that protect against infections in the future. This area of research is absolutely essential and is urgently needed for better preparation for future pandemics.We wish to express our appreciation to all the authors who have participated in this Special Issue and the reviewers for their comments.


Background: The prevalence of early initiation of breastfeeding (EIBF) was found to be low among adolescent postnatal mothers. EIBF is associated with improved parenting skills and better neurodevelopment in babies. We aimed to improve the prevalence of EIBF among teenage mothers to at least 90% through a quality improvement (QI) initiative.Materials and methods: It was a QI intervention conducted over a period of 15 months involving mothers under the age of 20 years at the time of present delivery. Six Plan-Do-Study-Act (PDSA) cycles consisting of multiple interventions, each lasting for one month, were conducted to improve the prevalence of EIBF. Rate of improvement in EIBF was noted and plotted against time. Post-intervention follow-ups of observations were done for six months.Results: The prevalence of EIBF among adolescent mothers was 28.5% during the pre-intervention baseline phase. QI team meeting was held and the barriers to EIBF among the adolescent mothers were discussed and depicted in the form of a fish-bone analysis model. The prevalence of EIBF increased during each intervention cycle to 50%, 60%, 62.5%, 72.7%, 88%, and 100%. At the end of six months follow-up phase, the prevalence of EIBF sustained at around 100%.Conclusion: This QI initiative has proven to be effective in improving the prevalence of EIBF with simple but effective measures. Adolescent women comprise of a vulnerable sub-population of high-risk mothers. Proper counseling and respectful maternity care will help them face the challenges of motherhood boldly. Early initiation of breastfeeding (EIBF) is known to reduce neonatal mortality and morbidity ,2. ColosThis QI initiative was conducted at a tertiary care teaching hospital in North India over a period of 15 months. It aimed to improve EIBF among neonates born to adolescent mothers. A baseline cross-sectional assessment over a period of three months showed that the rate of EIBF (within one hour of delivery) was low among teenage mothers compared to the adult population. Institutional ethics committee approval was obtained from Employee's State Insurance Corporation (ESIC) Medical College and Hospital, Faridabad, prior to the commencement of the study . Written informed consent was obtained from all participants. The study was conducted in the following three phases: a pre-intervention baseline phase (three months), an intervention phase (six months), and a post-intervention follow-up phase (six months).All deliveries being conducted  during the study time frame, where the newborns were shifted to mothers' side after initial resuscitation by the concerned pediatrician, were potentially eligible for recruitment. Mothers under the age of 20 years at the time of present delivery were included in the study. For women below 18 years, written assent was obtained. The cases where newborns required intensive care unit admission in the first hour of life or referral to other higher centers and babies born to mothers of age 20 years or more at the time of delivery were excluded from the study. During the pre-intervention phase, a baseline audit was conducted over a period of three months to assess the prevalence of EIBF among adolescent mothers delivering at our hospital. A QI team was constituted of six members under the guidance of the team leader. Team members included faculty guides, resident doctors, and nursing staff in the labor and postnatal wards. In the first meeting, the dedicated QI team formulated the problem statement, smart aim, and strategies to implement interventions. The problem statement tried to address the low rate of EIBF among teenage mothers in our hospital setup. The issues were poor knowledge, hesitation to initiate breastfeeding, lack of skill, and shyness at the level of adolescent mothers, and reluctance to initiate and guide the adolescent mothers at the level of nursing staff. No proper documentation of timing of breastfeeding and no designated staff to ensure EIBF were hospital-level policy issues. Presence of male relatives in the ward and gender discrimination were also noted. The smart aim was to increase the prevalence of EIBF to at least 90% through dedicated PDSA cycle. Each PDSA cycle included these various methods - (a) monthly training of the doctors and nursing staff was conducted through dedicated lectures and interactive question and answer sessions, (b) documentation of EIBF in the safe birth checklist along with mention of the interval between delivery and initiation of breastfeeding, (c) building a separate cubicle for postpartum mothers for breastfeeding counseling, (d) display of posters and charts depicting the importance and advantages of EIBF, and (e) counseling sessions for the antenatal women and their birth companions in the labor room when they got admitted with spontaneous onset of pain or for induction of labor. After the collection of baseline observations, a second team meeting was conducted and the possible reasons for the low prevalence of EIBF were discussed in detail. A conceptual framework of the possible reasons for low EIBF was noted and the same was plotted in the form of a fish-bone analysis chart  monthly training of the doctors and nursing staff was conducted through dedicated lectures, interactive question, and answer sessions; (b) documentation of EIBF in the safe birth checklist along with mention of the interval between delivery and initiation of breastfeeding; (c) building a separate cubicle for postpartum mothers for breastfeeding counseling; (d) display of posters and charts depicting the importance and advantages of EIBF; and (e) counseling sessions for the antenatal women and their birth companions in the labor room when they got admitted with spontaneous onset of pain or for induction of labor.Outcome measures were assessed by recording the monthly percentages of newborns breastfeeding within one hour of delivery , to assess the effect of interventions. Outcomes were noted over a period of six months. During this time, monthly training of the doctors and nursing staff was being undertaken regularly (on the first of each month). Data collectors  maintained records of the total number of deliveries conducted in adolescent women and the number of cases where EIBF was initiated. In the post-intervention follow-up phase, which lasted for six months, only observations were done without any intervention. All data were entered in a Microsoft Excel matrix and analyzed using IBM Statistical Package for Social Sciences Statistics, version 20.0.0 . Categorical variables were expressed in percentages and frequencies. The monthly rate of EIBF expressed in percentages was plotted in a line chart for better representation and comparison over the period of time.Overall 122 adolescent deliveries were included in the study, starting from the pre-intervention phase to the post-intervention phase. The prevalence of EIBF over the three months of pre-intervention baseline phase was only 28.5% (Figure This QI initiative aimed to improve the EIBF among adolescent mothers at a tertiary care hospital using the standard techniques of root cause analysis and PDSA cycles. A QI initiative conducted to improve early breastfeeding among newborns delivered in a secondary care hospital in Northern India increased the prevalence of EIBF from 52% to 97% . The preLike other QI initiatives which showed positive outcomes, our study also proved effective in improving EIBF despite the presence of so many hurdles. There were no proper hospital policies for EIBF, no documentation of timing of breastfeeding post-delivery, and no designated personnel to insist and supervise EIBF. With our QI initiative, nursing staff and resident doctors were trained and encouraged to insist on EIBF under the supervision of the nursing in charge of the postnatal wards. Dudeja et al. employed the same model of QI initiative which improved the first-hour breastfeeding after cesarean delivery from zero to 93% over a period of three months . The PDSThe strength of this study was that it included consultants, resident doctors, and nursing staff as a designated team undergoing rigorous training every month. All were involved in learning and teaching through active participation. It involved simple but most effective measures to improve the EIBF which can be employed in all healthcare centers with low-resource settings. The major limitation was it was single-centered. Owing to the huge patient load, maternal satisfaction could not be assessed in this study. Moreover, counseling of the family members and birth attendants could not be done effectively during the study period.This QI initiative was very effective in improving EIBF among adolescent mothers, thereby reducing neonatal morbidity and mortality. A designated team involving obstetric consultants, resident doctors, and nursing staff could improve breastfeeding initiation and overall maternal behavior among teenage mothers. The employment of simple steps by displaying posters and charts depicting the advantages of EIBF has proven effective in our hospital setting. It can be employed in low-resource health settings where teenage pregnancies are still a burden to sustainable development of women.


