Garlic skin, a by-product of garlic processing, was supposed to improve the fermentation quality of high-moisture silages because of its low moisture content and active compounds. Thus, fermentation and microbial characteristics of high-moisture Pennisetum hydridum ensiled with the addition of 0, 10, 20, and 30 wt% garlic skin (on a fresh matter basis) were analyzed during a 60-days fermentation. Results showed that the addition of garlic skin increased the dry matter content and lactic acid production, and decreased the pH and ammonia-N content of the silage. Adding garlic skin changed the relative abundance of bacterial communities with an increase in Lactobacillus and a decrease in Clostridium relative abundance. In conclusion, co-ensiling of high-moisture Pennisetum hydridum with garlic skin could be a simple approach to improve the silage quality and nutrients preservation. Allium sativum L.) is a flavoring ingredient universally used with an annual production of approximately 20 million tons in the world , also known as king grass or Hybrid Pennisetum, is a perennial gramineous forage species, which has been widely planted in the tropics and subtropics of China. P. hydridum has high fresh biomass yields (85–300 t ha–1) and strong resistance to environmental threats  of Southwest University, Chongqing, China. The collected materials were chopped into 2–3 cm with a manual forage chopper without wilting. Garlic skin was from Tengda Farming Co., Ltd., . The chemical composition and microbial population of materials before ensiling are shown in P. hydridum (GS0); (2) 90 wt% P. hydridum+ 10 wt% garlic skin (GS10); (3) 80 wt% P. hydridum+ 20 wt% garlic skin (GS20); and (4) 70 wt% P. hydridum+ 30 wt% garlic skin (GS30) on a fresh matter (FM) basis. After thorough mixing, each mixture (approximately 300 g) was packed into vacuum-sealed plastic film bags . In total, 48 bags (4 treatments × 4 ensiling time points × 3 replicates) were prepared and stored at ambient temperature (25–35°C).The Pre- and post-ensiling materials were oven-dried at 65°C for 48 h to determine DM content and then ground (1 mm sieve) for chemical analysis. The DM and crude protein (CP) contents were determined and calculated according to the methods of the Association of Official Analytical Chemists . The con3-N) concentration was determined using the phenol-hypochlorite reaction method . The filtrates were centrifuged for 10 min at 10,000 × g, and the supernatants were filtered with a membrane (0.22 μm). The lactic, acetic, propionic, and butyric acids were determined using high-performance liquid chromatography  . The ammn method  and expr–1 to 10–7 in sterile saline solution. The population of lactic acid bacteria and coliform bacteria were separately incubated on Man Rogosa Sharpe agar and Violet Red Bile agar, and yeasts and molds were separately incubated on Rose Bengal agar. Colonies were counted as numbers of viable microorganisms in the colony-forming unit (CFU) per gram of FM.The microbial population was analyzed by using the plate count method as described by PRJNA755145.Total DNA was extracted from the silage samples using the HiPure Soil DNA Kits  according to the manufacturer’s instructions. The PCR reactions were conducted in triplicate 50 μL mixture . The V3-V4 regions of 16S rDNA were amplified using primers 341F (CCTACGGGNGGCWGCAG) and 806R (GGACTACHVGGGTATCTAAT) by PCR  . RelatedQ-value > 20 by FASTP (version 0.18.0) and merged by FLASH (version 1.2.11), respectively. The paired and clean reads were merged as raw tags using FLASH with a minimum overlap of 10 bp and mismatch error rates of 2% as stated in The original sequencings were quality filtered by removing the sequence containing more than 10% of unknown nucleotides and the sequence containing more than 50% of bases with P. hydridum with garlic skin on chemical composition, fermentation, microbial population, the dominant genus of bacteria, and the predicted bacterial functions were evaluated by one-way ANOVA with Tukey’s HSD post-hoc tests in the R program (v. 3.1.1).1 Statistical significance of the principal coordinate analysis was performed using analysis of similarities (ANOSIM) and permutational multivariate analysis of variance (ADONIS) in R program. A P-value of less than 0.05 was considered to be a significant difference.Microbial populations of forage and silage were log-transformed prior to statistical analysis. The effects of mixing P. hydridum and garlic skin silage during the ensiling process are shown in P < 0.01), and the CP content was decreased (P < 0.01) with the increasing garlic skin proportion. The CP contents in all silage treatments remained relatively stable during the 60 days of ensiling in the current study. The NDF contents in P. hydridum silage mixed with garlic skin decreased (P < 0.01) compared with the P. hydridum silage ensiled alone, and the ADF contents were not influenced by the addition of garlic skin.The dynamic changes of the chemical composition of P. hydridum and garlic skin silage are shown in P < 0.01) with an intermediate value for GS10 silage (4.39). The lactic acid contents of GS0 silage decreased from days 7 to 60 of ensiling, and that of P. hydridum silage mixed with garlic skin increased days 7–30 of ensiling. The GS10 silage had the greatest lactic acid contents at days 60 of ensiling (P < 0.01). The acetic acid and NH3-N contents were greatest in GS0 silage (P < 0.01) at all-time points, and the contents of propionic acid and butyric acid were not detectable in GS10, GS20, and GS30 silages. In general, the lactic acid bacteria counts decreased as the ensiling progressed, and that was greatest in GS10 silage (P < 0.01), followed by GS10 and GS20 and least in