Vegetation played an important role in modulating multiple pressures on ESs. This study contributes to ecosystem management by effectively coping with anthropogenic and environmental pressures and sustaining the supply of ESs, particularly in alpine and plateau regions.Human activities and environmental change can impact the supply of ecosystem services (ESs) as pressures. Understanding the mechanisms of these impacts is crucial to support ecological conservation and restoration policy and applications. In this study, we highlighted the contribution of vegetation to mitigating these impacts on ESs in the Qinghai-Tibet Plateau (QTP) of China. First, we identified hot and cold spots of pressures from human activities and environmental factors and mapped the cumulative provision of five ESs . Then, we clustered these ESs into five bundles based on their supply level. Furthermore, structural equation modeling was used to quantify the pathways of multiple pressures on ESs. The results indicated that 1) for 2000, 2010 and 2019, the percentages of hot spots with high pressure were 28.88%, 27.59% and 45.66% respectively, with significant spatial heterogeneity from northwest to southeast; 2) both regions with high and low cumulative ES values experienced increased volatility; and 3) the joint effects of multiple pressures shaped ESs through pressure-ES (direct) and pressure-vegetation-ES (indirect) pathways. Specifically, precipitation had the largest positive effect on regulating services ( There i2 concentration to promote crop photosynthesis, but many regulating and cultural ESs were mostly negatively affected  and changes in environmental factors , ESs have shown many ways of changing over time . For exaaffected . A studyaffected . In addiaffected . Thus, haffected . Althougaffected . Anotheraffected . Most staffected . The impaffected . The hisaffected .While many studies have investigated various impacts of pressures on ESs , few stuThe QTP is a water tower area and an essential eco-safety shelter in the world, as well as a focus for ecological civilization construction in China. The QTP is of great significance to the sustainable development of the region and the ecological conservation of China and the world . As the Based on the above, we explored the following hypotheses: (1) the relationship between pressures and ESs changes over time, and (2) vegetation condition plays an indirect role in the impact of pressures on ESs. To this end, we first assess the spatiotemporal variation in multiple pressures. We then analyze the spatial patterns, bundle zones and temporal variations in ESs. Finally, we identify correlations between pressure, vegetation conditions and ESs, and propose corresponding ecosystem management and ecological conservation strategies.22.12 , and vegetation condition datasets ). All datasets are from 2000, 2010 and 2019, and are shown in 2.3(1) Human activitiesHuman activities have been proven to have effects on the sustainable provision of ESs . In this(2) Meteorological pressuresMeteorological pressures affect ESs through temperature and precipitation . Annual (3) Landscape fragmentationFragmentation weakens the resilience of ecosystems . Landsca2.4The CI and NDVI were chosen as the indices for vegetation condition. The CI is important in the determination of hydrological processes, photosynthesis, and canopy radiative transfer , which c2.5We specified five ESs for the QTP on a scale of 1 km. Five ES indicators  for 2000, 2010 and 2019 were chosen based on the Millennium Ecosystem Assessment , data av(1) Water yieldFreshwater provision is a crucial ES that is of great significance to human society . Moreove(2) Soil retentionThe sediment delivery ratio (SDR) module in InVEST was employed to simulate soil retention. The parameters needed for the run of this model include land use/cover data, erodibility data, terrain data, biophysical table, watershed division data, and threshold flow accumulation . The mod(3) Carbon sequestration2  Habitat qualityAn ecosystem’s habitat quality refers to its capability to provide wildlife or specific populations with the necessary resources and conditions to survive . The hab(5) Landscape aestheticsThe scenic views of landscapes contribute to local communities’ well-being in a variety of ways. To increase local economies, landscape aesthetics play an essential role in attracting visitors. In the study, the scenic quality module (SQ) of InVEST was used to assess landscape aesthetics. The SQ calculates the value of the impacted visibility. The parameters needed to run the model are described in detail in the reference .2.62.6.1*) hot spot analysis in ArcGIS was used to identify clusters where z-scores were high (hot spots) and low (cold spots). With hot spot analysis, local patterns of spatial associations can be easily identified, making it more suited for comparing and visualizing pressure patterns . By using SEM, we can implement conceptual models and estimate the relationships between their variables, where some latent variables are measured by manifest variables linked by linear regressions . A model33.1The spatiotemporal variation in pressures was identified by hot spot analysis. The three heterogeneous units shown in 3.23.2.1For the five ESs evaluated in the study, the cumulative value of ESs registered a maximum score of 2.9 Figure 43.2.2The grids of the QTP were divided into 5 bundles, where grids with similar ESs were bundled together. According to the supply of ESs, five bundles were named: “Habitat and carbon sequestration zone (HCSZ)”, “Landscape aesthetics zone (LAZ)”, “Habitat zone (HZ)”, “Carbon sequestration zone (CSZ)”, and “Insufficient ES zone (IESZ)”. The ES bundles are visualized in HCSZ was characterized by high NPP and habitat quality with a low delivery of soil retention, landscape aesthetics and water yield. In LAZ, landscape aesthetics were high, but NPP, water yield, habitat quality and soil retention were low. In HZ, habitat quality was high, but NPP, water yield, landscape aesthetics and soil retention were low. In CSZ, NPP was high, but water yield, landscape aesthetics, habitat quality and soil retention were low. In IESZ, all five ESs were low.3.32R ≥ 0.76), and only 1% of the variance in cultural service (2R = 0.01) in all three years. Human activities, temperature, precipitation, and landscape fragmentation together explained 42%, 13%, and 7% of the variability in vegetation conditions in 2000, 2010 and 2019, respectively. All the components analyzed in the SEM are strongly related, except for the relationship between human activities and vegetation conditions in 2010 . Landscape fragmentation had the largest effect on cultural service (-0.10 ≤ rα ≤ -0.07). Vegetation conditions had weak negative effects on cultural service . Additionally, vegetation conditions negatively affected regulating services in 2000 (rα = -0.26), while there was a positive relationship between vegetation conditions and regulating services in both 2010 and 2019 .The SEM results explained greater than three-fourths of the variance of regulating services R2 ≥ 0.76, In 2000, temperature, precipitation and landscape fragmentation had positive and significant effects on vegetation conditions and regulating services  The first pathway in which pressures influenced ESs was the direct pathway, namely the pressure-ES pathway (PESP) Figure 7(2) The second pathway considered vegetation conditions, namely, the pressure-vegetation-ES pathway (PVESP). The indirect pathway analysis indicated that multiple pressures could change the supply of ESs by influencing vegetation conditions Figure 74.3In this study, we found that both ESs and multiple pressures had spatiotemporal heterogeneity 4. This Considering the needs of ecological conservation, determining how to assess the relationship among pressures, vegetation, and ESs has become a crucial issue globally . The comThis study focuses on pressures and ESs and follows “spatiotemporal variation analysis, impact pathway identification, and ecosystem management support”, which has a good reference for the use of ES research to guide regional management in other alpine plateau regions. In addition, most of the data in this study are available through remote sensing and field sampling, making it easier to focus on spatial patterns and temporal changes in ESs and pressures, and thus better support ecosystem management decisions. The study primarily indicates rankings of ESs and pressures, rather than exact values, which can be easily performed in study of multiscale pressure-ES relationships and ecosystem management.Our assessment of ESs was mainly based on InVEST model, which was not compared with field site data. The unverified calibration of the simulation may lead to some uncertainty in the results of the models. However, it is difficult to verify the model with field data in such a large area as the QTP under current conditions. We will strengthen this in future studies to make the results more accurate and support regional ecological management.5Ecosystem management is challenged by multiple pressures. Synthesizing and learning pathways of “pressure-ES” and “pressure-vegetation-ES” is necessary to deepen the understanding of the impact mechanism of pressures on ESs and how they are used for landscape optimization and ecosystem conservation. Our study indicated that both spatial agglomeration and the amount of pressures on ESs increased in the QTP from 2000 to 2019. Although regions with cumulative ESs above the intermediate level accounted for 70% of the QTP with spatial heterogeneity, regions with poor ES supply were growing rapidly. Moreover, the results highlight the evidence that both PESP (direct) and PVESP (indirect) were vital pathways, providing a different perspective to dissect how pressures affect ESs. Our findings emphasize the need to consider the complex interactions between pressures, vegetation, and ESs in ecosystem conservation and management practices. Improvements in spatially targeted policy design are expected in the future by identifying ES bundles and the relative importance of multiple pressures affecting vegetation and ESs.The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding author.YXL: writing, reviewing, editing, and validation. YHL: conceptualization, editing, and resources. MZ: data curation, writing, and methodology. BF: conceptualization and reviewing. All authors contributed to the article and approved the submitted version.