Patient-centred care is an important part of quality healthcare and patient satisfaction has been shown to be associated with improved clinical outcomes. We aim to explore the satisfaction of patients with diabetic kidney disease (DKD) with their visits to the TCM physician and its association with patients' socio-economic characteristics.A questionnaire survey was conducted among patients aged >21 years with DKD. Participants' demographic, socioeconomic characteristics and satisfaction scores measured with the self-administered Medical Interview Satisfaction Scale (MISS) were collected after they visited the TCM physician. MISS is a 26-item questionnaire consisting of three domains – cognitive, affective and behavioural which was developed to assess patient satisfaction with medical consultation. Independent samples t-test and one-way analysis of variance (ANOVA) were used to analyse the data.137 participants completed the questionnaires and were included in the analysis. The mean satisfaction score was 3.1 out of 5, with the cognitive domain being significantly higher compared to the affective and behavioural domains. The mean satisfaction score of the cognitive domain differed significantly among participants staying in different types of housing and those with previous TCM encounters. The mean satisfaction score of the behavioural domain differed significantly among participants of different ethnicities. The mean satisfaction scores of all the domains were also significantly different among participants with different duration of follow-up with their TCM physicians.We found that ethnicity, types of housing, previous TCM experience and the duration of follow-up with the TCM physician may affect the satisfaction scores of patients with DKD. •Patient perspectives and satisfaction have been shown to be associated with improved clinical outcomes.•Patient satisfaction is multidimensional and consists of domains related to cognitive, affective and behaviour.•Communication, addressing patients' concerns and encouraging greater patient participation in healthcare encounters are important.•Ethnicity, type of housing, previous TCM experiences and the duration of follow up affect patient satisfaction. Patient satisfaction; Diabetic kidney disease; Traditional Chinese medicine The prevalence of DKD in patients in the local outpatient setting has been reported to be 53% . DKD is Management of DKD is challenging and a holistic approach is required for patient care. Conventional medicine is disease-targeted and has been proposed to be less interpersonal compared to complementary and alternative medicine (CAM) which concentrates on the harmony and restoration of balance between the body and mind , 4. PatiThe measurement of patient satisfaction has attracted growing attention and interest in recent decades. With the increasing emphasis on patient-centred care, more healthcare regulators are incorporating patient satisfaction as a key component of quality assessment , 10, 11.Traditional Chinese medicine (TCM) is one of the oldest forms of CAM that is commonly practised worldwide and is highly regarded in Asian societies such as Taiwan, Hong Kong and Singapore. Even though both Western medicine and TCM are widely available in Singapore, there is a lack of integration and corroboration between TCM and the standard healthcare system. Some studies showed that patients choose to visit TCM as they are not satisfied with the care and treatment outcomes provided by allopathic medicine . However22.1This prospective study was conducted at Singapore General Hospital (SGH) and Singapore Thong Chai Medical Institute from 18 May 2015 to 7 Dec 2016. Electronic medical records of the participants were reviewed and those that satisfied the inclusion and exclusion criteria were invited to participate in the study. Eligible participants were patients above 21 years of age, with type II diabetes mellitus (T2DM) and chronic kidney disease (CKD) stage II to V. Patients with dementia, recent increase in serum creatinine of more than two-fold, non-diabetic kidney disease, renal replacement therapy, chronic liver diseases, or severe cardiac failure were excluded. Eligible participants were scheduled for their TCM consultation at Singapore Thong Chai Medical Institute. The purpose of the TCM visit was to diagnose and assess the severity of DKD and no treatment was given .Hong Chang Tan was supported by Data will be made available on request.The authors declare no competing interests.No additional information is available for this paper.


Dealing with uncertainty is one of the critical topics in health technology assessment. The greater decision uncertainty in appraisals, the less clear the clinical- and cost-effectiveness of the health technology. Although the development of targeted cancer therapies (TCTs) has improved patient health care, additional complexity has been introduced in drug appraisals due to targeting more specific populations. Real-world data (RWD) are expected to provide helpful information to fill the evidence gaps in appraisals. This study compared appraisals of TCTs with those of non-targeted cancer therapies (non-TCTs) regarding sources of uncertainty and reviewed how RWD have been used to supplement the information in these appraisals.This study reviews single technology appraisals (STAs) of oncology medicines performed by the National Institute for Health and Care Excellence (NICE) over 11 years up to December 2021. Three key sources of uncertainty were identified for comparison . To measure the intensity of use of RWD in appraisals, three components were identified .p = 0.026), whereas mature survival data were available in fewer TCT appraisals (p = 0.027). Both groups showed similar patterns of use of RWD. There was no clear evidence that RWD have been used more intensively in appraisals of TCT.TCTs received more recommendations for provision through the Cancer Drugs Fund , whereas similar proportions were recommended for routine commissioning. With respect to sources of uncertainty, the external validity of clinical trials was greater in TCT appraisals (Some differences in uncertainty were found between TCT and non-TCT appraisals. The appraisal of TCT is generally challenging, but these challenges are neither new nor distinctive. The same sources of uncertainty were often found in the non-TCT appraisals. The uncertainty when appraising TCT stems from insufficient data rather than the characteristics of the drugs. Although RWD might be expected to play a more active role in appraisals of TCT, the use of RWD has generally been limited.The online version contains supplementary material available at 10.1186/s12885-022-10350-8. In England, the National Institute for Health and Care Excellence (NICE) has a role in assessing health technology, such as drugs and medical devices, in informing the best value of using the National Health Service (NHS) resources. Cost-utility analysis is the primary method to assess value for money in appraisals of cancer treatments. Uncertainty is unavoidable when appraising the clinical- and cost-effectiveness of new drugs. Uncertainty refers to the fact that we do not know the expected costs and effects of an intervention in a particular population of patients with absolute precision  — the moTargeted cancer therapy (TCT) refers to treatments that act on specific molecules associated with cancer growth, progression and spread guided by biomarker results . Lung caReal-world data (RWD) are suggested as a