GS0 silage at days 60 of ensiling. The counts of yeasts and molds were below the detectable levels after days 14 of ensiling. Although the counts of coliform bacteria declined consistently, it was still greater than 3 log10 cfu/g of FM in all silage treatments after the 60 days ensiling.The dynamics of the fermentation profiles and microbial population of P. hydridum ensiled with garlic skin is shown in P < 0.01, P. hydridum silage ensiled alone. At days 30 of ensiling, the Enterobacter (20.0–31.1%), Lactobacillus (5.7–46.6%), and Kosakonia (13.3–23.9%) were the dominant microbes in GS10, GS20, and GS30 silages while GS0 silage was dominated by Selenomonas (17.3%) and Enterobacter (10.5%). However, the relative abundance of Enterobacter at days 60 of ensiling decreased to 7.6–10.8% and the abundance of Lactobacillus increased to 45.8–68.2% in GS10, GS20, and GS30 silages.The alpha diversity of the bacterial community of Clostridium was greatest in GS0 silages, and garlic skin significantly increased the relative abundance of Kosakonia, Pantoea, Enterobacter, and Lactobacillus at days 30 of ensiling (P < 0.01). At days 60 of ensiling, the relative abundance of Lactobacillus and Kosakonia were still lower in GS0 silage compared with P. hydridum ensiled with garlic skin. Furthermore, the relative abundance of Prevotella and Caproiciproducens in GS0 silage was significantly greater than other silage treatments (P < 0.01).The relative abundance of P. hydridum silage ensiling process, bacterial function prediction analysis was performed by Tax4Fun based on the 16S rRNA gene sequences. At the first level of the KEGG pathway, the most abundance pathways were metabolism (∼79%) and genetic information processing (∼11%), followed by cellular processes (∼5%), environmental information processing (∼4%), human diseases (<1%), and organismal systems (<1%), which were not highly varied among silage treatments or between days 30 and 60 of ensiling. At the second level of the KEGG pathway, the relative abundance pathways of signal transduction, metabolism of cofactors and vitamins, energy metabolism, and infectious diseases were lower, and the relative abundance pathways of nucleotide metabolism, translation, and replication and repair were greater in GS10 silage compared with other silages at d 30 of ensiling  is similar to that reported in a previous study  and dried corn stover  suggests that the high mixing ratio of garlic skin might reduce the nutritional value of the silage. The P. hydridum has stemmy structures and accordingly has relatively high fiber content, which is in line with the previous study  or difficulty to pack and poorly fermented (DM > 40%) . The DM us study  and is fus study . Additio/g of FM , and WSC/g of FM . In the P. hydridum and garlic skin in the current study. In addition, DM contents in all silages consistently decreased as the ensiling progressed, which is in accordance with previous studies  was observed in the current study. The final pH value of the GS0 silage (5.08) after 60 days fermentation was still greater than the threshold pH value despite the consistent decline of pH during the fermentation process. The accumulation of acetic, propionic, and butyric acid content and decreased lactic acid in GS0 silage indicate the failure to inhibit the growth of spoilage organisms, such as clostridia that could ferment lactic acid to butyric acid  on P. hydridum silage fermentation quality  after 14 days of ensiling in the current study. Coliform bacteria are the major competitor of lactic acid bacteria and are responsible for silage nutrient loss (10 cfu/g FM) though the counts gradually declined with the decrease of pH during ensiling. It is suggested that coliform bacteria cannot be completely inhibited if the pH > 4.0  and in high-moisture corn stalk silage (28.9–39.3%), respectively. However, the relative abundance of Enterobacter at days 60 of ensiling decreased, which could be associated with the decreased pH value caused by the significantly increased relative abundance of Lactobacillus (45.8–68.2%). Ensiling is an anaerobic process, and the higher relative abundance of anaerobes, e.g., Lactobacillus, produces lactic acid to inhibit the acid-intolerant Enterobacter  were significantly different in GS10 silage compared with other silages at days 30 of ensiling. These differences might be related to the greater growth of entation .P. hydridum with garlic skin could significantly decrease pH value, butyric acid, NH3-N, and increase lactic acid production. Moreover, the relative abundances of favorable bacteria, such as Lactobacillus, were increased while that of undesirable bacteria, such as Clostridium and Enterobacter, were decreased. These results suggest that the addition of garlic skin could be a simple way to improve the fermentation quality and preserve protein of high-moisture P. hydridum silage via optimizing the moisture content and, thus, the inhibition of undesirable microbial growth during ensiling. Future research focusing on the utilization in animal husbandry and the related economic analysis are needed for better application in practice, and metagenomic and metabolomic analysis should also be considered to better characterize bacterial species and their functionality during ensiling.The current study illustrates that co-ensiling of high-moisture https://www.ncbi.nlm.nih.gov/, PRJNA755145.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: JC, GH, and ZZ designed the study and wrote the manuscript. HX, HQ, HZ, and YS performed the experiments. YL analyzed the data. YZ was involved in the manuscript revision. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.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.