This study systematically evaluates the effect of mindfulness meditation on depression during pregnancy. We provide evidence-based suggestions for preventing and reducing depression during pregnancy by exploring the most effective intervention mode, cycle, and frequency of mindfulness meditation.Records were retrieved from PubMed, Web of Science, EBSCO, and Science Direct. A total of 1,612 randomized controlled trial studies on the effect of mindfulness meditation on pregnancy depression were collected. 10 studies with 658 subjects meeting the inclusion criteria were extracted and analyzed by Revman 5.3 to evaluate study qualities. Stata 12.0 was used for the meta-analysis.P < 0.001]. Subgroup analysis showed that mindfulness-based cognitive therapy (MBCT) had the best intervention effect on depression during pregnancy (SMD = 2.795), the best intervention duration was below 4 weeks (SMD = 1.756), applied from the first to the third stage of pregnancy (SMD = 1.024), the frequency guided by experts was less than six times a week (SMD = 2.055) of <60 min each time (SMD = 1.068), and completing homework by themselves every day for 30 mins (SMD = 1.822) was most significant for the improvement of depression during pregnancy. We found high heterogeneity across studies . This may be caused by variance in measurement tools, among which Beck Depression Inventory-II was a significant source of heterogeneity.Mindfulness meditation had a positive effect on depression during pregnancy [standardized difference of the mean (SMD) = −0.786, 95% confidence interval , Mindfulness meditation can improve the prevention, remission, and reduction of depression during pregnancy and can be used as an auxiliary measure for the clinical treatment of pregnancy depression. According to a report by the World Health Organization, depression has become one of the major causes of disability worldwide  guidelines  basic information: first author, literature title, country, publication year; (II) baseline characteristics of subjects: age, population, sample size, pregnancy cycle; (III) specific intervention mode, cycle, frequency, and time; (IV) measurement tools and outcomes.Two reviewers independently evaluated the quality of the literature using Review Manager 5.4.1. Quality was evaluated as “low risk,” “unclear,” or “high risk” depending on the following seven aspects: random sequence generation, allocation concealment, blinding of participants and personnel, blinding of outcome assessment, incomplete outcome data, selective reporting, other bias. In case of differences, a third researcher assisted in the judgement.2, and the heterogeneity was divided into negligible heterogeneity (I2 = 0–24%), medium heterogeneity (I2 = 25–50%), large heterogeneity (I2 = 50–74%), and non-negligible heterogeneity (I2 = 75–100%)  as an index of effect sizes for measurement data. The effect sizes were categorized as small (0.2 ≤ SMD <0.5), medium (0.5 ≤ SMD <0.8), and large (SMD ≥ 0.8). Heterogeneity tests were performed by calculating χA total of 1,612 articles were obtained through database searches and reference collection from relevant papers. After deleting duplicate articles, 1,591 of these were screened. Through reading their titles, abstracts and full text, articles with incomplete, repeated, or inappropriate non-RCT studies were excluded. Finally, ten qualified studies were included. The literature screening process is illustrated in The basic characteristics of the included studies are presented in P < 0.001]. Through the heterogeneity test , non-negligible heterogeneity was detected; thus, a random effects model was selected for the meta-analysis to explore the source of heterogeneity, as shown in The results of the meta-analysis using a random effects model showed that mindfulness meditation had a significant positive effect on depression and significantly reduced depression symptoms during pregnancy [SMD = −0.786, 95% CI , P > 0.05).We performed sensitivity analysis of the included studies, as shown in P-value above 0.05, indicating that these variables had no significant impact on the heterogeneity among the included studies, while the results show that measurement tool had P-value below 0.05.To further explore the source of heterogeneity, we conducted univariate meta-regression analysis based on the aspects of country, measurement tool, expert-guided intervention frequency, intervention time, homework time, intervention cycle, the participants' pregnancy cycle, and intervention method. The results of the meta-analysis in Of the six depression measurement scales used across the studied papers, only the BDI-II showed a significant effect. In a systematic review of the prevalence of depression during pregnancy, the Beck Depression Scale, EPDS, and structured interviews were used to evaluate depression status to evaluate differences between the scales. The results showed that the Beck Depression Scale had more physical items than the Edinburgh Postpartum Depression Scale  A reduction in the level of cortisol, the main hormone of stress, reduces adverse psychological symptoms and negative emotional reactions, increases subjective wellbeing, improves behavioral regulation, and changes brain activity accordingly, indicating that mindfulness meditation has a direct positive effect on the brain . The number of expert-guided interventions could be less than six times a week, each time ≤60 min. Moreover, daily self-meditation of 30 min according to the guidance of experts is also recommended. Future research should supplement this information and provide more targeted suggestions for pregnant women with different statuses.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.YL, JC, and BC formed the conception of the study and drafted the manuscript. TW, ZW, and XH collected and analyzed the data. SL contributed to the conception and revision of the study. All authors contributed to the article and approved the submitted version.This paper is supported by the National Social Science Fund Projects The research on intelligent elderly care service mode of sports and medicine integration (No. 21XTY006) and Beijing Social Science Fund Major Projects Theoretical and practical research on the deep integration of national fitness and national health in the new era (No. 20ZDA19).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Portal hypertension monitoring is important throughout the natural course of cirrhosis. Hepatic venous pressure gradient (HVPG), regarded as the golden standard, is limited by invasiveness and technical difficulties. Portal hypertension is increasingly being assessed non-invasively, and hematological indices, imaging data, and statistical or computational models are studied to surrogate HVPG. This paper discusses the existing non-invasive methods based on measurement principles and reviews the methodological developments in the last 20 years.First, we used VOSviewer to learn the architecture of this field. The publications about the non-invasive assessment of portal hypertension were retrieved from the Web of Science Core Collection (WoSCC). VOSviewer 1.6.17.0 was used to analyze and visualize these publications, including the annual trend, the study hotspots, the significant articles, authors, journals, and organizations in this field. Next, according to the cluster analysis result of the keywords, we further retrieved and classified the related studies to discuss.n = 43), while “Journal of Hepatology” had the highest citations. “Liver fibrosis” published in 2005 was the most influential manuscript. Among the 20,558 cited references of 1,088 retrieved manuscripts, the most cited was a study on liver stiffness measurement from 2007. The highest-yielding country was the United States, followed by China and Italy. “Berzigotti, Annalisa” was the most prolific author and had the most cooperation partners. Four study directions emerged from the keyword clustering: (1) the evaluation based on fibrosis; (2) the evaluation based on hemodynamic factors; (3) the evaluation through elastography; and (4) the evaluation of variceal bleeding.A total of 1,088 articles or review articles about our topic were retrieved from WoSCC. From 2000 to 2022, the number of publications is generally growing. “World Journal of Gastroenterology” published the most articles , CT, and magnetic resonance imaging (MRI) support the non-invasive assessment of PH by providing both hemodynamic and fibrotic information. Elastography, mainly USE, is the most important method of PH monitoring. Portal hypertension (PH), defined as portal pressure gradient (PPG) or hepatic venous pressure gradient (HVPG) > 5 mmHg, nearly 90% is cirrhotic PH, which belongs to intrahepatic PH . CirrhotHVPG, the golden criterion to assess the presence and severity of portal hypertension, is the difference between wedge-shaped hepatic venous pressure (WHVP) and free hepatic venous pressure (FHVP) . GradingThe high technical difficulty, high cost, invasiveness, and low feasibility of multiple measurements restrict the widespread clinic application of HVPG . OtherwiThis article reviews the development of non-invasive detection of portal pressure with the assistance of bibliometric-analysis visualization and analyzes the current shortcomings and future expectations, in hopes of the routine monitoring of PH being a reality in the clinic.On 8 April 2022, we retrieved a total of 1,209 records (publication time from 2000 to 2022) on the Web of Science Core Collection (WoSCC) by searching the term Topic  AND Topic (“non-invasive” or “non-invasive” or “non-invasive”). A total of 121 meeting abstracts, editorial materials, letters, corrections, and reprints were excluded. Approximately, 1,088 articles or review articles were included in this study. The full record and cited references of the 1,088 articles were exported in the format of a plain text file. Next, the bibliographic data were imported to VOSviewer 1.6.17.0 to create analytic maps. VOSviewer was a feature-rich bibliometric analysis tool, and the abundant bibliometric information in WOS data made it play completely. The analytic results were saved in the format of a figure and were exported to Microsoft Office Excel 2021. The analysis of the trend of annual publications and the rank of authors, regions, organizations, journals, literature, and keywords were finished in Excel. Based on keyword clustering, the four clusters inspired further discussion of non-invasive methods for PH detection.There were in total 306 journals of 1,088 searched pieces of literature, and the top 10 high-yielding journals are listed in There were in total retrieved 1,088 documents, and the 10 documents in Totally, of the 68 countries/regions of the 1,088 documents, 34 countries/regions met the requirement of ≥ 5 publications. Of the 34 countries/regions, 32 countries/regions had connections with others and constructed the network as n = 26). The top 10 high-production organizations are shown in Totally, of the 1,400 organizations, 100 organizations met the restriction condition of publishing over 5 pieces of literature, of which 91 organizations had connections and consisted of the co-authorship network . The 91 n = 42), and had a connection with other 7 clusters (except for yellow Cluster 4 and pink Cluster 10). The top 10 high-yield authors are listed in In total, there were 5,779 authors of 1,088 pieces of literature; 130 authors met the threshold of over 5 publications, of which, 100 authors have a connection and composed the co-authorship network as n = 254) and was often co-cited with “Transient elastography accurately predicts the presence of significant portal hypertension in patients with chronic liver disease” (rank 8th) published in 2008, “Liver stiffness measurement selects patients with cirrhosis at risk of bearing large esophageal varices” (rank 7th) published in 2006, and “Transient elastography for diagnosis of