means of overcoming evidence gaps and helping appraisal of innovative drugs in light of the challenges of obtaining the required information from randomised controlled trials (RCTs) , 12. ForThe uncertainty in appraisals is one of the significant concerns in HTA decision-making. RWD has received attention as a means of reducing uncertainty. However, there are caveats with using RWD due to confounders, biases and data quality . Also, in = 229). NICE technology appraisal guidance is publicly available (https://www.nice.org.uk/guidance). The appraisals were manually screened to identify the relevant appraisals. This study uses data extracted following a protocol developed to record information about the use of RWD in NICE appraisals of oncology medicines [This study compared single technology appraisals (STAs) of TCT and non-TCT in terms of appraisal recommendations, the size of clinical trials, types of uncertainties and use of RWD. Chi-square tests were used to show whether any differences between TCT and non-TCT were statistically significant. This analysis includes NICE STAs of oncology medicines for which guidance was issued between January 2011 and December 2021 . Any STAs issued after TA399 were regarded as ‘after 2016 CDF’.The size of clinical trials was also reviewed in this study. The number of patients included in the trials was summarised in a histogram to look at the distribution of the trial size. Kernel Density estimation was used to approximate the histogram with a continuous distribution. This estimation compared the similarities and differences between TCT and non-TCT appraisals, focusing on the average number of patients in the trials.questionable external validity. External validity is considered moderate if the ERG raises a few minor concerns. A comment such as “younger and fitter patients” without mentioning performance status is classified as a minor concern. External validity is classified as acceptable if there are no specific critiques.This study focuses on three potential sources of uncertainty in NICE appraisals: the external validity of clinical trials, the availability of direct treatment comparisons, and the maturity of survival data. The sources of uncertainty identified by Morrell et al.  were claThe type of treatment comparison made by manufacturers in their evidence submissions is reviewed to identify the availability of direct treatment comparisons. A sixfold classification of treatment comparisons in NICE appraisals can be made using the information on the availability of head-to-head comparison for all comparators, indirect treatment comparison, anchored/unanchored treatment comparison and population-adjusted treatment comparison. The possible combinations of treatment comparison are presented in Additional file Lastly, the maturity of survival data is highlighted as a source of uncertainty. This study uses three categories  based on the percentage of death events in the primary clinical studies. 20 and 50% were used in this study to classify appraisals, adapting the findings from Tai et al. . If the There are many potential uses of RWD in an appraisal and several ways of reporting the use of RWD. Simple counts of the number of occasions when RWD are used in an appraisal may not be a good guide to how differently one appraisal utilises RWD compared to another. This study used a few different methods, such as pattern review and intensity analysis, to review the use of RWD. Figure The patterns of use of RWD were reviewed to provide a clearer picture of how RWD have been used. The extraction protocol distinguished 31 economic evaluation components where RWD might be used, giving rise to many different patterns. The patterns were reviewed by distinguishing between the parametric and non-parametric use of RWD. Parametric use involves basing the numerical value of specific variables in the economic model on RWD. For example, the use of data to provide values for overall survival (OS) or resource use in the economic model is categorised as parametric use. Non-parametric refers to using RWD to develop the model structure and support or validate assumptions in the model. Using RWD to select comparators or validate the survival distribution choice are examples of non-parametric use. This separation provides a more comprehensive review of how RWD have been used in appraisals. All components where RWD could be used are presented in Additional file The intensity of use of RWD in different appraisals was investigated by classifying different patterns in terms of the extent to which RWD are drawn upon in different economic evaluation components. Three components  are identified as major uses of RWD, which are likely to have a high impact on the outcome of the economic evaluation, the incremental cost-effectiveness ratio (ICER). The remaining components are regarded as minor uses of RWD. The identified patterns were categorised into seven groups by distinguishing major and minor uses of RWD Fig. . Two claSingle Technology Appraisals of TCT and non-TCTn = 229). The number of STAs of oncologic medicines has generally increased over time except for 2019 and 2020. Of included STAs, 36% were TCT appraisals. Although there were fluctuations, the TCT proportion has increased over time. The highest proportion of TCT appraisals was in 2019–57% of oncology appraisals. Note there were no TCT appraisals published in 2011.Figure Figure The TCT and non-TCT appraisal recommendations are reported in Table The number of patients in the clinical trials upon which treatment effectiveness in the economic models was based was reviewed to compare the sizes of the overall trials between TCT and non-TCT. Most clinical studies had fewer than 1000 patients. Right skews were found Fig. A. These 2.Sources of uncertainty in NICE appraisalsPotential sources of uncertainty are summarised in Table The external validity of the clinical study2)Types of treatment comparison in manufacturer submissionsThe uncertainties concerning external validity raised in the appraisals were reviewed. These factors  are usually addressed in the ERG reports when assessing the generalisability of trial outcomes to NHS practice. Twenty-seven appraisals were identified, where the ERG highlighted the high level of uncertainty with respect to the external validity of the clinical evidence. Ten of these appraisals were TCT. Problems were identified with respect to the study population (70%), the comparators (20%) and subsequent treatment received by trial participants (10%). In appraisals of non-TCT, the external validity of evidence was heavily questioned in seventeen appraisals. The main reason was the study population (53%), followed by the issue of subsequent treatment received by trial participants (35%). The general problem of trial populations being younger and fitter than routine practice is widely noted by ERGs. However, this was not a major reason for the high level of uncertainty unless subgroups in the trial were very different from those in routine practice. More often, the issues with respect to the study population arose from differences in prior treatment, which might impact survival outcomes. For example, in an appraisal of nivolumab (NICE TA530), the ERG expressed serious concerns regarding the representativeness of the trial population to the UK population. One of the reasons was a mismatch of prior therapies. More than 75% of patients in UK clinical practice received a previous gemcitabine platinum-based therapy, while less than 40% of