Vehicular edge computing (VEC) is a potential field that distributes computational tasks between VEC servers and local vehicular terminals, hence improve vehicular services. At present, vehicles’ intelligence and capabilities are rapidly improving, which will likely support many new and exciting applications. The network resources are well-utilized by exploiting neighboring vehicles’ available resources while mitigating the VEC server’s heavy burden. However, due to the vehicles’ mobility, network topology, and the available computing resources change rapidly, which are difficult to predict. To tackle this problem, we investigate the task offloading schemes by utilizing vehicle to vehicle and vehicle to infrastructure communication modes and exploiting the vehicle’s under-utilized computation and communication resources, and taking the cost and time consumption into account. We present a promising relay task-offloading scheme in vehicular edge computing (RVEC). According to this scheme, the tasks are offloaded in a vehicle to vehicle relay for computation while being transmitted to VEC servers. Numerical results illustrate that the RVEC scheme substantially enhances the network’s overall offloading cost. The progress in the Internet of Things and wireless technologies put forward us prevalent smart devices like smart vehicles, which can execute numerous powerful and innovative applications . These aMobile cloud computing is introduced to cope with extensive computation requirements and greatly enhances computation performance and resource utilization. However, It also has its limitations due to the latency constraints while communicating with backbone networks. Since cloud servers are at a great distance from moving vehicles, this would lead to offloading effectiveness. Edge computing is presented as a way out, whereby it brings the services of the cloud to the edge of the network  and faciDue to the advancements in the internet of vehicles, there are more smart vehicles. These vehicles are armed with the computation unit, multi-communication technology, sensor platform, and human-machine interaction devices. With such progress in technology, it is more feasible for these smart vehicles to provide smart traffic applications, e.g., parking decisions, monitoring road traffic, and automatic management. Moreover, different multimedia onboard applications exist for both passengers and drivers . Such apThe applications having too stringent computation requirements in computation present many constraints to the vehicular terminals, specifically their computational resources. Since vehicles have low computation capacity and limited storage compared to VEC servers . To addrVEC servers provide reduced transmission cost and quick response to the computation offloading service due to proximity. Nevertheless, compared to the conventional cloud, the VEC server still faces resource limitations. VEC servers have a tight time duration to perform computation tasks, particularly for the VEC servers, which are situated in highly dense road segments and experience various computation demands. Also, vehicles can communicate in a relay via V2V multi-hop connections as a considerable number of vehicles present on the road. Therefore, exploiting the benefit of V2V communication and using underutilized computational resources leads in terms of load balancing and latency reduction in vehicular networks. Moreover, unlike that, some operators provide RSU access services, V2V communication is self-organized by moving vehicles, and its cost is lesser than Vehicle-to-Infrastructure (V2I) communication.Motivated by the facts mentioned above, we present a relay task/computation offloading scheme in vehicular edge computing (VEC). They are otherwise based on the precise file transmission prediction and period spent by the vehicle while on the road. The other vehicle may enter the transmission range of RSU_n the moment its task is completed. The computation result is transferred straightly from RSU_n to the other vehicle via V2I transmission without multi-hop backhaul. Hence, using both computation and communication resources of the vehicle can reduce the overall system time and cost for the task offloading and network congestion. However, the duration of transmission delay of V2V and V2I modes depends on the state of wireless channels, the size of the input file, and the density of the road since beacons are the packets sent periodically in a broadcast by vehicles to share its information like type, speed, direction computation capacity, and channel state .This paper aims to minimize the overall offloading cost, including the computation and communication cost, while meeting the VEC environment’s delay constraints to ensure vehicular user quality of experience (QoE).We proposed an RVEC scheme, which allows vehicles to transfer the task to their neighboring vehicle in their moving direction to the VEC servers via V2V multi-hop. The V2V computation is done in the neighboring vehicle case with sufficient resources, and the output returns to the vehicle in a relay fashion. This is conditional on the maximum delay tolerance and the vehicle’s stay time. This scheme the overall offloading latency and cost and network congestion.A distributed RVEC algorithm is designed for the proposed scheme. We evaluate the impact of various parameters on the RVEC algorithm by comparing it with different approaches. The results confirm that RVEC outperforms compared to the existing solutions to minimize offloading cost, i.e., computation latency and communication resource utilization.This paper put forward a promising relay task offloading scheme in vehicular edge computing (RVEC). In this work, both the heterogeneous demands of computation and communication tasks are examined by taking into account the vehicle dynamics. This scheme enhances transmission efficacy while meeting the computation tasks’ delay constraint. Moreover, RVEC reduces the overall cost of the offloading process. The significant contributions of this article are summarized as follows:The rest of this article is structured as follows. We review the related work in “Related Work”. The system model is described in “System Model”. In “Relay Task Offloading Scheme in VEC”, our proposed RVEC scheme is discussed, comprised of computation mode and communication mode. The distributed RVEC algorithm is presented in “The Distributed RVEC Algorithm”. We demonstrate numerical results in “Numerical Results” and conclude the paper in “Conclusion”.Recently, plenty of research has been done on the vehicular networks to use the underutilized vehicular resources. In The most encouraging technical method to enhance cloud computing efficiency, Mobile edge computing (MEC), has gathered the researchers’ substantial focus lately. Recent works on MEC aims at minimizing latency by offloading computation-intensive and latency-sensitive tasks to the neighboring RSUs or base stations for executing it remotely. In A vehicular cloud network can manage massive computation-extensive tasks flexibly and in a virtual manner . The worHowever, several studies emphasize vehicular cloud networks and MEC technologies; in contemporary literature, few papers provide MEC research with vehicular cloud networks. Moreover, the effect of several vehicular communication schemes on task offloading performance and vehicle mobility has been overlooked. However, in The RSUs connect each other via an optical-fiber backbone, and a central controller is also installed that controls the RSUs . Each RSc represents the required computational resources to complete task T. The c can be measured as the required number of CPU cycles, and the d in the equation shows the input file size of the computation task along with some necessary information on the computation task, e.g., the programing instructions, shared