advanced fibrosis and portal hypertension in patients with hepatitis C recurrence after liver transplantation” (rank 14th) published in 2006. Moreover, “Expanding consensus in portal hypertension: Report of the Baveno VI Consensus Workshop: Stratifying risk and individualizing care for portal hypertension” (rank 2nd) was more recent.In total, there were 20,558 references of 1,088 retrieved pieces of literature, from 2,908 journals, having 13,108 first authors. Of the 20,558 references, there were 309 references cited over 20 times by the 1,088 retrieved literature. The top 10 high-cited references are listed in There was a total of 2,806 keywords retrieved from 1,088 publications. Among them, 102 keywords occurred over 20 times and formed the network as Blue Cluster 3 consisted of 26 keywords shown in Portal hypertension-related fibrosis included liver fibrosis and spleen fibrosis, the former was the leading component of PH, and the latter reflected the extrahepatic secondary alterations caused by PH. This part mainly depicted the PH evaluation based on liver fibrosis. Liver fibrosis was characterized by the chronic inflammatory insult of liver parenchyma and excessive collagen deposition and was also accompanied by the occurrence of endothelial dysfunction . CirrhotThe injury of liver parenchyma was the feature of liver fibrosis, related liver function indices including single platelet count (PLT), aspartate aminotransferase to alanine aminotransferase (AAR) ratio, aspartate aminotransferase to platelet count ratio index (APRI), fibrosis index based on 4 factors , glutamyl transpeptidase to platelet (GPR), King’s score , Lok score , and the CSPH risk score  have been validated to evaluate PH. Using HVPG ≥ 10 mmHg as the golden standard, the area under the receiver operating characteristic curve (AUROC) of the above indices to diagnose CSPH was 0.6–0.8 and hardly exceeds 0.8 –29.In addition to the above conventional biomarkers, the more proprietary markers like increased osteopontin (OPN) accompanying liver fibrosis have also been discussed . Plasma A recent study  of NASH R = 0.77, p < 0.0001) in 2007, but missed further study  and was independent of the Child-Pugh score prediction. When the taken cut-off value of vWF-AG was ≥ 241%, the positive predictive value of detecting CSPH was 87%, and the negative predictive value was 80% . Another study   . On this≤ 0.001)  measured≤ 0.001)  predicteComamonas, Cnuella, Dialister, Escherichia/Shigella, and Prevotella, but poor Bradyrhizobium, Curvibacter, Diaphorobacter, Pseudarcicella, and Pseudomonas. There was no significant difference in peripheral and hepatic venous blood in patients with cirrhosis. Concentrations of Bacteroides, Escherichia/Shigella, and Prevotella were found in patients with severe portal hypertension (SPH). Another study  and correlated with HVPG . When the LSN scores cutoff value of 2.8 was used to detect CSPH, the positive predictive value was 88%. The AUROC of LSN scores was 0.88 ± 0.03, which was higher than most blood tests. LSN scores could also be obtained from MRI, and the result was similar  could also assess the level of fibrosis through related morphological features, and then evaluate PH. Liver surface nodularity (LSN) has been proven to differentiate cirrhotic from non-cirrhotic livers with AUROC = 0.929 . Based o similar . But thep < 0.001). ECV values showed the same trend. The results showed a high correlation between ECV and portal hypertension . Subsequently, clinical data by Mesropyan et al.  and directly with HVPG , and had a good diagnostic effect for CSPH (AUC = 1.000). In the validation cohort, the diagnostic performance of mapping parameters was comparable. Splenic ECV has a good predictive value for portal hypertension in patients with advanced liver disease.In 2014, MR T1 mapping of the liver was firstly reported to provide high diagnostic accuracy in the non-invasive assessment of liver fibrosis . Extracen et al.  showed tArtificial intelligence and machine learning, which seem to have nothing to do with medicine, have been gradually changing clinical diagnosis and treatment by fully utilizing the information from routine imaging, serological, and other examinations. Bosch et al.  conducteIn summary, the ML algorithm could identify fibrous matrix features and correlate with HVPG measurements. In addition to morphology-based features, texture features and deep convolutional neural network-based imaging analysis also showed a potential performance in assessing PH , 57. HowRed Cluster 1 includes 30 keywords shown in The PH assessment methods based on fibrosis and hemodynamics were not completely separate, as hemodynamic indices like hepatic perfusion could also be used to evaluate the level of liver fibrosis. PH-related hemodynamic factors include intrahepatic and extrahepatic blood flow, the former mainly included portal vein, hepatic vein, and hepatic artery; while the latter included splenic vein, renal artery, and collateral circulations. US, CT, and MRI could provide static indices as vessel diameter and dynamic indices as flow velocity to reflect portal pressure. Moreover, ascites, spleen sclerosis, and splenomegaly could also be regarded as extrahepatic hemodynamic factors.Although imaging was the most common method of estimating blood flow, blood tests like the filtration test of indocyanine green (ICG) and cholate could also reflect the status of hepatic blood flow, which worked as the contrast in imaging. The ICG retention test was first used to evaluate the residual liver function before hepatectomy and was determined by the liver secretory function and liver blood flow . In 2014Cholate, similar to ICG, was high hepatic extraction, and its extraction was determined by hepatic blood flow instead of hepatic metabolism . The morAlthough the principle was similar to ICG, there was no comparison between the HepQuant SHUNT test and ICG-R15 in predicting portal pressure. Additionally, the relatively complicated and costed blood tests, such as ICG-r15 and HepQuant SHUNT tests, required larger confirmatory studies to check the cut-offs and the feasibility in different clinical scenarios.The introduction of Doppler ultrasound (DUS) was an important breakthrough in evaluating organ hemodynamics . DUS wasContrast-enhanced ultrasound (CEUS) with microbubbles improved the imaging quality of the US with a higher signal-to-noise ratio. When the ultrasound pulse reached the microbubbles, the microbubbles resonated and scattered the fundamental wave, subharmonics, second harmonics, and superharmonics, and the harmonics created contrast imaging with tissue echoes. Among these harmonics, the second harmonic was the strongest signal . The hemr = 0.82), and the correlation was greater (r = 0.97) when HVPG ≥ 12 mmHg (n = 6). In 2021, a larger-scale clinical study in 125 patients with chronic hepatitis of various etiologies was carried out by Gupta et al. (r = 0.68), and the correlation was lower (r = 0.41) when the HVPG ≥ 12 mmHg (n = 14), which was contradictory and possibly because of the low sample size or the changed transmit pulses. Nonetheless, SHAPE ≥ −0.11 dB could discriminate HVPG ≥ 10 mmHg with sensitivity of 91% and specificity of 82% and discriminate HVPG ≥ 12 mmHg with sensitivity of 90% and specificity of 80%.The received intensity of the ultrasound signal was determined by transmitting pulses and microbubble scatter areas. Due to the compressibility of a microbubble, ambient pressure could change its size and scatter area. Hence, the change of harmonic amplitude could reflect ambient pressure, and the subharmonic was the most sensitive harmonic . Based oa et al. . Comparein vivo had not been fully studied, and the interpretation of the contrasting results also required further study , and in the validation set with 10 patients still kept a high correlation. More recently, Hectors et al.  or three-dimensional (3D) MRI have also proved the correlation with HVPG –89, and in vivo measurement of 3D blood flow dynamics in the heart and the large vessels  with strong vascular network connectivity, the clustering coefficient was 0.4447; while, in the set with HVPG ≥ 10 mmHg (n = 11), the clustering coefficient was 0.0237. The correlation coefficient between the predicted HVPG of this model and measured HVPG was greater than 0.8. Xiao Long Qi and his team proposed an interesting idea (n = 29), the AUROC of v-HVPG in predicting CSPH was 0.83. In the validation cohort (n = 73), diagnostic performance was prospectively confirmed with an AUROC of 0.89. The inter- and intra-observer agreement was 0.88 and 0.96, respectively, indicating good reproducibility of virtual HVPG measurements.In 2015, Amat-Roldan et al.  construcing idea , that waYellow Cluster 4 consists of 20 keywords shown in 2 could be diagnosed as CSPH  of only 62.8%, which might derive from the technical accuracy influenced by the fat thickness and probe size (M or XL), or from the physical increased stiffness of fat accumulation, which lowers the correlation between liver stiffness and cirrhosis  was the best recognized and the most widely used method to surrogate HVPG. The past study  had supp as CSPH . Under tirrhosis . Notablyirrhosis , 101, soirrhosis , 40. Howirrhosis .Although LSM has gained widespread acknowledgment in assessing PH, the past study proved that the relationship between LSM and PH was blunted with increasing pressure when HVPG ≥ 10–12 mmHg, which might be from the stronger involvement of extrahepatic factors like collateral circulation . MoreoveSpleen stiffness measurement (SSM) holds the potential to address the above issues with the ability to reflect extrahepatic hemodynamic changes. In 2012, a study  in 100 pIn summary, USE was the most significant hot spot in predicting PH, which is comprehensively proper in terms of technical difficulty and risk, consumption of time and money, feasibility, and accuracy. The studies about USE in assessing PH could be mainly grouped into three directions:1.The technical questions, including operation protocols , the influence of subcutaneous fat thickness, spleen size, the alanine aminotransferase level, and other diseases.2.The comparison of LSM and SSM3.The combination of USE with other markersFor the first direction, there had been specialized recommendations , 112 to In addition to ultrasound elastography, MRE could also measure tissue stiffness in clinical MRI examinations by encoding shear wave propagation into magnetic resonance phase signals. MRE showed an excellent ability to detect liver fibrosis and cirrhosis with a low failure rate . The EAS•n = 155)Liver biopsy with no evidence of portal hypertension Patients with portal hypertension who were tested for HVPG Healthy subjects  ; other iThis review was based on bibliometric analysis and reviewed the non-invasive methods of PH monitoring according to the keywords clustering results of VOSviewer. The limitations included that the search strategy could not cover all the literature about non-invasive tools on PH. Besides, inequality in the quality of the retrieved literature might influence the credibility of the analysis results. However, the retrieved literature was enough to show the basic architecture of this field and supported further system retrieval.The portal vein was away from the surface of the body, making direct puncture a high-risk operation. WHVP was first proposed as an indirect method to measure portal pressure in the 1950s , from thAs listed in As far as detecting changes in portal pressure after various therapies, LSM was unreliable in evaluating the effect of NSBB, and NSBB could not significantly change the stiffness of the liver . While hMN and XL contributed to the conception, design, and final approval of the manuscript. JX, SW, AA, LW, and TR contributed to sequencing data. XS and HN wrote the final manuscript. All authors have read and approved the final version of the manuscript.