the trial population did. Another example is an appraisal of durvalumab (NICE TA578). The ERG identified that the population in the clinical trial (PACIFIC) was narrower than in the scope (patients expressing PD-L1 > 1%). Also, they received different types of chemoradiation therapy cycles. UK patients received sequential rather than overlapping treatment, potentially affecting the treatment effect.The treatment comparisons made were not statistically different between TCT and non-TCT appraisals. The availability of head-to-head RCTs was reviewed to understand the patterns of indirect treatment comparison. The proportion of single-arm trials in TCT appraisals is higher than that of non-TCT. Nineteen TCT appraisals did not use RCTs as primary clinical evidence . Several possible ways to compare treatments were found in these appraisals Fig. . In gene3)Maturity of survival data in clinical trialsThe maturity of survival data showed a statistical difference between TCT and non-TCT appraisals. The proportion using extremely immature survival data was similar between the two groups, whereas immature survival data were used more in TCT appraisals. The changes in the use of extremely immature, immature and mature survival data over time are shown in Fig. 3.The use of RWD in the economic models of TCT and non-TCTPattern reviewThere is no dominant pattern of use of RWD in these appraisals. Fifteen different patterns of use of RWD can be identified, which appeared in three or more appraisals. These patterns cumulatively account for 51% of all appraisals Intensity analysisWhen looking at the non-parametric and parametric use of RWD separately, more diverse patterns were found for parametric use than for non-parametric use. Sixty-two per cent of all appraisals involved no non-parametric use of RWD , classification B does not provide a significant difference . Over time, the major use of RWD has increased in both groups of appraisals. In 2020, about 60% of TCT and non-TCT appraisals made at least two major uses of RWD. The cases of three major uses of RWD were observed in the non-TCT group in 2018. Such a major use of RWD was not observed in the TCT group. Using classification A  inhibitors  are good examples of likely future challenges. In these appraisals, the main clinical trials were basket trials, which is a novel trial design to evaluate the treatment effectiveness of TCT for one or more targets regardless of the pathology . Also, cTo date, the targeting of treatment populations has not introduced significantly different appraisal challenges. However, the next generation of TCT, such as histology-independent therapy, might present more decision-making challenges, including identifying the eligible population and appropriate prices across the different populations in the future . OverallUncertain clinical outcomes due to immature survival data are commonly encountered in NICE appraisals . The immThe limited availability of direct treatment comparisons was identified as a source of uncertainty across appraisals. Regardless of the treatment type, obtaining head-to-head estimates of comparative effectiveness from a single trial becomes more challenging since the treatment options are rapidly expanding. When direct treatment comparison is not available in a trial, network meta-analysis has been used to identify the treatment effect indirectly. However, a network is not always available unless a common comparator links the available trials . The indThis study found that the evidence used in appraisals of new cancer drugs was uncertain across both TCT and non-TCT appraisals. The sources of uncertainty observed in TCT appraisals were not essentially different from those in appraisals of non-TCT. The uncertainties decision-makers face are ones they have faced previously. Given the novelty of targeted therapy, a new approach was required, such as an innovative clinical trial design and strategy for early decision-making to improve operational efficiency . HoweverRWD have been identified as supplementing RCT data. As the pattern review showed, RWD were used in diverse ways. However, while many are optimistic about the potential contribution of RWD , the useAlthough limited use was made of RWD, it is notable that the intensity of use of RWD has increased over time. Among the patterns that appeared in three or more appraisals, five patterns included using RWD for estimating OS. It is a noteworthy result given the strong signal of NICE’s interest in the use of RWD . AlthougThis study explored several aspects of appraisals of TCT and non-TCT from an HTA perspective. Given the increased interest in using biomarkers to identify treatment groups, there will likely be growing challenges in appraising TCT. Although the findings of this study could change over time as more TCT are developed, this study is the first to document systematically the differences and similarities in sources of uncertainty and use of RWD between appraisals of TCT and non-TCT by reviewing over two hundred appraisals. However, this study has a few limitations. First, the information about external validity relies on the ERG reports. Although appraisal committees agree with ERG’s assessments in general, committees do not necessarily always agree on all points with ERGs. What committees critically emphasise regarding external validity could be different.Another limitation is the classifications of uncertainties and intensity of use of RWD. Although all the information used in this study was obtained from appraisal documents, how to categorise this information was based on the data extraction protocol. The maturity of survival data was classified using two values, 20 and 50%. However, these points are not agreed criteria to define data maturity. Committees can make different judgements with respect to maturity. With respect to the intensity classification, this study focuses on a specific assumption that the use of RWD in three major components would be intensive use of such data. However, the criteria to measure intensity are not universally agreed upon. Also, decision-makers might not be concerned about which RWD inform components of the economic model. More likely, they would concentrate on how RWD would help to address the decision problem. How to classify uncertainty and intensity of use could differ across researchers and decision-makers.Finally, it is noted that there might be a difference in the number of appraisals depending on which criteria were used. In this study, the STAs for treating side effects of cancer drugs were excluded. When appraisals are collected, potentially also affects their number. Some appraisals available in this study might not be available later due to the replacement of appraisals  Likewise, previously available appraisals might not be included in this study as the guidance was withdrawn. Despite this potential difference, this study included all STAs of cancer therapy which were available as of December 2021.Some differences in uncertainty were found between TCT and non-TCT appraisals. The appraisal of TCT is generally challenging, but these challenges are neither new nor distinctive. The same sources of uncertainty were also often found in the non-TCT appraisals. The uncertainty in appraising TCTs is more likely to stem from insufficient data rather than the inherent characteristics of the drugs. Although RWD might be expected to take a more active role in appraisals of TCT, the use of RWD has generally been very limited.Additional file 1.