variables, methods, or link libraries, whereas Every Vehicle runs with uniform speed along the road, facilitated by vehicular communication protocols like dedicated short-range communication standards . The PoiN represents a total number of vehicle types. According to the type of computation task, the vehicles are similarly categorized into We classify the S types of the tasks and define the tasks as The RVEC comprises computation and communication modes. We discuss each mode in detail, which are as follows. Display style equations should be numbered consecutively, using Arabic numbers in parentheses.The task computation mode selection depends on the processing time requirement and its impending cost. Each vehicle can choose to accomplish its computation task either locally on its vehicular terminal, on another vehicle in a relay, or a VEC server. The task-completion mode selection depends on the processing time requirement and the corresponding cost.C0 represents the local vehicle resources. The vehicles with type-i tasks are categorized as Here, we assume that all vehicles have homogeneous resources for computation. By satisfying We denote the relay vehicle computation resources as Moreover, if When portions of the vehicles decide to select VEC computing mode, they offload VEC servers’ tasks. Let location .The integral part of the offloading process’s delay is the time consumption of file transmission from the vehicles to the VEC servers. Furthermore, to transmit data, the distant VEC server implies increasingly wireless transmission hops and greater time latency. We denote The operators always provide different services, i.e., maintenance of VEC servers, computation, offloading, etc., by charging some fees against those facilities. Where the computing cost for a type-i task on the VEC servers is defined as We compare V2I transmission with our proposed relay transmission approach concerning time consumption and cost to validate our approach. This work’s communication scheme is used to generate a realistic vehicular environment to study the task offloading problem. Moreover,we examine practical assumptions and measure the transmission rates for relay transmission and V2I transmission according to our previous work .We consider a scenario in which a type-i vehicle travels on a road segment Let the time delay and the cost for transferring the type-i task output through one road segment are expressed by As the vehicles’ speed is high and tasks take more time in their execution, the vehicle gets out of the VEC server’s proximity that executes the task. A longer delay might incur additional cost and transfer process.In such cases, when a vehicle offloads its task, it is first transmitted through the multi-hop V2V relay transmission. As a worst-case scenario, if no vehicles have enough computational resources in the relay, they keep on transferring the task to the next vehicle in a relay. Until the file is transferred to an RSU via V2I by the vehicle at the transmission relay’s destination hop. Therefore, the computation task is offloaded in advance of time to the VEC server in the vehicle’s headway compared to the vehicle’s current position. After the computation result/output reception, the VEC server saves the output to the RSU associated with it. Whenever the offloading vehicle enters into the communication range of RSU, it takes the output directly from the RSU.Let Contrary to this, if any vehicle in the relay has sufficient computation resources available, it will compute the task file while moving on the road. After completing the task, the vehicle sends the output to the relay’s desired vehicle and earns some reward for both computation and transmission. This process can reduce the overall cost and time to execute the task. The time consumption of the type-i task through relay vehicles will be defined as:Likewise, the cost would be defined as:By comparing The total offloading cost for our scheme is defined as:Thus, offloading cost minimization problem for the proposed relay task offloading scheme can be formulated as:Here, Here, by taking advantage of the neighboring vehicles’ communication and computational resources, we design an effective algorithm identified as a distributed RVEC Algorithm. The main objective of the distributed RVEC Algorithm is to minimize the network’s offloading cost.Moreover, Line 9–13 represent that if the time in relay computation is less than that of maximum delay tolerance, then the vehicle having sufficient computation resources computes the task and provides the output to the initial vehicle in the relay and gets the reward for providing its communication and computational resources from the service provider. The reward is received in terms of, i.e., electronic cash, free provision of network resources in the future, etc. On the other hand, Line 14–17 illustrates that if the vehicle is unable to be computed, it transmits the task to the next vehicle in the relay and earns its reward for only providing communication resources. Line 19–21 indicates that if none of the relaying vehicles can compute. Then the task will be transferred to the VEC server for computation. After the desire computation, the VEC server transfers back the initial vehicle results via the corresponding RSU.The flowchart of We demonstrate the performance results of our RVEC scheme in this section. We take five RSUs positioned alongside a unidirectional urban road. The vehicles are moving at a speed of 120 Km/h, and the vehicle’s density on the road is considered as λ = 0.3. The vehicle’s computational tasks are categorized into four types with the probabilities {0.01,0.2,0.3,0.4}. We figure out each task type’s computation resource as {9,18,26,33} units, respectively, other simulation parameter values are shown in V2I Direct scheme: In this scheme, the task is directly transmitted to the vehicle’s VEC server. Therefore, this scheme ignores utilizing the neighboring vehicles’ computation resources.PCMS Scheme: This scheme utilizes the vehicular communication resource while ignoring the vehicular computation resources. Thus, the task is only computed on VEC servers in a V2V relay manner.The proposed RVEC scheme is evaluated against the following benchmark schemes.and PCMS . It can On the other hand, in the case where the (In In For the reasons mentioned above, using the RVEC scheme will relieve the communication and computing burden of the VEC servers, thus avoiding network congestion. The higher priority task is more likely to prefer the RVEC scheme to reduce the offloading cost.In contempt of the rapid response rate, the VEC servers usually confront the resource limitation compared to the conventional core-cloud with a comparatively large computational capacity. The VEC servers associated with RSUs located at the road segments have to serve a high density of vehicles, leading to increased latency and network congestion. To avoid this situation, we designed a task offloading strategy while taking advantage of a vehicle’s transmission and underutilized-computational resources. Furthermore, we investigated the time consumption and the offloading cost of different transmission modes and proposed a promising relay task offloading mechanism in VEC. Also, numerical results reveal that our scheme significantly reduces the offloading cost, i.e., computation latency and communication resource utilization, and outperforms other benchmark schemes. In our future work, we will study how to shift VEC server burdens to the vehicular cloud through incorporating a machine learning algorithm. By considering the dynamic vehicular nature, diverse application needs, and application stringent latency constraints in more practical and real trace-based complex road scenarios in our future work.10.7717/peerj-cs.486/supp-1Supplemental Information 1Click here for additional data file.