Due to diagnosis of gastric cancer in advanced stages as well as its poor prognosis, finding biomarkers is essential. In this study, using the TCGA RNAseq data of gastric cancer patients, we evaluated the diagnostic value of lncRNAs that had differential expression.P-value, FDR, and log fold change for whole transcripts. Next, by comparison of the RNAseq gene names with the total known lncRNA names, we identified differential expressed lncRNAs. Following this, specificity and sensitivity for lncRNAs coming from the previous step were calculated. For more confirmation, we predicted target genes and performed GO and KEGG signaling pathway analysis. In the end, we examined reliability and consistency of expression of this signature in three gastric cancer cell lines and one of them in twenty tumors and tumor-adjacent normal tissue samples using qRT-PCR.We evaluated in vitro.Five lncRNAs had proper sensitivity and specificity and had target genes involved in cancer-related signaling pathways; however, they showed different expression patterns in TCGA data and PART1, UCA1, DIRC3, HOTAIR, and HOXA11AS require more investigation to be confirmed as diagnostic biomarkers in gastric cancer.The results of our study demonstrated that the five-lncRNAs  Gastric cancer (GC) is one of the most important leading causes of cancer-related death worldwide . AccordiLong noncoding RNAs (lncRNAs) are attributed as a class of non-coding RNAs (ncRNAs) of greater than 200 nucleotides in length, which have a specific expression in various tissues and diseases such as cancers –9. LncRNPART1, UCA1, DIRC3, HOTAIR, and HOXA11AS) as a novel potential biomarker for diagnosis of GC patients. We also examined the expression of this signature in three GC cell lines and one of them in twenty tumors and tumor-adjacent normal tissue samples. In this study, we attempted to perform a comprehensive study using transcriptomic data analysis of patients with GC that are freely available in the Cancer Genome Atlas database (TCGA) (HTTP:// cancergenome.nih.gov) and to provide a new and effective signature for the diagnosis of GC patients. By conducting data mining in RNAseq data of GC patients and thereafter determination of differentially expressed genes and calculating the AUC of the ROC curve, we identified a five-lncRNA signature  contained 450 transcriptomes, of which 415 were tumors and 35 were tumor-adjacent normal tissue samples.P-value, and FDR were calculated for each transcript. In the next step, by applying appropriate filtration  on these components, the list of genes with differential expression was obtained. This list included all types of transcripts with differential expression, from which lncRNAs were to be identified. For this purpose, a list of all identified lncRNAs was obtained from the HUGO Gene Nomenclature Committee (HGNC) website and aligned with the list of differential expression genes to identify the lncRNAs with differential expression. The result was 14 lncRNAs, which were nominated for the next step of the analysis. The RNAseq data processing was continued using the R program. At first, we performed secondary data trimming by determination of the third data quantile and omission of the transcripts that had read counts of fewer than 25 in three-quarters of the data. Applying this command resulted in the removal of 4639 transcripts from the original data set with 20531 transcripts. Then, by calling the edgeR library, logFC, log CPM, ROC curveThis step was done using SPSS software to determine the sensitivity and specificity of the lncRNAs from the previous step, which ultimately led to selection of 5 lncRNAs as potential diagnostic biomarkers. Target gene prediction and functional enrichment analysisP-value less than 0.01 was considered statistically significant.The target genes of the five-lncRNA signature were predicted by the usage of two online tools including LncRRIsearch and lncRNA2Target. The sum of the identified target genes for these 5 lncRNAs was 238 by LncRRIsearch, after the removal of duplicates, and 115 by LncRNA2Target. Furthermore, the functional analysis of the GO annotation, DO and KEGG signaling pathways were performed. A Protein-protein interaction networkTo predict the relationships between the lncRNAs target gene products in-network, the ID number of the lncRNAs target genes and STRING online tool were used. The highest confidence score (a combined score >0.900) was considered significant. Globe-shaped proteins have been characterized based on their association with other proteins. The target genes with multiple connections to other target genes appear to play important roles in the protein-protein interaction network. Experimental designAppropriate experimental design is a necessity for gene expression studies. Since RNA is a sensitive molecule and prone to damage, strict conditions must be applied in dealing with it. The most important difference between the experimental and control groups was in the presence or absence of gastric malignancy. In this study, 3 GC cell lines as experimental group 1 with 20 healthy tissue samples as control group 1, and also 20 tumor tissue samples as experimental group 2 with 20 tumor-adjacent normal tissue samples as control group 2 were compared for definite gene expression. All information about experimental procedures, control groups, replicates, experimental conditions, and methods of working with samples in each group were carefully determined and recorded before starting gene expression studies. The qPCR assay was performed at the Immunology Research Center, Avicenna Research Institute, Mashhad, Iran.SamplesNormal tissue samples as a control group were obtained from 20 non-cancerous individuals that had been scanned for gastroesophageal diseases via upper endoscopy in Imam Reza hospital, Mashhad, Iran. Healthy control tissue samples were confirmed by pathological examination. The fresh specimens were immediately transferred to RNA later solution and then stored at -70 °C before RNA extraction. RNA extraction was normally performed from 40–50 milligrams of tissues until two days after tissue sampling. Written informed consent was obtained from all participants in this study before tissue sampling.5 cells from AGS, MKN-45, and EPG cell lines were grown in DMEM-high glucose , DMEM-low glucose  and RPMI 1640  media containing 10% fetal bovine serum , and Penicillin-Streptomycin , respectively. For all used cell lines, STR profiling was performed for determination of cell line identity; the report was compared with standard cell lines by the usage of matching criteria based on an algorithm that compares the number of shared alleles between two cell lines, expressed as a percentage. Cell lines with ≥80% match are considered to be related, derived from common ancestry. Cell lines with between 55% and 80% match require further analysis for authentication of relatedness; all three cell lines had an acceptable resemblance. Finally, we extracted RNA from 5×105 cells from each cell line.Three GC cell lines, MKN-45, AGS, and EPG were obtained from the National Cell Bank of Iran, Tehran. 5×10RNA extractionTotal RNA was extracted with RNX-Plus solution , chloroform , 2-propanol , and 75% ethanol  according to the common protocol used in molecular laboratories. Next, the concentration and purity of the extracted RNAs were evaluated by Spectrophotometer, Biwave II  which had a concentration and 260/280 ratio of 170–415 ng/µl and 2-2.1, respectively. We also used 2% Agarose gel electrophoresis to ensure RNA integrity by seeing the 18s and 28s bands related to rRNA. Extracted RNAs dissolved in DEPC water were stored at -70 °C until used for cDNA synthesis. For eliminating probable DNA pollution and just before starting cDNA synthesis, we treated the extracted RNAs with DNase1  according to the manufacturer’s protocol .Reverse transcriptase Reverse transcription reactions were performed in two steps using the NG dART RT kit  according to the manufacturer’s protocol. The reactions were done in a total volume of 20 µl, detailed in qPCR target informationPART1, NR_028509.1), urothelial cancer-associated 1 , disrupted in renal carcinoma 3 , HOX transcript antisense RNA  and HOXA11 antisense RNA . PART1, UCA1, DIRC3, HOTAIR, and HOXA11AS have 5914, 2314, 3910, 2364, and 1628 base pairs (bp) of lengths, respectively. Except for PART1 and HOTAIR, which have three splice variants, the rest of the non-coding RNAs have only one variant. The product sizes of qPCR for the lncRNAs were 181, 151, 73, 175, and 89 bp for PART1, UCA1, DIRC3, HOTAIR, and HOXA11AS, respectively. The designed primers were assessed using NCBI Primer-BLAST and showed 100% specificity and acceptable secondary structure scores. These primers were designed to amplify part of exon 1 of PART1, exon 3 of UCA1, exon 1 of DIRC3, exon 7 of HOTAIR, and exon 1 and 2 of HOXA11AS. The proposed diagnostic signature of this study included five lncRNAs prostate androgen-regulated transcript 1 (qPCR PrimersThe reference sequence of the lncRNAs PART1 (NR_028509.1), UCA1 (NR_015379.3), DIRC3 (NR_026597.2), HOTAIR (NR_003716.3), and HOXA11AS (NR_002795.2) was achieved from the NCBI gene database. The primer design was performed using the primer3 online tool, and specificity and secondary structure formation of the primers were checked by NCBI Primer-BLAST. All primers used in this study were synthesized by Pishgam Biotechnology Company and purified by the HPLC method. The sequences of primers used along with their PCR product length are shown in qPCR protocolGAPDH. qPCR analysis of the five lncRNAs was performed on a Light Cycler 96  using the RealQ Plus 2X Master Mix Green , which is composed of TEMPase Hot Start DNA Polymerase, dNTPs, fluorescent dye and an optimized buffer system. When the fluorescent green dye is free in the solution, it emits a very low fluorescent signal. As soon as the dye binds to the double-stranded DNA, the signal increases significantly (thousandfold), which makes the fluorescent signal of the dye directly proportional to the amount of amplified dsDNA. In this study, 40 temperature cycles were used for amplification of the targets. The amount of materials used for a 0.2 ml qPCR tube stripe in each reaction is shown in qPCR validationWe optimized the qPCR reactions by the use of the Labcycler Gradient machine . Negative control was used to ensure the absence of contamination and the accuracy of qPCR reactions. Also, to determine the specificity of the amplified products, a melting curve was drawn for all reactions after the completion of the reaction cycles. Each product or amplicon, depending on its specific Tm, will create a unique peak in the melting curve. The primer dimer also creates a peak in the melting curve, which due to its small size and low Tm is below 80 °C, and thus in most cases will be separable from the desired PCR product. In cases where the melting curve was not a single peak or was different from the expected