Diving implies cardiovascular adaptations in marine mammals. The work aimed to analyze the aortic wall in nine cetacean species with deep diving habits belonging to four families. We hypothesize that variations in the aortic wall will reflect the diving capability of each species. The analyses showed that the elastic component was higher in the initial part of the artery, the muscular elements increased in the distal portion in all cases, and the apparent organization of the components in the aortic wall did not show essential modifications between the four families. All species presented a decrease in the arterial wall thickness along the aorta. The reduction was dramatic between the initial and thoracic aorta in the sperm whale specimens; meanwhile, the species of the other three families, beaked whale, pygmy sperm whale, and deep-diving dolphins, showed a more uniform decrease between the aortic segments. Because of the diving habits described for the different species, our findings support that a decrease in the arterial wall thickness between the aortic segments is the most relevant characteristic related to diving adaptation.This study analyses the aortic wall structure in nine cetacean species with deep diving habits belonging to four Odontoceti families: Ziphiidae, Kogiidae, Physteridae, and Delphinidae. Samples of ascending, thoracic and abdominal aorta were processed for histological and morphometric studies. The elastic component was higher in the proximal aortic segments, and the muscular elements increased distally in all cases. Morphometric analyses showed that all families presented a decrease in the thickness of the arterial wall and the tunica media along the aorta. The reduction was dramatic between ascending and thoracic aorta in the Physeteridae specimens; meanwhile, the other three families showed a more uniform decrease between the ascending, thoracic and abdominal aorta. The decline was not correlated with a reduced elastic or lamellar unit thickness but with a loss of lamellar units. The organization of the elements in the aortic wall did not show essential modifications between the four families, resembling the structure described previously in the shallow and intermediate diving dolphins. Our findings support that the difference in the morphometric characteristics of the different segments in the aortic wall is likely related to the diving habit more than the absolutes values of any other parameter. The critical bradycardia and peripheral vasoconstriction during diving suggest a need for cardiovascular system adaptations in marine mammals. Studies on the properties of the aorta in some marine mammals, primarily pinnipeds and some species of toothed whales, showed macroscopic and morphometric differences in the dimensions and structural wall composition in the ascending and other segments of the aorta ,2,3,4,5.Mesoplodon densirostris [De Blainville 1817], Mesoplodon bidens [Sowerby 1804], Mesoplodon europaeus [Gervais 1855], Ziphius cavirostris [Cuvier 1823]), Kogiidae (Kogia breviceps [De Blainville 1828] and Kogia sima [Owen 1866]), Physeteridae , and Delphinidae . Most of these families are characterized by deep-diving species, except for the Delphinidae family, composed mainly of shallow or intermediate divers. However, Globicephala macrorhynchus and Grampus griseus are deep divers, so we will refer to them as deep-diving species of the Delphinidae family. Animals studied belonged to different sex and age categories and were stranded along the Canary Islands coasts between 2008 and 2020  and anti-von Willebrand factor (RRID: AB_2811207) antibodies were visualized using a Chemmate Dako EnVision detection kit (Glostrup-Denmark). The Peroxidase™/DAB, Rabbit/Mouse System  was used following the manufacturer’s instructions. Sections of human aortas were used as positive controls for actin. Vascularized pheochromocytoma tumors from chromaffin cells of the adrenal medulla with large micro-vessels sections were used as positive controls for the von Willebrand factor. The tissue bank of the Pathologic Anatomy Service from the Complejo Hospitalario Insular Universitario Materno Infantil de Las Palmas de Gran Canaria (Spain) provided the human positive control samples for immunohistochemical positive controls. An antihuman monoclonal anti-actin alfa-smooth muscle antibody produced in mice was used at a 1:2000 dilution for actin immunodetection. For the von Willebrand factor, immunodetection was performed with an antihuman polyclonal anti-factor VIII-related antibody produced in rabbits at a 1:100 dilution. In both cases, endogenous peroxidase was blocked for 30 min with 3% hydrogen peroxide in methanol. The samples were analyzed with a Labophot-2 Nikon microscope. Images were captured with a Sight DS-5M digital camera.The 4× and 10× measurements of the aortic wall were performed in randomly selected fields of one Orcein or one Masson trichrome stained histological section of each case, using a Digital Sight Camera Control Unit coupled to a Labophot-2 Nikon microscope. In addition, four different measurements were performed directly in each histological section using digital camera tools. At low magnification (4×), the thickness of the tunica intima, tunica media, and total wall (IM) were measured. The tunica adventitia was not measured because it was impossible to determine its outer limit in all cases. The tunica media thickness was considered as the perpendicular distance between the innermost and outermost elastic lamina. The thickness was measured at four equidistant points within the same aortic ring section. The mean value of the four measurements was calculated. The elastic laminas’ thickness and the tunica media’s lamellar unit thickness were measured separately at higher magnification (10×). The size of the lamellar unit thickness included smooth muscle cells (smcs), the elastin layer, collagen, and the extracellular matrix. We counted the thickness of the lamellar unit in the transversal section from the adluminal side of a smooth muscle cell to the abluminal side of the elastic layer that limits the luminal side of the following smooth muscle cell. Two of the four measurements were performed in the inner part of the aortic wall and two in the outer. Again, the mean value of the four measurements was calculated. The lamellar unit in the abdominal aorta was measured, avoiding the sizeable longitudinal muscle bundles in the inner part of the tunica media. The number of lamellar units was calculated considering each sample’s total wall thickness and the lamellar unit thickness in each sample.Mesoplodon densirostris, Mesoplodon bidens, Mesoplodon europeaus, Ziphiius cavirostris), Kogiidae (Kogia breviceps and Kogia Sima), Physeteridae , and deep-diving species of the Delphinidae family . Statistical data analysis was performed by the IBM SPSS Statistics 27 . Categorical variables were expressed as frequencies and percentages, and numerical variables were summarized by mean ± standard deviation (SD). Three means of independent samples were compared using the non-parametric Kruskal–Wallis test due to non-compliance with data normality by using the Shapiro–Wilk test. The results were considered statistically significant if the p-value < 0.05.For statistical purposes, the species were grouped and compared within families as follows: Ziphiidae  and is not directly translational to living organisms. However, comparative analyses under the same conditions should render translational differences in living organisms. Given the ethical, logistical, and technical difficulties of obtaining these measurements from these cryptic and elusive animals, studies on dead-stranded animals with the same decomposition code represent a fair approximation and a first step in understanding the vascular morphological adaptations of marine mammals to dive.The findings of this work point out that the decrease in the IM and tunica media thickness and the loss of lamellar units between the ascending and thoracic aorta may be the most representative of aortic wall morphometric and morphological adaptation to diving among the analyzed parameters. The higher thickness and elasticity of the proximal segment would allow more elevated and more sure storage of systolic flow during diving; meanwhile, the thinner and less elastic thoracic aorta would allow a more continuous peripheral circulation of the diastolic flow during diving. Therefore, the difference between these two segments could imply better capacitation for diving.