Giardia duodenalis , is one of the most commonly-identified parasitic diseases worldwide. Chronic G. duodenalis infections cause a malabsorption syndrome that may lead to failure to thrive and/or stunted growth, especially in children in developing countries. Understanding the parasite/epithelial cell crosstalk at the mucosal surfaces of the small intestine during human giardiasis may provide novel insights into the mechanisms underlying the parasite-induced immunopathology and epithelial tissue damage, leading to malnutrition. Efforts to identify new targets for intervening in the development of intestinal immunopathology and the progression to malnutrition are critical. Translating these findings into a clinical setting will require analysis of these pathways in cells and tissues from humans and clinical trials could be devised to determine whether interfering with unwanted mucosal immune responses developed during human giardiasis provide better therapeutic benefits and clinical outcomes for G. duodenalis infections in humans.Human giardiasis, caused by the protozoan parasite  Giardia duodenalis , is one of the most prevalent enteric parasitic protozoan infections globally, with prevalence rates ranging from 2-5% in the developed world and 20-30% in the developing countries  of which only assemblages A and B are typically identified in both humans and in other mammalian hosts, whereas assemblage E, for example, is predominantly identified in the livestock  manifested extensive histological abnormalities as well as chronic inflammatory responses in intestinal and extra-intestinal organs (Tight junction (TJ) proteins represent major components of the intercellular adhesion molecules and regulate the permeability of epithelial  and endothelial barrier functions [for a review see ref ]. These l organs , the genl organs , 42.Escherichia coli  as well as Helicobacter pylori, secrete virulence factors that target TJ proteins in order to induce pathogenesis (Entamoeba histolytica cysteine protease A5 (EhCP-A5) elicited a pro-inflammatory profile, as characterized by increased expression of IFN-γ, TNF-α, and IL-13 that correlated with impaired expression of TJ proteins claudin-2, occludin, and ZO-1 , increased intestinal permeability, and reduced transepithelial electrical resistance (TEER) in both murine models of giardiasis as well as in humans  was strain-dependent and could be reversed by using caspase-3 inhibitors or the pre-treatment with the epidermal growth factor (EGF)  . Furtheror (EGF) . It has ardiasis . Notablylaudin-4 . HoweverG. duodenalis infection leads to loss of intestinal epithelial barrier integrity during human giardiasis as well as in murine models of human Giardia infection , capable of degrading multiple components of the host immune system  in IECs and are also capable of degrading chemokines expressed by parasitized IECs  as well as increased intestinal permeability observed during human giardiasis, especially in younger children, further investigations are warranted  staining assay , and its two major ultimate metabolites  by IECs represents another defensive mechanism employed against a wide range of lumen-dwelling enteric pathogens at the intestinal epithelium surface , 75. Giaparasite . Additiopoptosis , 75.G. duodenalis strains differ in their ability to induce pathological changes at the upper intestinal epithelial surface (tpi), glutamate dehydrogenase (gdh) and β-giardin (bg), commonly used to assign Giardia parasites to specific genotypes/assemblages are not associated with virulence   [for a review see ref ]. The AMin vitro .via the regulation of these peptide secretions through interaction with its receptor, IL-22R, which is solely expressed on non-hematopoietic cells, including epithelial cells in the intestine [for a review see ref  and trefoil factor 3 (TFF3)  in vitro , however, Giardia trophozoites become invasive and are spread into intestinal mucosa extending into submucosa layer as well as extra-intestinal sites  in greater numbers, accompanied by depleted GCs mucin stores as compared with their uninfected controls (Muc2-/-) showed significantly higher trophozoite burdens in the small intestine and had impaired weight gain as compared with control animals (Citrobacter rodentium-induced colitis (Meriones unguiculatus) receiving a high-fiber (20%) diet were more resistant to infection with G. duodenalis as compared with those gerbils maintained on a diet with low fiber (5%) contents  as well as in the lamina propria lymphocytes (LPLs) of the small intestine during an early phase of the parasite’s colonization  by these cells in vitro  in a primary mouse model of human infection , lacking both arms of the adaptive immunity  cleared G. duodenalis infection similar to their wild-type controls, whereas they did not exhibit defects in disaccharidase activity  from the Department of Biomedical Sciences, School of Medicine and Health Sciences, University of North Dakota (to SS-M), a Dean’s Meritorious Pilot Grant, School of Medicine and Health Sciences, University of North Dakota (to SS-M), and by NIH/NIGMSP20GM113123 (to SS-M).The author declares 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.


Crispness is among the most important food textures that contribute significantly to palatability. This study investigated the association between the perceived crispness and palatability of five types of Japanese rice crackers known as “kakinotane.” Two experiments were conducted using the temporal dominance of sensations (TDS) and temporal drivers of liking (TDL) methods. As descriptors for the TDS evaluation, we used 10 Japanese onomatopoeias to indicate various attributes of crispness. We also measured the mastication sounds and electromyography (EMG) activity during mastication. Principal component analysis data revealed that principal component 1, representing moisture characteristics, contributed more than 60% in both experiments. The palatability of the stimulus, which was described as having a very soft, moist, and sticky texture, BETA-BETA, was significantly lower than the others. However, there was no significant relationship between the amplitude of mastication sound or EMG activity and palatability. We demonstrated that naïve university students can discriminate the fine nuances of the crispness of “kakinotane” using the TDS and TDL methods. Our findings also suggested that the onomatopoeias used as descriptors in the TDS method had a greater influence on describing the nuances of food texture than the physiological data. Crispness is an important factor in food perception and evaluation and plays a key role in food palatability . Many prWhile the contribution of the physical properties of food to its crispness has been examined, few studies have examined the perception of crispness. Specifically, Japanese people prefer crispy foods and are more sensitive to different degrees of crispness than North Americans . This stA useful methodology for investigating the sensory characteristics of foods is the temporal dominance of sensations (TDS) method , which eMimetic words are an example of the sound symbolic aspect of language, where the acoustic representation of the word has a connection with its meaning. Moreover, mimetic words imitate sounds or other sensory experiences, rather than describe higher cognitive concepts. As texture is a sensory property, it can be described more effectively by sensory words such as onomatopoeias, rather than by semantic words . PreviouApproximately half of the 135 Japanese onomatopoeic words for texture have a recognition rate of at least 75%, and 66 out of these words were correctly recognized by over 90% of the nonprofessional population . As therAnother important reason for using onomatopoeic words in this study