values, the qPCR product was also run on 2% agarose gel. If non-specific bands are seen on the agarose gel, these bands can be partially removed by changing the PCR reaction parameters. For example, by reducing the amount of reaction primers, the band observed in the primer dimer range can be removed, or non-specific bands that have a long length can be removed by reducing the extension phase time or increasing the temperature of this phase.Data analysisP-value<0.05 was considered statistically significant. ROC curve analysis was performed using IBM SPSS Statistics v. 25 to determine the sensitivity and specificity of potential biomarkers. An AUC greater than 0.7 was considered a cut-off. Changes in gene expression in tumor tissues or cell lines compared with normal tissues were evaluated using ∆∆CT calculation and Log Fold Change determination. For statistical analysis of real-time results, the ANOVA test was used for cell lines and paired t-test was used for tissue samples, which was done using SPSS software. A Differentially expressed GenesThe data demonstrated that 3722 genes (1840 up-regulated and 1882 down-regulated) were differentially expressed in STAD. Moreover, 14 lncRNAs (139 up-regulated and 49 down-regulated) were identified that were most deferentially expressed in patients. The data are presented in ROC curve indicates good performance for the suggested diagnostic signatureP-value less than 0.05 was considered a significant level. According to AUC and P-value, eventually, five lncRNAs  were selected as the candidate diagnostic signature. These selected lncRNAs had good potential sensitivity and specificity for diagnosis of GC patients  and LncRNA2Target  online tGO annotation, disease ontology, and KEGG signaling pathwaysFor more investigation of the potential biological function and the mechanism of the suggested diagnostic signature, we took advantage of GO annotation, disease ontology, and KEGG signaling pathway analysis for 353 target genes from and disease ontology (DO) indicate activation of cancer-related pathways such as GC, prostate cancer, small cell lung cancer, pancreatic cancer, breast cancer, bladder cancer, colorectal cancer, melanoma, glioma, connective tissue cancer, bone cancer, renal carcinoma, etc. .Functional predictions and PPI network constructionCDKs, TWIST1, CDH1, MMPs, TP53, RB1, WNT7A, GSK3B, ATM, PTEN, BCR, ERBB2, VEGFR, SMAD, TGFB2, MAPK1, FGF1, AKT1, and HIF1A.To predict and visualize the protein-protein interactions among the target genes of the five-lncRNA signature, we used the STRING  softwareFunctional analysis via co-expressed target genesCDKs, TWIST1, CDH1, MMPs, TP53, RB1, WNT7A, GSK3B, ATM, PTEN, BCR, ERBB2, VEGFR, SMAD, TGFB2, MAPK1, FGF1, AKT1, and HIF1A that were involved in cancer-related pathways and therefore, dysregulated in many cancers. At the beginning of this study and before selecting the appropriate lncRNAs as biomarkers, a list of genes with differential expression of TCGA data was obtained and then a list of target genes was predicted by online tools for this signature. To probe the functional value of the signature in GC (stomach adenocarcinoma), we searched the predicted target genes that were co-expressed with our suggested signature and we found a list of 212 genes with this characteristic. In this new list, there are many master genes such as Real-time PCR verificationPART1 and DIRC3 had no detectable expression in any of the three cell lines and normal tissue samples. On the other hand, UCA1 was expressed in whole samples. Two lncRNAs, HOTAIR and HOXA11AS, were similarly expressed only in the EPG cell line and normal tissue samples. We investigated the expression level of the suggested five-lncRNA signature in the three GC cell lines and 20 healthy tissue samples  for TWIST1. This study showed that there is no significant correlation between the expression level of UCA1 and TWIST1 in tissue samples which is in contrast to our expectations for UCA1 expression. We also examined the expression level of PART1, UCA1, DIRC3, HOTAIR, and HOXA11AS have more differential expression in the tumor tissues versus normal counterpart margins. Moreover, ROC curve analysis showed that these lncRNAs have significant sensitivity and specificity (diagnostic) values. Furthermore, we investigated the role of the lncRNAs through enrichments and real-time PCR.In our investigation, we used the TCGA RNAseq data to introduce a lncRNA signature that can be utilized as a diagnostic biomarker. Our bioinformatics data illustrated that lncRNAs et al. declared that a long non-coding RNA signature can predict the prognosis of the glioblastoma multiform patients  by inhibiting miR-200b expression in NSCLC . This examination showed that UCA1 was up-regulated and down-regulated in 8 and 3 samples, respectively. In the other 9 tissue samples, UCA1 did not show altered expression. Altogether, the evaluation of UCA1 using paired t-test did not show a significant statistical relationship in twenty tissue samples; this was another important contradiction that we encountered during this study. Discrimination in sampling (selection of samples with high TWIST1 expression) seems to be one of the reasons for this unexpected result; However, based on bioinformatics analysis, our sampling was correct and the results should have been different.  Activation of the PI3K-Akt pathway is reported in many malignancies. Inhibition of the PI3K-Akt pathway can induce apoptosis and decrease cell division via negative regulation of  in vivo , 52. One process . TWIST1 Overall, like many other studies, our study had its limitations. First, the data used for this study to introduce a diagnostic signature were extracted from a single database (TCGA); the use of more databases such as GEO certainly makes the results more reliable. Second, due to the use of cell lines in this study for investigation of suggested signature expression and the differences between these cells in terms of growth conditions and cumulative mutations, as well as the low number of cell lines used, it is necessary to examine the expression of these 5 lncRNAs in more tissue samples.PART1, UCA1, DIRC3, HOTAIR, and HOXA11AS showed more differential expression in the TCGA RNAseq data of GC patients. Additionally, drawing ROC curves exhibited them properly as biomarkers. Enrichment analysis confirmed the role of the signature in the critical biological processes and pathways. Furthermore, we predicted the target genes of the signature and protein-protein interactions among them. So far, all surveys have supported the biomarker role of this signature, however, the results of real-time PCR in cell lines and tissue samples were inconsistent with the findings of the analysis of RNAseq TCGA data, which is difficult to fully interpret using the findings of this study. The bioinformatics analysis of TCGA RNAseq data presented a lncRNA signature that seemed to be useable as a diagnostic biomarker. The proposed lncRNA signature including We could not conclusively present a five-lncRNA signature with diagnostic potential for GC because of some identified contradictions in bioinformatics and laboratory study. More investigations should be performed for ultimate validation or rejection of this signature: investigations that cover our faults using a large sample size, examination of these results in tissue samples, and also a further exploration of the biological and molecular mechanism of the suggested five-lncRNA signature in GC progression. GM, PA, and AMR Conceived the study. GM, PA, AMR, AA, and TN Performed interpretation and analysis of data. GM and PA Prepared the manuscript. AMR Revised the manuscript for important intellectual content. AMR provided supervision.This study was funded by The Vice Chancellor for Research at Mashhad University of Medical Sciences (no. 13986). The authors declare that they have no competing interests.


Lophomonas spp., is an emerging protozoan parasite that belongs to the Parabasalids  which infects the respiratory tracts of humans. Despite the presence of a few reports of human lophomoniasis, the true burden of Lophomonas infection is unknown. This systematic review aimed to elucidate the latest global status of publications reporting human cases of lophomoniasis as a new emerging protozoal disease.  A comprehensive and systematic search was performed in 10 (five English and five Persian) databases for studies reporting cases of lophomoniasis between 1993 and March 2020 (27 years). Then, the selected articles were carefully reviewed and screened based on the inclusion and exclusion criteria. n = 171; 55.7%, P < 0.04) significantly belonged to the juvenile age group (aged ≤18 years). The male to female ratio of the cases was almost equal, and no statistically significant difference between them was observed. The maximum number of cases (n = 237) was reported from Iran. Most cases  had no history of underlying diseases/organ transplantation (P < 0.001). Moreover, the BAL specimen was the most commonly used clinical sample to diagnose lophomoniasis (P < 0.001).  Overall, 32 eligible publications reporting 307 lophomoniasis cases from around the world are included in this review. The patients were aged between 1 month and 84 (mean age = 23.7 years). The highest number of cases (Lophomonas spp. should not be considered as an opportunistic infection. Thus, current work sheds light on some controversial issues regarding the epidemiological aspects of lophomoniasis. Our findings reveal that the prevalence of lophomoniasis is likely to be markedly underestimated when evaluated based on published case reports. Additionally, our data, at least for the time being, supports the idea that  Lophomonas spp. is an anaerobic protozoan parasite that belongs to the Parabasalids  [Trichomonas vaginalis and Trichomonas tenax), Tritrichomonadidae (Tritrichomonas foetus), Lophomonadidae (Lophomonas blattarum and Lophomonas striata), and Holomastigotoididae (Spirotrichonympha) [Lophomonas has been isolated from the human respiratory system and the guts of certain insects, mainly cockroaches, as part of their microbiome [Lophomonas as an emerging infectious pathogen remain unknown [onadida) . The Paronympha) , 2. To dcrobiome . Humans crobiome . Followicrobiome , 4. Howe unknown .L. blattarum, has increased worldwide [During the last decades, the incidence of human lophomoniasis, caused by orldwide . The firorldwide . Similarorldwide , Turkey orldwide , India [orldwide , Peru [9orldwide , and Mexorldwide .This protozoan parasite can infect both the upper (mainly sinuses) and lower (mainly lungs and bronchia) respiratory tracts, with nonspecific clinical features such as chronic cough, hemoptysis, dyspnea, and wheezing , 4.Currently, microscopic examination is used as a gold standard for the diagnosis of lophomoniasis using several clinical samples such as bronchoalveolar lavage (BAL) fluid, bronchial/tracheal aspirate, and sputum samples –11. HoweLophomonas