The interfacial interface between fibers and matrix plays a key role for epoxy matrix composites and short recycled randomly arranged fibers. This study used short recycled carbon fiber (RCF) as a filler. Plasma treatment was used for carbon fiber surface treatment. This treatment was performed using radio (RF) and microwave (MW) frequencies at the same pressure and atmosphere. Appropriate chemical modification of the fiber surfaces helps to improve the wettability of the carbon fibers and, at the same time, allows the necessary covalent bonds to form between fibers and the epoxy matrix. The effect of the plasma treatment was analyzed and confirmed by X-ray photoelectron spectroscopy, Raman microscopy, scanning electron microscopy, transmission electron microscopy and wettability measurements. Composite samples filled with recycled carbon fibers with low concentrations  and high concentrations (20 wt% and 30 wt%) were made from selected treated fibers. The mechanical properties  were analyzed on these samples. It was found that the modulus of elasticity and bending stress increase with the increasing content of recycled carbon fibers. A more significant change in impact strength occurred in samples with low concentration. In recent years, the production of carbon fiber-filled composites has increased significantly, and thus the topic of recycling these materials has become increasingly important . When deFurthermore, this study analyzes the mechanical properties of epoxy composites reinforced with recycled short carbon fibers (RCF) with and without plasma treatment. Plasma treatment usually increases the adhesion between fibers and matrix, and the incorporation of the fibers into the matrix provides an ideal reinforcement effect. The mechanical properties of the RCF-filled composites were analyzed using impact toughness and flexural strength tests. These tests were chosen because, based on previous knowledge of short fiber reinforced composites, it can be assumed that the tested specimens have low ductility. In tension, the applied tensile force would act primarily on the matrix of the material, and short fiber reinforcement would not significantly help the tensile strength.2 or He or Ar, and the by-products allow the formation of bonds to adjacent chains  are shown in Figure 8. The following formula was used for the calculation:h [m] is thickness and b [m] is width of the composite samples.The flexural strength was measured according to EN ISO 14125. A schematic of the test equipment is shown in F [N] is the loading force and l [mm] is the distance between the supports. To calculate the elastic modulus:s is the deflection [mm].SEM images. Together with XPS analysis, these images helped select a suitable filler type for the fabrication of the composite samples. Comparing the SEM images of RCF in Comparing the SEM images of RCF, TEM images. Due to the nature of TEM imaging, we focused on the ends of the fibers, where details of the longitudinal arrangement of the carbon fiber surface can be seen. No significant changes were observed between fibers with and without plasma treatment at the fiber ends. In Measurement of wettability. The contact angle of WCA recycled carbon fiber is affected by plasma action. Due to the roughness of the surface, the measured contact angle is only indicative of the plasma treatment trend. The measured values show that plasma treatment of CF has a significant effect on the change of hydrophilic properties. Untreated fibers show hydrophobic behavior, while plasma treated fibers show hydrophilic behavior. The increased hydrophilicity of the fibers is probably due to the binding of oxygen functional groups to the fiber surface, together with the higher surface roughness caused by the plasma treatment.However, in general, roughness is considered a relatively unimportant factor causing contact angle hysteresis . The hysComparing the WCA results shown in Raman spectroscopy was used to assess the effect of plasma treatment on the structural changes of carbon fibers. The Raman spectra (RS) for different types of carbon structures show two peaks at approximately 1355 and 1580 cm−1, corresponding to the defective carbon (D band) and graphite mode (G band) [(G band) . Figure (G band) . RecycliXPS analysis. The plasma treatments modify surface of carbon fibers significantly. The main observed effect is functionalization of fibers by introducing oxygen functionalities on the surface. The plasma treatment increased oxygen content twice from 10% on untreated fibers up to approximately 20% on plasma treated in both types of discharges. The nitrogen content is not significantly changed by the plasma treatment.The detailed study of chemical bonding on the surface was made on high resolution photoelectron spectra, see Literature reported Raman analyses showed both G and D bands typical for sp2 and defects/sp3 hybridization . TherefoThe nitrogen related N 1 s peek has two components  located The oxygen photoelectron peek O1s can be successfully fitted by three components. Where the component at lowest energy 531 eV is O=C, component at 532.9 eV is O–C similarly as in polymers , the finCharacteristics of fracture surfaces. SEM analyzed the surface of the fracture surfaces after impact toughness measurements. After impact toughness measurements on FRE composite specimens, the fracture surface images obtained by SEM were analyzed. The dispersion state of the recycled carbon fibers in the epoxy resin was evaluated. Impact toughness of FRE composites. The impact toughness results are shown in Three-point bending. The three-point bending results are shown in  Flexural strength is the ability of material to resist a bending force applied perpendicular to the longitudinal axis of the composite specimen. The stress induced by the bending load is then a combination of compressive and tensile stresses [stresses . Figure Fiber-reinforced polymers, particularly carbon fiber reinforced epoxies, are expected to account for more than 50% of the structural weight of motor vehicles in the future. As the number of applications increase, the amount of ancient carbon fiber structures will grow, and recycling of these materials will become more important . TherefoThis study analyzed the mechanical properties (represented by impact toughness and three-point bending) of epoxy composites filled with short RCFs. Two types of plasma treatment were used to modify the RCF surface. These treatments were performed using radio frequency (RF) and microwave (MW) radiation at the same pressure and atmosphere. WCA measurements and XPS analysis confirmed that plasma treatment changes the surface layers of carbon fibers, as reported in studies ,11. The The suitability of plasma treatment was verified in mechanical tests performed on composite samples prepared from these fibers. It was found that the modulus and bending stress increased with increasing recycled carbon fiber content, as also reported in the study ,11. HoweFurthermore, in this work, the spectrum of different concentrations of epoxy-filled RCF composites has been analyzed. The increase in the adhesion between fibers and matrix is due to the increase in the surface roughness of the fibers and the oxidation of their surface. For impact toughness, it was found that there is an optimum limit. A suitable balance between elasticity and toughness was achieved for composite samples with 1 wt% RCF. and both types of plasma treatment further improved the tested mechanical properties by this concentration of RCF. However, RF plasma treatment of the fibers had a more significant effect on the improvement of impact toughness and modulus.The selected RF plasma treatment procedure is readily applicable for plasma treatment of short recycled fibers and represents a significant advantage (for the lowest concentration samples) over the use of untreated RCF. FRE composites reinforced with plasma-modified RCF fibers can be obtained for various applications in the composites industry.