is their connection with the visual cortex area in the brain, whereby experiences are felt just by reading or hearing the word . BecauseAnother important method used in this study is the temporal drivers of liking (TDL) method, developed by Thomas et al. ; a TDS sDifferent psychological factors may impact our evaluation of foods. Interestingly, it has been suggested  that vocTwenty-one university students  participated in this study after providing written informed consent. All participants were native Japanese speakers. One participant was excluded from the analysis because of missing data.The sample size of the study was determined with the reference of those in the preceding studies ,22. All Five types of “kakinotane” were presented as samples labeled with the name S, N, K, Kr, and Kc. Three of the samples  were produced by representative brands of “kakinotane” and chosen based on varieties of textural characteristics by the sensory panel of the Japanese manufacturer. These samples were commercially available products produced by different manufactures . Sample A pilot study was performed to select the most appropriate onomatopoeic words for describing the crispness of “kakinotane.” Ten university students and researchers wrote down the onomatopoeias describing the crispness of each sample while eating it for 90 s.The 10 most frequently used words were selected and used as descriptors in the TDS session: BARI-BARI, BORI-BORI, GARI-GARI, ZAKU-ZAKU (for hard textures), PARI-PARI, PORI-PORI, KARI-KARI (for more crumbly textures), SAKU-SAKU, SARA-SARA (for light crispy textures), and BETA-BETA (wet and sticky texture). For a detailed explanation of the terms see The experiment was conducted using the sensory analysis software MagicSense 3.1.6  for Windows. The participants responded using a touchscreen-based personal computer. Responses were recorded at a sampling rate of 5 Hz with MagicSense.The experiment consisted of the TDS session and the liking session. Both sessions were conducted on the same day. The order was counterbalanced: half of the participants started with the TDS session and the other half with the liking session.In the TDS session, the participants were asked to keep chewing a sample for 60 s and evaluate the crispness using the given descriptors. Four types of samples were presented in paper cups containing five pieces of each product (approximately 2 g). Within 5 s of placing a sample in their mouth, the participants were asked to touch the button on the screen to start the trial. While masticating at a free pace, the participants were requested to select the most appropriate descriptor of the crispness from the 10 descriptors displayed on the screen. When their perception of crispness changed, they were asked to select another descriptor. The participants were asked not to choose any of the descriptors if they had already swallowed the sample or if none of the words described the perception. After each trial, a 1-min rest period was provided, during which, the participants rinsed their mouths with water.To measure the palatability of the samples, the liking session was conducted in accordance with the TDL method . In the The TDS data were analyzed using the MagicSense software to obtain the TDS band plots of the perceived crispness. Additionally, for the duration of each descriptor chosen for each sample in the TDS session, principal component analysis (PCA) was performed. One-way analysis of variance (ANOVA) was performed for the mean liking values over time in each trial. A post-hoc comparison was performed using Ryan’s method when a significant main effect was observed.t-test. When a CLWD score was significantly (α = 5%) higher or lower than 0, it indicated a positive or a negative driver of liking, respectively  (ps < 0.05 with Ryan’s test) and that the liking score for N was significantly lower than that for S (p < 0.05). = 0.63] b. A postps > 0.05) because of the small samples .The CLWD scores for the five samples are depicted in In the current experiment we measured the perceived crispness of five samples of “kakinotane” As previously confirmed by the TDS data of the preceding studies, hardness and crispness are dominant sensations at the beginning of mastication, but stickiness becomes more dominant at the end of the mastication period .Based on the obtained TDS data, PCA was performed. Our findings suggested that two PCs are mostly responsible for the crispness sensation: PC1 is the moisture characteristic of the sample, and PC2 is the hardness, with a high predominance of PC1. As moisture, expressed by BETA-BETA, is the main factor in crispness, the CLWD score revealed that palatability could be expressed negatively using only one descriptor. The remaining descriptors were not effective reflections of palatability.Consistent with previous findings ,4, moistExperiment 1 had inherent methodological flaws. First, the duration of the trials in the TDS and liking sessions were different because the liking sessions were completed when the participants no longer perceived crispness or after swallowing the sample. In contrast, the duration of the TDS sessions was fixed at 60 s. The CLWD is the mean liking score weighted by the dominant duration of each descriptor. In the calculation, the durations of the sessions were averaged. Therefore, differences in the duration of both sessions might have affected the results. To solve this issue, in the next experiment, we investigated the temporal dynamics of crispness evaluation in a shorter time window of 30 s. We set the pace of mastication to 1 Hz to control the speed with which crispness was changing.Second, in Experiment 1, we considered only the subjective evaluation of crispness, and we did not investigate the physical and physiological properties of the crispness of “kakinotane.” As suggested by previous studies ,27, measTwenty university students  participated in the study after providing written informed consent. All the participants were native Japanese speakers, untrained on TDS. The participants did not include those from Experiment 1. The experimental protocols were approved by the ethics committee of Tohoku University (No. 2019-008).The food samples were identical to those used in Experiment 1. The experimental procedure was conducted with the same program and a personal computer. Mastication sounds were recorded using a handheld audio recorder  with a windproof bore close to the mouth and binaural microphones  inserted into the ears . Sound sThe experiment consisted of three sessions: the TDS, liking, and sound/EMG recording sessions. All the sessions were conducted in the above order in 1 day. In the TDS and liking sessions, the procedures were identical to Experiment 1 except for the pace of mastication and the duration of each trial. The participants were asked to masticate at 1 Hz in time with metronome clicks. The duration of each trial and the rest period were 30 s.In the sound/EMG recording session, the participants placed the binaural microphones into their ears and masticated the samples while keeping their mouths close to the audio recorder. Additionally, the bipolar electrodes for EMG recording were attached to the masseter muscles. The participants touched the button on the screen to begin the trial. After placing the sample into their mouths for 3 s, they were asked to masticate the sample for 30 s at 1 Hz monitored by a timer displayed on the computer screen. They began the next trial after rinsing their mouths during a 30-s rest period.The TDS and liking data were analyzed as in Experiment 1. Power spectrum analysis of the mastication sound data was performed using only the first 5 s of each trial as the amplitude of sounds decreased over time . The ampEMG data for each sample were analyzed