infection remains controversial.Based on published studies, metronidazole is a drug of choice for lophomoniasis treatment with satisfactory outcomes , 13–16. Lophomonas infection is unknown. Therefore, determination of the epidemiological aspects of lophomoniasis worldwide could be useful to estimate the global public health burden and to manage control plans. Given that there are some controversial issues regarding the epidemiological aspects of lophomoniasis, our study aims to obtain data which will help to address some of these gaps. Thus, the purpose of this systematic review was to estimate the global prevalence and distribution pattern of this enigmatic parasitic disease.Despite the presence of a few studies on the subject of human lophomoniasis around the world, the true burden of the Lophomonas,” “Pulmonary,” and “Respiratory” alone or in combination with “OR” and/or “AND.”To achieve all reports on human lophomoniasis, we performed a systematic review of the literature, following the preferred reporting items for systematic reviews (PRISMA) guidelines . Two resA search of the related literature in 10 databases was conducted to evaluate the reports on lophomoniasis. Five English databases  and five Persian databases , and Scientific Information Database) from 1993 to March 2020 (27 years) were searched.The selected articles, after completing the search, were reviewed by two scholars independently. After reviewing the title, abstract, and full text of the articles, all the duplicate and unrelated studies were eliminated. Furthermore, to avoid republish bias, the results of the articles were attentively investigated and duplicates were excluded. Additionally, to avoid omitting any additional qualified studies, all references cited in the original and review articles were checked. Because of the limitation of access to theses and conference papers, they were not deliberated on in this paper. All the references listed in the selected articles for completing the checklist were investigated manually.All chosen articles were commentaries to recognize the potentially eligible articles by the two researchers using a piloted form. The definitive determination of eligibility or exclusion from studies was made separately. Disagreements were resolved by a third reviewer with insight. After duplicate entries were removed, data were extracted from selected studies with at least one of the following inclusion criteria: case report, case series, cross-sectional, case-control studies, and letters to the editor corresponding to determining the prevalence of lophomoniasis. The exclusion criteria contained the following: (1) abstracts of articles that were not available in the English language; (2) review articles; (3) summaries of articles presented as proceedings at conferences; and (4) studies that were carried out on insects. The PRISMA flowchart of the study plan is shown in Of the retrieved articles, 32 were eligible for inclusion in this systematic review. The following information was extracted: first author, year of publication, place of study, number of patients, gender, age, specimen, and patients' past medical history (underlying disease and/or organ transplantation).t-test were applied to comparisons of proportions in each group, and P < 0.05 was considered to indicate significance.Data analysis were performed using SPSS v16  and revealed as proportions (%). Furthermore, the chi-squared test and two-tailed We identified that among 846 studies in the literature search, 32 records were potentially appropriate for inclusion in this systematic review. About 30 of the articles were full text, and 2 were a letter to the editor . Figure n = 237; 77.2%), China , Turkey , India , and Malaysia . Twenty-two cases were reported from South American countries: Panama , Peru , and Mexico . Three cases were from Spain  and Egypt   .P < 0.04). The patient's age was not recorded in 21 (6.8%) cases. Moreover, of all the examined cases, 130 (42.3%) were female and 155 (50.5%) were male (P < 0.13). The patient's gender was not recorded in 22 (7.2%) cases.The patients were aged between 1 month and 84 years, with an average age of 23.7 years. Of the 307 cases, 171 (55.7%) were significantly juvenile (aged ≤18 years) (n = 196) of cases significantly had no history of underlying diseases and or organ transplantation (P < 0.001). In contrast, one hundred eleven (36.1%) of the patients had a history of underlying diseases and or organ transplantation as follows: 24 (21.6%) suffered from cancer, 15 (13.5%) asthma, 12 (10.8%) organ transplantation, 10 (0.9%) tuberculosis, 8 (7.2%) COPD, 6 (5.4%) heart failure, 7 (6.3%) corticosteroid therapy, 5 (4.5%) sinusitis, 5 (4.5%) renal failure, 4 (3.6%) diabetes, 1 (0.9%) HIV, and 14 (12.6%) others, respectively. Furthermore, BAL samples were the most  commonly used specimens to detect Lophomonas infection than other ones (P < 0.001). In addition, the microscopic examination was used to diagnose the infection in all studies, except one.Moreover, the most remarkable result to emerge from the data is that the majority . Lophomoniasis is a neglected protozoan parasitic disease which occurs only in a few countries of 4 continents. However, far too little attention has been paid to lophomoniasis in view of new emerging protozoa.This review, to the best of our knowledge, is the first global lophomoniasis systematic study based on published reported cases. Our findings demonstrated a 91.85% prevalence of Lophomonas parasite worldwide, its true burden remains underestimated as the most neglected tropical infection in various parts of the world. However, most of the studies conducted on lophomoniasis almost exclusively focused on case reports worldwide [The data reveal significant differences in the number of Iranian patients. In a total of 307 examined patients, 237 (77.2%) were from Iran, 29 9.4%) China, 19 (6.2%) Panama, 11 (3.6%) Turkey, 4 (1.3%) India, 2 (0.65%) Mexico, 2 (0.65%) Spain, 1 (0.3%) Egypt, 1 (0.3%) Malaysia, and 1 (0. 3%) Peru. Since many researchers and physicians are not familiar with % China, It should be noted that Li and Gao  in a pren = 155) and female (n = 130) lophomoniasis cases (P=0.13). Overall, these findings are in contrast to most of the findings that have been recorded by other researchers [Moreover, our data revealed that there were no significant differences between male . Therefore, young age can be a risk factor for lophomoniasis. This has been previously assessed only to a very limited extent because studies on Lophomonas in juveniles have recently increased [A further novel finding is that there was a significant difference between adults (44.3%) and juvenile patients (55.7%) with ncreased , 16, 35.Lophomonas infection and subjects without underlying disease or organ transplantation (P < 0.001). It means that, according to the findings of the present study, this infection occurred significantly more often in immunocompetent versus immunocompromised subjects . Even so, there is no evidence regarding the opportunistic essence of this newly emerged parasite. However, our results contrast with the idea that most researchers believe that Lophomonas is an opportunistic parasite [Our findings show a remarkable correlation between parasite , 40. ThuLophomonas and COVID-19 should be ruled out. Hence, early diagnosis and treatment of this dual infection are critically important because misdiagnosis of this protozoan infection could cause severe complications and increase the duration of hospitalization [Moreover, we believe that during the coronavirus (COVID-19) pandemic, comorbidity of lization –43.Lophomonas significantly more frequently than other respiratory specimens (P < 0.001). This is in good agreement with previous studies [Furthermore, our data showed that the BAL specimen was the most commonly used to detect  studies , 40, 43.Lophomonas was confirmed using a PCR test. Although the microscopic examination is currently used as a routine technique for detecting Lophomonas, it has low sensitivity and specificity when compared to a molecular method for distinguishing the protozoa from bronchial ciliated epithelial cells [Lophomonas proliferation, on the other hand, was quite unsuccessful, and there is not available media for this parasite [Lophomonas is strongly recommended [The most surprising finding from the current work is that only in one study, the presence of the al cells , 41, 43.parasite . As a reommended , 43.Lophomonas, however, reveals a number of gaps and shortcomings. Despite decades of reporting on the first Lophomonas human infection, there are key questions and notions that are still not discussed in the literature, such as its vertebrate and invertebrate hosts, transmission modes, virulence, and diagnosis method. This is a critical issue for future research in order to provide a comprehensive picture of the epidemiological aspects and clinical manifestations. Advances in diagnostic methods and examination of Lophomonas infection in animal hosts (as a possible reservoir) will be helpful for our knowledge about risk factors related to lophomoniasis and the development of appropriate public health interventions.A closer look at the literature on Lophomonas showed that the parasite still remains an enigmatic issue for many scientists around the world in the present era [This systematic review indicates that lophomoniasis is common in several tropical countries, mainly in Asian countries such as Iran, where there is an appropriate status for the breeding of cockroaches and termites , 44. We sent era .Lophomonas infection than immunocompromised ones and adults. Thus, these data support the idea that Lophomonas spp. should not be considered as an opportunistic pathogen. However, current work sheds light on some controversial issues regarding epidemiological aspects of lophomoniasis. Additionally, our findings revealed that the incidence of lophomoniasis is likely to be markedly underestimated when evaluated based on published case reports. Overall, further investigations are required to estimate the exact burden of the disease in other parts of the world. As a whole, this review provides valuable information regarding some epidemiological aspects of lophomoniasis worldwide, which will likely be very favorable for management and control programs of this disease.Our systematic review showed that lophomoniasis has been reported from 10 countries in 4 continents, mostly from Asia. Moreover, immunocompetent and juvenile subjects are more susceptible to Due to practical constraints, this work cannot provide a comprehensive review of lophomoniasis cases. In this investigation, a number of important limitations need to be considered: first, a significant lack of access to articles in other languages, particularly Chinese. Second, some publications were not included in the analysis when nondefined parasite names had been cited, such as protozoa forms, ameboflagellates, and hypermastigotes.