Cattle productivity in tropical and subtropical regions can be severely affected by the environment. Reproductive performance, milk and meat production are compromised by the heat stress imposed by the elevated temperature and humidity. The resulting low productivity contributes to reduce the farmer’s income and to increase the methane emissions per unit of animal protein produced and the pressure on land usage. The introduction of highly productive European cattle breeds as well as crossbreeding with local breeds have been adopted as strategies to increase productivity but the positive effects have been limited by the low adaptation of European animals to hot climates and by the reduction of the heterosis effect in the following generations. Gene editing tools allow precise modifications in the animal genome and can be an ally to the cattle industry in tropical and subtropical regions. Alleles associated with production or heat tolerance can be shifted between breeds without the need of crossbreeding. Alongside assisted reproductive biotechnologies and genome selection, gene editing can accelerate the genetic gain of indigenous breeds such as zebu cattle. This review focuses on some of the potential applications of gene editing for cattle farming in tropical and subtropical regions, bringing aspects related to heat stress, milk yield, bull reproduction and methane emissions. Tropical and subtropical regions are home of about 40% of the world’s human population and where nations with the highest growth rates and poorest populations are located . These rIn an attempt to improve cattle productivity, breeds from European origin have been introduced in Africa, Asia and Latin America. However, the introduction of breeds that were proved successful in developed nations located in regions of temperate climate usually results in lower efficiency because of the heat stress, low pasture quality  and paraGene (or genome) editing tools have been developed and improved in the last two decades. These tools allow the precise introduction of mutations in a given gene , for whaThe applications of the CRISPR system in different fields of biology has been shown in several reports . In cattAssisted reproductive technologies such as somatic cell nuclear transfer  or in vitro fertilization (IVF) are required to generate gene-edited embryos. When IVF is applied, a large number of gene-edited animals can be produced in one or two generations by commercial in vitro fertilization laboratories. The gene-edited embryos can be biopsied so that the genomic evaluation can be performed in order to select the ones with high estimated genomic values before transferring them into recipients. Thus, gene editing together with assisted reproductive technologies and genomic selection can play a major role in genetic breeding programs by either reducing the generation interval, increasing selection intensity and accuracy, and/or by increasing genetic variation .In this review we will focus on how cattle farming can benefit from gene editing technologies in the tropical and subtropical regions. This review will cover aspects related to gene editing applications regarding heat stress, milk yield and composition, bull reproduction and methane emissions.Bos taurus cattle from central and south America such as Senepol, Romosinuano, Criollo Limonero and Carora have been selected for adaptation to tropical conditions. One of the first studies examining the thermotolerance of B. taurus cattle from the tropics was performed by Hammond et al. in subtropical Florida to compare the rectal temperature (RT) of Senepol, Angus, Hereford and Brahman cattle during summer  responsible for the short (slick) hair coat phenotype  compared to wild-type females . For that, a single strand ODN (ssODN) donor template designed with one of the SLICK mutations can be used. As the HDR is not the usual DSB repair pathway employed by cells, the chances to introduce some of the SLICK mutations are limited. However, as the different mutations of SLICK alleles found in the different breeds are located between BTA20:39099113 and BTA20:39099321 positions (ARS-UCD1.2 genome assembly)  , it is lSLICK animals can be generated from embryos produced by nuclear transfer performed with gene-edited somatic or embryonic stem cells , or from in vitro fertilized zygotes injected or electroporated with CRISPR/Cas9 . The aniIt has been challenging to maintain efficient dairy production systems in tropical and subtropical regions. Because of heat stress, high productivity European breeds cannot produce milk as they usually do in temperate climates. On the other hand, despite the adaptation to the tropical environment, local indigenous (or adapted) breeds have low productivity, in part due to the lack of well-established genetic improvement programs. Heat stress also affects milk composition. Protein and fat content can be reduced, which may alter the coagulation properties of the milk used to make cheese, affecting cheese yield .GHR) gene. One of the actions of the growth hormone (GH) is to stimulate milk and protein production, which can occur indirectly through systemic changes, such as food intake, blood flow and nutrient delivery to the mammary gland . The resulting allele (Y) is associated with higher milk production  , althoug content  but also content . The K a content . The A a content , which i content . It was  content . The A a content , and the content . The A a content . In cont content . Low fre content . As resuSCD1 gene that results in an alanine to valine substitution in the enzyme and it is associated to changes in milk fatty acid composition in Holstein cows  is an enzyme responsible for fatty acid desaturation in the mammary gland and other tissues, playing an important role in lipid metabolism of mammary tissues by introducing a cis double bond at the C-9 position of a wide range of fatty acids . The preein cows . Milk fa:1 cis-9 , and alseic acid , althougeic acid . The freeic acid  and Jerseic acid  breeds,  in milk .GHR , DGAT1  and SDC1  genes are the result of point mutations presented in higher frequency in European dairy breeds compared to indigenous breeds, and are associated to milk yield and fat content. Thus, those point mutations are potential candidates to increase the milk yield and improve milk composition in adapted indigenous breed in tropical and subtropical zones, such as zebu dairy breeds. The introgression of those alleles in the genome of dairy breeds can be performed by gene editing, preserving other racial features of indigenous breeds.As shown above, the alleles found in The HDR mechanism is required to insert these point mutations into the genome. For that, the DSB caused by the Cas9 enzyme (or other nucleases) can be repaired using a ssODN donor template designed with the target mutation and homology arms (upstream and downstream of the mutation site). Moreover, the donor template must contain a silent mutation to avoid the re-cut of the repaired DNA by the Cas9. Finally, to increase the chances of successful HDR, the DNA cleavage site needs to be as close as possible from the mutation insertion site . The proStrategies can be employed to increase the chances of HDR over the NHEJ mechanism for DSB repair. Some small molecules can act to inhibit NHEJ while others can be used to stimulate HDR. One of the NHEJ inhibitor molecules is SCR7, which acts by inhibiting the DNA ligase IV enzyme, necessary for double-strand break repair . In miceThe high temperature and humidity index typical of tropical and subtropical climates alters the behavior of breeds from European origin; examples of behavioral changes are a decrease in dry matter intake and the seeking for shade . MoreoveNANOS2 gene in pigs to generate male offspring without germline cells (spermatogonia) but with preserved testicular development . In fact, the phenotype has been replicated and proved feasible in mice, pigs, goats and cattle  belongs to a family of zinc-finger motif-contained RNA-binding protein and it is necessary for generating the spermatogenic cell lineage and for spermatogonial stem cell (SSC) self-renewal . The CRIelopment . The autd cattle . Adult Nindustry .NANOS2 knockout offspring associated with SSCT may impact positively the field of cattle production. Surrogate sires generated from adapted indigenous breeds (such as zebu breeds) could carry sperm from thermosensitive high genomic value sires (from European breeds) and be used to breed cows by natural mating in the tropical or tropical regions , although it has a short lifetime (2 to reduce carbon dioxide (CO2) and produce methane, which is released to the atmosphere mainly through eructation and breathing , SDN-2 and SDN-3 . The appIn the European Union, the Court of Justice decided that products developed by gene editing techniques are subject to the same regulation of GMOs regardless of the approach employed,, although the matter is still under discussion . In the For SDN-3, the DNA repair is also performed by HDR, but usually with a large donor template where an exogenous DNA sequence  is included. Organisms generated by SDN-3 approach are uniformly considered as GMOs.Thus, depending on the country, gene-edited products generated by SDN-1 or SDN-2 approaches can be classified as non-GMO on a case-by-case analysis by local regulatory agencies . That caCattle farming in tropical and subtropical regions have several challenges imposed by the environment, one of the main ones being heat stress. Heat stress results in low productivity and is one of the main constraints for efficient cattle farming activity in such regions. Gene editing technologies can be applied to decrease the negative effects of heat stress on productivity. Mutations associated with heat tolerance can be inserted in thermosensitive European breeds; similarly, mutations associated with milk yield and composition can be inserted in thermotolerant but low productivity indigenous breeds. Surrogate sires from adapted breeds can carry sperm from non-adapted, high genomic value bulls for natural mating. Increasing cattle productivity in the tropics and subtropics will contribute to produce more animal protein without significantly increasing methane emissions. Finally, gene editing could also be applied to modify the expression of genes in Archaea in order to modulate methane production in the rumen. Those potential applications are summarized in the


Case Description. A 45-year-old male presented with a significant painless decreased vision in the right eye following an electrical injury of the right hand in his workplace. The best-corrected visual acuity (BCVA) of the right eye was 20/40. Funduscopic examination of the right eye revealed diffuse superficial and deep intraretinal hemorrhages, mild venous tortuosity, and an area of the pale retina. Optical coherence tomography (OCT) demonstrated hyperreflective band-like lesions in the middle retinal layers. Patchy areas of vascular flow void in deep capillary plexus seen in OCT angiography of the right eye were compatible with PAMM. Fluorescein angiography of the right eye was indicative of delayed venous filling suggestive of CRVO. The left eye was completely normal on exam and imaging.  To report a case of central retinal vein occlusion (CRVO) and paracentral acute middle maculopathy (PAMM) following electric shock injury.  This report illustrates the occurrence of CRVO associated with PAMM following electric shock injury. Electrical injury leads to a wide range of retinal manifestations. Clinicians need to pay attention to any hyperreflectivity and thinning of middle retinal layers in OCT in cases with the presentation of sudden visual loss following electrical injuries. Electrical injuries lead to a wide range of ocular manifestations, from the eyelid, conjunctival, corneal, and lens involvement to posterior segment manifestations. Complications such as vitreous hemorrhage, retinal detachment, retinal edema, chorioretinal rupture, macular hole, Purtscher-like retinopathy and retinal vascular occlusive diseases are examples of retinal pathologies reported following electrical injuries –3.The severity of the electrical damage depends on the electric current intensity, the exposure time, and the tissue resistance to the electricity . The optSeveral mechanisms have been proposed to explain how tissue damage occurs following electrical injuries. These mechanisms include direct transmission of the current, heat effect, tissue ischemia, and mechanical injury from electrical shock waves . The eleA 45-year-old male, an electrical technician by profession, presented to us with marked painless decreased vision in the right eye (RE) since the last day following an electrical injury in his workplace. A current of 400 V passed through his right hand before the presentation. There was no history of unconsciousness or falling after injury. No significant family or past medical history was documented. His general physical examination was normal.On ocular examination, his best-corrected visual acuity (BCVA) was 20/40 for the right eye (RE) and 20/20 for the left eye (LE). The pupil examination revealed round and reactive pupils and a negative relative afferent pupillary defect.Extraocular movements were normal. Intraocular pressure measured 12 mmHg for the RE and 13 mmHg for the LE. Anterior segment examination was unremarkable. We found diffuse superficial and deep intraretinal hemorrhages and mild venous tortuosity compatible with CRVO and an area of the pale retina in the macula of the RE. The examination of the LE was entirely normal.Multimodal imaging including spectral-domain optical coherence tomography (SD-OCT) , OCT angiography , and fluorescein angiography  has been performed to make a more accurate diagnosis. SD-OCT of the RE showed no macular edema and hyperreflective band-like lesions in the middle retinal layers suggestive of paracentral acute middle maculopathy (PAMM). . OCT angOur patient's BCVA was 20/20 in both eyes after a month of follow-up, and no neovascularization of the retina or iris had developed.PAMM was first described in 2013 by Sarraf et al. as a hyperreflective band-shaped lesion in the inner nuclear layer (INL) and the outer plexiform layer (OPL), leading to thinning of the INL. PAMM first was considered as a variant of acute macular neuroretinopathy (AMN), but these are now known as two distinct entities. Unlike PAMM, AMN lesions are red wedge-shaped intraretinal lesions that point toward the fovea and are better seen with infrared reflectance images. On the other hand, AMN lesions in OCT are placed deeper than PAMM and involve the outer nuclear layers and the OPL and may lead to thinning and destruction of the outer retinal layers. Although the exact mechanism of PAMM is still under investigation, OCT-A findings have suggested vascular changes in the intermediate and deep capillary plexus due to hypoperfusion. The INL and OPL are located in the watershed area of the retinal blood supply. In a hypoperfusion state, the maximum blood flow is directed toward the superficial capillary plexus and choroid, so INL and OPL are more prone to ischemia . Also, tAnother mechanism that is responsible for INL damage in PAMM is reperfusion injury. Reperfusion occurred following a transient hypoperfusion results in increased production of reactive oxygen species, nitric oxide neurotoxicity, inflammation, and extravascular leakage. Reperfusion injury is associated with chronic changes of PAMM .Previous studies have shown the association between some retinal vascular diseases and PAMM including retinal artery and venous occlusion .Electric current can damage tissue in several ways. The direct passage of electricity through the tissues leads to the destruction of the cell membrane and direct cell damage. Also, the heat produced by the conversion of electrical energy causes vascular contraction, resulting in tissue ischemia and eventually necrosis. Also, the direct mechanical injury caused by an electric shock wave is noted. The tissue conductivity of electrical current also affects the severity of the damage, so tissues such as the retina and optic nerve are more vulnerable to electrical injury. The macular region is more prone to electrical injury due to several reasons. The avascularity of the fovea makes it more sensitive to ischemic damage. Due to the greater accumulation of melanin in the macula, the heat produced in the macular region is higher, and the risk of tissue ischemia is more , 5.It seems that in our patient, the transmission of electric current led to vascular constriction followed by tissue ischemia and presented with central retinal vein occlusion. Findings of the fundoscopic examination including diffuse superficial and deep intraretinal hemorrhages and mild venous engorgement and delayed venous filling in FA support the diagnosis. According to the abovementioned mechanisms, both hypoperfusion and reperfusion injury may disturb vascular flow toward the deep capillary plexus (DCP) and cause PAMM in this patient. Although FA has been the gold standard for the evaluation of retinal blood circulation, it is not able to show blood flow in the vascular layers of the retina separately. OCT-A is a noninvasive modality, which is helpful for this purpose. In our patient, hyperreflective, band-like lesions in the middle retinal layers of the macula in SD-OCT, and patchy areas of vascular flow void in the DCP in OCT-A are suggestive of PAMM.There is yet to be a treatment that has been proved to reverse PAMM; thus, treatment should be focused on the underlying cause of PAMM when it is present . In our As in our patient, the majority of PAMM cases have a good visual prognosis, with good final VA and no known neovascularization or rubeosis iridis in concomitant CRVO and CRAO cases .The present case illustrates a patient with a sudden unilateral marked decrease in visual acuity following an electrical injury at the workplace. Findings of ocular examination and multimodal imaging support the diagnosis of PAMM and nonischemic CRVO. Electrical injuries lead to a wide range of retinal presentations that can be nonspecific. Ophthalmologists need to be familiar with the various ocular manifestations of electrical injuries. Paying attention to a history of electrical injury in an otherwise healthy person presented with PAMM may be helpful. This is the first report of PAMM following electric shock injury. Currently, this report does not prove any definite relationship between electrical injury and PAMM, and further studies are required for preparing a causal relationship.