Xanthomonas campestris pv. campestris (Xcc), would be desirable to avoid significant crop yield losses. The main aim of this work was to develop a method of detection of Xcc infection on broccoli leaves. Such method is based on the use of imaging sensors that capture information about the optical properties of leaves and provide data that can be implemented on machine learning algorithms capable of learning patterns. Based on this knowledge, the algorithms are able to classify plants into categories . To ensure the robustness of the detection method upon future alterations in climate conditions, the response of broccoli plants to Xcc infection was analyzed under a range of growing environments, taking current climate conditions as reference. Two projections for years 2081–2100 were selected, according to the Assessment Report of Intergovernmental Panel on Climate Change. Thus, the response of broccoli plants to Xcc infection and climate conditions has been monitored using leaf temperature and five conventional vegetation indices (VIs) derived from hyperspectral reflectance. In addition, three novel VIs, named diseased broccoli indices (DBI1-DBI3), were defined based on the spectral reflectance signature of broccoli leaves upon Xcc infection. Finally, the nine parameters were implemented on several classifying algorithms. The detection method offering the best performance of classification was a multilayer perceptron-based artificial neural network. This model identified infected plants with accuracies of 88.1, 76.9, and 83.3%, depending on the growing conditions. In this model, the three Vis described in this work proved to be very informative parameters for the disease detection. To our best knowledge, this is the first time that future climate conditions have been taken into account to develop a robust detection model using classifying algorithms.A rapid diagnosis of black rot in brassicas, a devastating disease caused by  Brassica oleracea var. italica) has become increasingly attractive and profitable. It is highly regarded for its nutritional value and also its antioxidant and anticancer properties .The cultivation of broccoli . The most notable pathovar of X. campestris is campestris (Xcc), which is the causal agent of black rot of crucifers and affects all cultivated brassicas. Indeed, and according to the report elaborated by EIP-AGRI Focus Group for the European Commission, Xcc is a threat to the production of broccoli, cauliflower, and cabbage throughout Europe . Moreover, Xcc can be subdivided into nine races on the basis of the responses they induce on different cultivars. Among the nine races described for Xcc, races 1 and 4 are considered the most virulent and spread worldwide  is an indirect measurement of the vegetation transpiration rate , which is related to vitality of canopies Tucker, . Other Va et al.  used a cC-TA) and five already known VIs. Moreover, three novel VIs specifically designed for detecting the Xcc infection were extracted from the reflectance spectra of healthy and diseased broccoli leaves. They were named diseased broccoli indices (DBI1-DBI3). This set of nine parameters were implemented on a selection of algorithms widely used on precision agriculture for their success in classifying infected plants: the multilayer perceptron-based artificial neural network (MLP), the support vector machine (SVM), and the k-nearest neighbor (kNN). Finally, the suitability of the trained models was evaluated by comparing their performances in correctly classifying new samples as healthy or diseased leaves under a range of climate conditions, including intermediate and extreme climate change scenarios, as well as current climate conditions. Furthermore, the relevance of every input parameter for the detection of Xcc infection in broccoli plants was evaluated.The main aim of this work was to develop an efficient method for the detection of Xcc infection in broccoli plants based on thermal and hyperspectral reflectance imaging on individual leaves. For this purpose, six parameters were recorded: leaf temperature  were grown in a growth chamber in a 16/8-h day/night regime with 60% relative humidity, 200 mol photon m−2 s−1 of PAR light. The ambient temperature and CO2 concentrations (The Ctrations  were choXanthomonas campestris pv campestris (Xcc) race 1 and race 4 were grown for 24 h at 28°C in LB (Luria-Bertani) plates. Bacterial suspensions were prepared in sterile 10 mM MgCl2 at 108 colony-forming units per ml (cfu mL−1) by adjusting the optical density at 600 nm to 0.1.2 or inoculated with bacterial suspension by clipping four secondary veins per leaf with rat tooth tweezers previously dipped in the corresponding solution (The third leaf of four-week-old plants (under CCC or RCP 4.5) or five-week-old plants (in case of RCP 8.5) was mock-inoculated with sterile 10 mM MgClsolution . For eacThermal images of whole leaves were recorded using a FLIR A305sc camera  vertically positioned 30 cm above the leaf, according to Pérez-Bueno et al. . For eacReflectance spectra  of broccoli leaves were recorded using a Pika L hyperspectral imaging camera  in the visible (400–700 nm) to near-infrared spectral range , with a spectral sampling at 2.1 nm and a spectral resolution  of 3.7 nm. The camera was positioned vertically 45 cm over the sample, which was placed on a translation stage. Thus, a datacube with 281 images was built for each attached leaf.Leaves were illuminated with four calibrated xenon lamps with homogeneous light intensity between 400 and 1,000 nm, positioned above the samples and around the camera. Previous to leaf measurements, dark and light corrections were made in darkness and illuminating a white homogenous calibration tile provided by Resonon, respectively. Dark and light corrections, build-up of datacubes and analysis, were carried out with the software Spectronon v. 2.134 (Resonon).Reflectance spectra averaged for whole leaves were obtained and used to calculate images corresponding to several widely used VIs, as summarized in Numerical data obtained from thermal and reflectance images (including reflectance spectra and VIs) were managed using Microsoft Office Excel 2016 . Aiming to design a simple method of detection, values were averaged from whole leaves rather than regions of interest.t-test (Microsoft Excel) was performed to compare, for every treatment and at every dpi assayed: (a) spectra reflectance profiles; (b) values of novel VIs (DBIs). The null hypothesis was that there were no differences between treatments. This hypothesis was considered false at p < 0.05, and variables were treated as different when p-value was below this value. Figure graphs were plotted using Microsoft Excel.Two-tailed Student's C-TA, NDVI, PRI, ARI, CRI, WBI, DBI1, DBI2, and DBI3) at every dpi (1–9 dpi) and treatment . In addition, data were rescaled from zero to one to ensure comparison between treatments and days, according to the equation: rescaled value = (x-minimum)/maximum. Then, the three databases were randomly partitioned into training and test datasets, in a proportion of 7:3, respectively , one per climate condition. Each database contained values of selected parameters  organized in layers transmit information to each other, learning from both input and output data Hahn, . In contF-measure . All the three feedforward backpropagation MLPs tested were designed to have one hidden layer with four neurons . Higher number of hidden layers or their neurons did not result in an improvement of the performance. A polynomial kernel was used for the SVMs, with bias = 1 and gamma = 1. More complex spatial kernels did not improve the performance of the algorithm. Finally, the optimal number of neighbors for the kNN algorithm was k = 5 using the Euclidean distance. Regarding SVM and kNN libraries, we have used the basic nodes  implemented on Knime software v. 4.3.2. This process was performed independently for each of the three climate conditions under study.Broccoli leaves were categorized into mock-control, Xcc race 1-, and Xcc race 4-infected plants using the classifying algorithms MLP, SVM, and kNN. The performance of classification was evaluated in terms of (i) sensitivity (true positive rate); (ii) specificity (true negative rate); (iii) accuracy (percentage of right guesses); and (iv) C-TA, NDVI, PRI, ARI, CRI, WBI, DBI1, DBI2, and DBI3), that is, how informative was a given parameter for the model to make a correct decision. For such a purpose, global surrogate random forest (RF) models were trained to estimate the variable importance using the Global Feature Importance component developed for Knime software . RF was trained on the standardly pre-processed input data. Feature importance was then calculated by counting how many times it had been selected for a split and at which rank (level) among all available features (candidates) in the trees of the RF.Finally, the global variable importance was calculated for each parameter , whole leaf reflectance spectra were registered at 1, 2, 3, 6, and 9 dpi. Those profiles revealed specific spectral patterns for each treatment, showing clear differences between them from the first timepoint measured , 2. Thusp < 0.05) between treatments. Thus, three spectral ranges were found to show maximal differences between treatments at every dpi tested: 400–500, 600–700, and 900–1000 nm. On the contrary, the regions 520–570 and 730–890 nm were very stable and not affected by the infection until 9 dpi. For this reason, they were selected for “normalization” of the designed parameters. Several wavelengths were chosen within these spectral regions of interest. To avoid redundancy of adjacent wavelengths in the reflectance spectra, only wavelengths separated by at least 40 nm were used in this process. Finally, the selected wavelengths were combined by different mathematical calculations  in order to find novel VIs showing statistical differences (p < 0.05 according to Student's t-test) between treatments along entire experiments at each climate condition. Among the large collection of proposed VIs, three of them, named diseased broccoli indices 1–3  for every treatment and dpi assayed. The three databases were normalized and then randomly split in two datasets: training (70%) and testing (30%). Each of the three training dataset was used to feed supervised classifying algorithms  to classify samples into the following categories: mock-control, Xcc race 1-, and Xcc race 4-infected leaves; each of the three testing dataset was used to calculate their performance of classification.For each experimental condition, an independent database was built containing the values of selected parameters  was inefficient. This was probably due to the underrepresentation of mock-control samples in the datasets , whereas in three-category models, every group had the same size.The MLPs provided the highest accuracy for every climate condition . They al1, DBI2, and WBI. In contrast, under RCP 4.5 conditions, TC-TA, DBI3, and PRI obtained the highest global variable importance. Finally, under RCP 8.5 conditions, DBI2, CRI, and NDVI were the most instructive parameters.The suitability of the designed VIs for the identification of Xcc-infected leaves was evaluated in terms of global variable importance in the classifiers, calculated by a surrogate RF algorithm . The accImaging techniques appear to be essential for precision agriculture due to their fast time-spatial response to biotic and abiotic stress in a non-destructive manner , the symptoms developed under CCC triggered by races 1 and 4 were similar. However, Xcc race 1 showed a slower timecourse than race 4 under the same ambient conditions. The process of undergoing climate change could affect considerably plant biochemistry and therefore plant defense responses. For this reason, it is relevant to include future climate conditions in disease detection studies. Indeed, each disease may respond differently to these variations, and thus, climate change would cause neutral, positive, or negative effects on plant responses to diseases , as well as on the physiology of Xcc races, and/or their interaction with host plants.According to Fargier and Manceau , when a In literature, a number of classic VIs derived from multispectral  imaging can be found. This VIs can be used to detect, classify, and quantify specific diseases with varying degrees of success  and other common hyperspectral reflectance parameters , were implemented in three different supervised classifiers  for each experimental condition. Since the learning process of each algorithm differs from each other, so will the quality of its predictions on the new samples. In this sense, it is a common procedure to compare the performance of several algorithms when sorting new samples  after training on the same dataset. Metrics such as specificity (true negative rate), sensitivity (true positive rate), accuracy (percentage of right guesses), or F-measure (harmonic mean of precision and sensitivity) evaluate the performance of classification, that is, the estimation of the true risk of error of the output prediction of a machine learning algorithm  to identify infected plants at conditions resembling CCC. For example, Sankaran et al.  with 94–100% accuracy depending on the pathogen. Pérez-Bueno et al. (Rosellinia necatrix) with accuracies up to 82.5%. Nguyen et al. , which in turn reduces significantly the influence of the unknown variability. Therefore, MLPs are usually more robust models that often outperform other classifying algorithms in solving a variety of classification problems  of the infections, as discussed above.DBI1, DBI2, and DBI3 here presented are good reporters for Xcc infection in broccoli leaves. Furthermore, the model based on MLP and the set of parameters DBI1, DBI2, and DBI3 along with common VIs  and TC-TA would be an effective procedure for the identification of Xcc-broccoli infected plants. In addition, this model proved to be robust regardless of the climate conditions.The parameters DBIThe original contributions presented in the study are included in the article/MP, MLP-B, and MB conceived and designed the experiments and wrote the manuscript. MP and MP-B conducted the experiments. MP analyzed the data, interpreted the results, and mounted images. MB contributed to the materials, equipment, and analysis tools. All the authors reviewed it and approved the final version.This work was supported by grant RTI2018-094652-B-I00 funded by MCIN/AEI/10.13039/501100011033 and by “ERDF: A way of making Europe”. The free open access publication was partially funded by Consejo Superior de Investigaciones Científicas (CSIC) through the Unidad de Recursos de Información Científica para la Investigación (URICI).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Furthermore, there was evidence of an association between various susceptibilities, particularly between measles and chickenpox (OR: 4.38). Conclusion: this study showed that even if the majority of our healthcare professionals are immunized for MRV, it is necessary not to underestimate the seronegativity of non-immune ones. All health professionals should be vaccinated to ensure safety for patients, especially the weakest.Immunization is the best protection against chickenpox, measles and rubella. It is important to identify and immunize susceptible healthcare workers to prevent and control hospital infections. Our aim was to estimate the susceptibility level of healthcare workers at a Teaching Hospital in Rome concerning these diseases and the factors associated to the susceptibility. Methods: a cross sectional study was carried out at the Department of Occupational Medicine of the Umberto I General Hospital of Rome. Participants were recruited during routine occupational health surveillance. As far as inclusion criteria, the following professionals were considered: doctors, nurses, laboratory technicians and other health professionals. Concerning the exclusion criteria, patients with HIV, cancer and diseases of the immune system, and acute illness or fever more than 38.5 °C, were not included in the study. A blood sample was tested for the presence of antibodies against measles, rubella and chicken pox. Results: 1106 healthcare professionals were involved in the study : 25 (2.3%), 73 (6.6%) and 35 (3.2%) of these were susceptible to measles, rubella and chicken pox, respectively. The only variable associated with susceptibility of measles was age ( Vaccination plays a fundamental role in eliminating the diseases for which it is available , includiFor populations considered to be at high risks, such as healthcare workers (HCW), vaccination is recommended by the national vaccine prevention plan (PNPV) . In thesVaccination is important because these diseases can infect not only healthcare workers but also hospitalized patients, fragile by definition .In general, there is a lack of a unified global policy towards vaccinations for the diseases examined, an indication of a low level of attention regarding the issue, even among healthcare workers (HCW) , as repoTwo surveys conducted in Central and South Italy have shown that healthcare workers (HCWs) have a significantly greater susceptibility to preventable disease vaccines (VPDs) ,10. FurtIn addition, it must be considered that seronegativity is not exclusively related to non-vaccination, but could also be a consequence of a failure to respond to the primary vaccination course.A study conducted in Italy on the seroprevalence of anti-measles and anti-rubella IgG among medical students and trainees vaccinated in childhood with two doses of MMR showed seronegativity rates of 15% and 9% respectively, seroconversion after a booster dose of MMR occurred in 74% of cases for measles and 98% for rubella .At the European level, a seronegativity rate for measles in HCWs is estimated to be around 6%  while a (a)The susceptibility level of healthcare workers at a Teaching Hospital in Rome, as a high-risk population, concerning VPD and in particular towards chickenpox, measles, and rubella;(b)The factors associated to the susceptibility.The study aims to estimate:A cross sectional study, according to the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement, was carried out between February 2017 and January 2020 .Participants were recruited during routine occupational health surveillance in teaching hospital ‘Policlinico Umberto I’ of Rome. As far as inclusion criteria, the following professionals were considered: doctors, nurses, laboratory technicians and other health professionals. These health care workers work in 15 different integrated care departments (DAI) such as surgical, clinical and health services. Concerning the exclusion criteria, patients with HIV, cancer and diseases of the immune system, and acute illness or fever more than 38.5 °C, were not included in the study. During medical examination, social-demographic, clinical and occupational data were collected. After obtaining informed consent from the health care workers, in order to control and prevent nosocomial transmission of measles, rubella and chicken pox, a blood sample was taken by the staff of the Occupational Medicine Unit. This sample was then tested for the presence of antibodies against measles, rubella and chicken pox. Antibody titration was performed by the staff of the Microbiology and Viral unit using a semi-quantitative immune-enzymatic test (ELISA method) following the protocol suggested by the producer .-Negative 0—<0.100-Grey zone 0.100–0.200-Positive > 0.200The ranges used for the evaluation of VZV-IgG and anti-measles IgG are:-Negative 0.0–0.89-Grey zone 0.9–1.09-Positive > 1.10The ranges used for the evaluation of anti-Rubella IgG are:Sensibility and specificity of the tests were 97%, 99%, 90.6%, and 98.2%, 97%, 100% for measles, rubella and chickenpox, respectively.-Population size: 6550 HCWs;-Population proportion of seroprevalence: 80%;-Margin of Error: 3%;-Confidence level: 99%.Sample size was calculated using the following parameters:Sample size calculations indicated the need to recruit at least 1003 HCWs.t-test or the Mann-Whitney U test was applied for two-group comparisons, and ANOVA and the Kruskal-Wallis test were used for comparisons of more than two groups. The Kolmogorov-Smirnov test was used to verify the normal distribution of quantitative variables. The Pearson’s correlation coefficient was computed to estimate the direct or indirect correlation between variables. Statistical analysis was performed using mean, standard deviation (SD), median, and minimum and maximum values for quantitative variables. For qualitative variables, frequencies and percentages were computed. Student’s Differences in susceptibility to exanthematic diseases were assessed using univariate analysis.Multivariate analysis was conducted using a multiple logistic regression model, considering the following explanatory variables: gender, age, type of department, qualification and typology of company . Susceptibility to infection was considered as a dependent variable.Results of the logistic regression models were presented as Odds Ratio with 95% confidence interval.p-value of 0.05.All statistical analyses were performed using SPSS for Windows, release 25.0 . The statistical significance was set at a A total of 1,106 healthcare professionals were examined including 336 doctors (30.4%), 462 nurses (41.8%), 136 laboratory technicians (12.3%), 167 other health professionals (15.1%). Of these 460 were males (41.6%) and 646 females (58.4%). Age ranged from 25.4 to 70.9 years (mean 54.1 ± 8.8 standard deviation).History of prior infection was reported by 51.8%, 40.7% and 77.4% for measles, rubella and chickenpox, respectively.In p < 0.001).Of the 1106 healthcare workers surveyed, 25 (2.3%) 73 (6.6%) and 35 (3.2%) were susceptible to measles, rubella and chicken pox, respectively. These data are indicative of a better situation than that reported a decade ago by other Italian studies ,13,17,18Except for a lower chance of being susceptible to measles with increasing age, no significant differences were found for other socio-demographic or occupational variables, such as gender, use in medical/surgical areas (type department) or the relative structure. Different situation reported in a similar study of Qatar hospital, in which the seropositivity rate of measles in physicians (93.8%) was significantly higher than nurses (83.9%) and allied HCWs (80.5%). As for the age, those aged > 40 years were more seropositive than 30–39-year-old workers (79.0%) and HCWs aged < 29 years (81.5%), with a predominance of varicella seropositive rate in HCWs with longer service .p < 0.0001), indicating the age over 16 months for a better MMR vaccine efficacy due to a better humoral and cellular immune responses. The Cox analysis suggests that a younger age at the time of the first and second doses of MMR vaccine, an older age at enrollment, and a longer time from the second dose of MMR vaccine to the antibody titer evaluation seem to be risk factors for the persistence of circulating antibodies. However, this study shows that after an MMR vaccine booster dose there was a seroconversion of 74% of seronegative HCWs and the overall seroconversion rate after a second dose (booster) was 93% [Results of an Italian retrospective cohort study in which the long-term immunogenicity of measles vaccine was evaluated in a sample of two thousand immunized (2 doses of measles-mumps-rubella [MMR] vaccine) students and residents of the University of Bari, reported that in 305 of these (15%) no protective anti-measles IgG were detected and this result was not in line with that of our study and with the evidence of the literature. This proportion was higher among subjects who received vaccination at ≤15 months (20%) than in those who received vaccination at 16–23 months (17%) and at ≥24 months (10%)  where it was assessed whether Danish pediatric healthcare workers were protected against selected severe vaccine-preventable diseases (VPD). In particular, protective levels of IgG were found for measles (90.3%), rubella (92.3%). Moreover, they found seropositivity for all three MMR components in 421 75.9%) HCW, lowest in those younger than 36 years (63.3%). The immunity gaps found primarily in young HCW indicate a need for a screening and vaccination strategy for this group. Considering the poor correlation between self-reported immunity and seropositivity, efforts should be made to check HCW’s immune status in order to identify those who would benefit from vaccination [5.9% HCW,Several studies conducted in European and non-European countries were carried out to determine the seroprotection against measles, rubella and chickenpox among HCWs, but the strength of our study is the large sample on which to perform the statistical analysis, and there is evidence that training on vaccination and mandatory measures may be needed in order to achieve better coverage .In the occupational setting a screening and vaccination strategy for healthcare workers is needed, especially considering that there is usually a low correlation between self-reported immunity and seropositivity . Some AuThe present study has some limitations. First of all, the study design is cross-sectional and a causal relationship between susceptibility and risk factors cannot be established. Moreover, the sample represents one fourth of all HCWs in the Teaching hospital and a selection bias cannot be fully excluded due to the voluntary participation in the survey. Finally, due to practical reasons related to the occupational health surveillance of HCWs, the study was carried out in a three-year period, and this can have introduced some confounding factors.In conclusion, even if our study shows that the majority of healthcare professionals are immunized for MRV, it is necessary not to underestimate the levels of seronegativity, that could be the reservoir to the developing of hospital-acquired and occupational infections. All health professionals should be vaccinated to ensure safety for patients, especially the weakest such as children, the elderly and immunosuppressed, remembering that in this segment of the population, a nosocomial infection can lead to the development of serious complications, sometimes even with a fatal outcome. In relation to the results obtained, we therefore suggest implementing the health surveillance program with a careful screening of seroprevalence towards MRV, to promptly recognize susceptible subjects and invite them to vaccination.