using MATLAB and Signal Processing Toolbox . Data from the electrodes on the left and right masseter muscles were averaged and converted to the averaged rectified value by smoothing using a 5-Hz low-pass filter. Data from 10 participants were excluded from the EMG analysis because of a failure in collecting all data, as the electrodes were removed from some of the participants.TDS band plots for each of the five samples are depicted in PCA of the TDS data indicated that two PCs that had variance combined to a contribution rate of 98%. PC1 (86%) was the moisture characteristics of the sample (expressed by BETA-BETA–SAKU-SAKU), and PC2 (12%) was the feeling of the easiness of mastication (expressed by BARI-BARI–ZAKU-ZAKU) .F = 72.04, p < 0.05, p2η = 0.79] (ps < 0.05), and the liking score for N was significantly lower than that of S (p < 0.05).As in Experiment 1, liking scores for all the samples decreased with time a. A one- = 0.79] b. A postThe CLWD scores for all five samples are presented in F = 2.54, p < 0.05, p2η = 0.15]. A post-hoc multiple comparison test (Tukey’s test) revealed that the amplitude spectrum of Kc was significantly higher than that of Kr (ps < 0.05). The mean amplitude spectrum of the mastication sounds was positively correlated with the mean liking results .F = 0.81, p = 0.53, p2η = 0.083]. The mean peak voltage of the EMG was positively correlated with the mean liking results .Examples of EMG signals from a single participant are presented in The liking score in Experiment 2 was similar to that in Experiment 1. Thus, it appears that mastication speed and ingestion time do not affect the evaluation of palatability.The TDS band plot indicated that the participants mostly chose BETA-BETA from the beginning to the end in evaluating sample Kr. It was assumed that they used this descriptor in two meanings. At the beginning of the evaluation, they used it to indicate stickiness of the product surface, and later, they used it to indicate moisture.The PCA results of the TDS data indicated that the participants evaluated crispness according to the moisture characteristics in Experiment 2 as in Experiment 1. However, PC2 in Experiment 2 was not the hardness of the sample but rather the perception of the size of pieces during mastication. This result likely illustrates the effect of the time window difference between the two experiments. As the duration of the TDS session in Experiment 2 was only 30 s and all the samples remained hard during all trials, the participants could focus more on the size differences of the masticated samples. However, in Experiment 1, the participants mostly evaluated the hard-soft properties during the 60-s time frame, as the hardness decreased with mastication because of saturation with saliva.The difference between the results of Experiments 1 and 2 could be attributed to the metronome usage, which may have influenced participants’ attention during evaluation. However, previous experimental studies have indicated that the number of chews and the time from ingestion to the first swallow did not significantly change the crispness level ; thus, rSeveral previous studies have demonstrated that jaw movements ,32 and mHowever, this study failed to detect other significant effects of mastication sounds and the EMG activities of the masseter muscles on crispness. Inconsistencies between the present and previous studies are likely a result of differences in the mastication action, whereby, previous studies ,27,36 haFrom the results of Experiment 2, we can suggest that in the case of “kakinotane,” onomatopoeic expressions were more positively correlated with crispness and palatability than the acoustic measurements or EMG activity of mastication. It is hypothesized that these results were caused by the impact of onomatopoeic words describing crispness that exceeded the effect of mastication sounds or EMG activities of the masseter muscles. Similarly, previous research indicated that onomatopoeias activate visual cortical areas in the brain  and creaThis study revealed that naïve Japanese university students discriminated between similar samples of “kakinotane” based on their crispness. Moreover, they evaluated their palatability differently, in consistence with previous experimental studies demonstrating that untrained volunteers could successfully distinguish between the different varieties of apples with scores similar to the trained panel . Thus, tThis study also revealed that the palatability of “kakinotane” depended mainly on its crispness, especially on the moisture characteristics. Moreover, onomatopoeias, describing the moisture characteristics, were the key descriptors of the crispness of “kakinotane” because PC1 in both experiments was described by the words BETA-BETA and SAKU-SAKU (“very moist”–“not moist”). Consistent with this observation, two other studies have reported that the moisture characteristics of snacks significantly affected their crispness ,4. HowevThe results of the LWD scores in both experiments revealed that BETA-BETA was a negative factor of crispness. BETA-BETA describes the moisture characteristics and appeared in the latter half of the TDS session in all samples. As “kakinotane” is famous for its crispness, BETA-BETA describes a quality that is opposite to its characteristic crispness. It should be noted that Japanese people like the stickiness of cooked rice and have a large glossary of terms describing it . HoweverWhile common negative factors are easily identified, the results of the LWD scores in the two experiments failed to identify common positive factors. This can likely be explained by the differences in the proposed conditions, such as the trial durations or the fixed mastication pace. “kakinotane” is a rice cracker with a porous structure that readily absorbs moisture. Thus, the number of masticatory strokes and the time required for a sample to be saturated with saliva are critical factors in determining the crispness of “kakinotane.” Therefore, the most reasonable time window for the TDL analysis to evaluate the relationship between palatability and crispness should be investigated.Another important factor of the current investigation was the acoustic measurements of mastication to elucidate the physical indices of crispness; however, no significant differences were observed. These results contradicted the findings of the review by Vickers, which revealed correlations between acoustic factors and tactile cues of crispness  and suggPrevious studies ,43 have While no specific differences in sound measurements were observed, the participants could identify and evaluate different crispness sensations based on very similar sensation stimuli. Thus, the perceived difference in crispness, as the perceived palatability, was mainly associated with the presented descriptors; therefore, semantic expressions may broaden our sensation sphere. As it has previously been suggested, words, such as “crispy” and “crunchy,” are not descriptive enough to describe a complex of sensory sensations and properties of food comprising sight, flavor, taste, texture, and sound. Moreover, a more peculiar sensory vocabulary would aid in distinguishing and investigate crispness levels in more detail ,46.In this study, the participants successfully evaluated palatability by using certain onomatopoeic words; therefore, these words can be an effective instrument for exploring the influence of vocabulary on perception. It has been suggested that various words are required to describe those nuances because of different textures in Japanese food ; as it iRecently, many studies have demonstrated that perceptual experience is significantly affected by the linguistic background. For example, Winawer et al. revealed that language affects color discrimination . ExperimAn additional, important limitation should be mentioned. This study was conducted with Japanese participants and used specific Japanese verbal expressions. To validate our hypothesis, further investigations focusing on additional connections between language and perceptual experience, are required. Thus, a range of consequent experiments is being prepared to explore the understanding of Japanese onomatopoeias to describe food texture by non-native Japanese speakers with different linguistic backgrounds and to measure the impact and use of a particular onomatopoeic expression on the perception of food. The results of this study can be sufficiently used as a part of a methodological base for education and an explanation of linguistic and semantic nuances of Japanese onomatopoeic words to non-Japanese speakers in future research. Similar experimental investigations, which were conducted among non-Japanese speakers using the onomatopoeias responsible for various nuances of pain, confirmed that these words were interpreted similarly in many dimensions without knowledge of the Japanese language  and the While a big number of mimetic words is a specific feature of the Japanese language and their usage can be acknowledged as a limitation of this study, continuing research on sound symbolism may highlight the universal character of such words and their ability to express tiny perceptional nuances. Thus, English onomatopoeic words were used in a range of preceding studies, showing that there was a considerable effect of mimetic words usage on consumer judgements  of a cerThere is also strong evidence that English onomatopoeic words follow common patterns in sound and meaning with Arabic  and PersHanada reported that semantic images for food perception occurred in participants when they were presented with pictures of food without the real sensation of the food stimuli . Our stuAnother important hypothesis that requires further research is that the knowledge of a certain vocabulary can stimulate the level of perception ,17. FurtTo our knowledge, this study is the first to evaluate crispness by the TDS and TDL methods using Japanese onomatopoeic words as descriptors. While it has been demonstrated that onomatopoeias can enrich the description of a sensory experience and have a significant influence on gustation, their use in experimental evaluation remains insufficient. Our results indicated that Japanese onomatopoeias are associated with various crispness sensations and would help native speakers to discriminate between similar kinds of foods. PCA revealed that onomatopoeias for moisture characteristics are key descriptors of perceived crispness. According to our results, several words, such as SAKU-SAKU, provide pleasant associations and are responsible for positive evaluations of snacks. Conversely, other words, such as BETA-BETA, are associated with overly sticky and moist textures and, thus, provided an unpleasant image and negative evaluation of palatability. We observed that several onomatopoeias were more positively correlated with palatability and likeness scores compared to objective measures, such as loudness and mastication, and a predictive framework for evaluating palatability based on onomatopoeias was provided.As this study was devoted to the nuances of the same product and specific Japanese expressions were used in the experiments, we recognize the limitations of the research. Especially, it remains unclear whether the association between described patterns of verbal expressions for sensory characteristics is present among non-Japanese participants. Further studies are required to investigate the impact of onomatopoeic words on the perception of foods on a broader level.


The determination of precise displacement vector fields is an important tool for deformation analyses in in-situ X-ray micro-tomography time series. The goal of the work presented in this publication is the development and validation of an optimized algorithm for 3D least-squares matching saving computation time and memory. 3D least-squares matching is a gradient-based method to determine geometric  transformation parameters between consecutive cuboids in voxel data. These parameters are obtained by an iterative Gauss-Markov process. Herein, the most crucial point concerning computation time is the calculation of the normal equations using matrix multiplications. In the paper at hand, a direct normal equation computation approach is proposed, minimizing the number of computation steps. A theoretical comparison shows, that the number of multiplications is reduced by 28% and the number of additions by 17%. In a practical test, the computation time of the 3D least-squares matching algorithm was proven to be reduced by 27%. Volume data analysis in a voxel space representation is a logical extension of 2D image data processing. In the field of Photogrammetry and Computer Vision, image correlation techniques are often used for fine measurement of corresponding points in image sequences or stereo image pairs. Early contributions applied cross-correlation to image data . In the It is also possible to use more complex geometric models for the perspective transformation . Grün anMaas et al.  transferIn the last years, also free DVC code was provided. For instance, TomoWarp2 ensures the computation of 3D displacement fields using 3D cross-correlation . They alThe least-squares matching algorithm contains an adjustment procedure where normal equations are computed. To speed up the process, Sutton et al.  showed aThe publication at hand is based on the work of Maas et al.  and LiebIn the following, some formulae of Liebold et al.  are reusSimilar to The range of the cuboid in the reference volume is:Equation  shows thBetween the reference and the deformed state, a coordinate transformation is performed. Similar to  in 2D anThe reduced reference coordinates are computed as follows:Thus, the vector of unknowns is: The parameter vector The first terms of the Taylor series expansion are used to linearize the observation equation, see Equation  where inThe volume gradients of the gray values are written with their short forms in Equation . The fulThere are different ways to obtain these gradients: One possibility is the computation of central differences Equation ) at inteAnother approach is the use of tri-cubic spline interpolation to compute the derivatives  vT·The solution of this problem can be obtained using the normal equations:As the computation of the normal matrix After setting up the matrices Equation  is solveThe steps of the interpolation of the gray values and their derivatives as well as the computation of After the iterative adjustment routine, the residuals are determined by rearranging Equation . The graEquation  with thes0 = standard deviation of the unit weightn = number of voxel of the cuboidn = number of unknownsThe standard deviation of the unit weight is:The model in Equation  is not lOften, mainly translations and only small rotations occur between different epochs of measurements. In these cases, 3D cross-correlation can be used to obtain initial shifts . An overAs mentioned above, the normal matrix (Equation ) as wellEquation  shows thEquation , the rigIn fact, the geometric and radiometric parameters are usually independent on each other and are therefore often solved independently in LSM. If only the affine parameters are unknowns and the radiometric parameters are excluded from the model, the last two columns and last two rows of For each voxel, the following terms are computed and saved in temporal variables because they are used several times: Substituting the terms of Equation , the subThe computational costs are compared between the direct computation and the standard way that includes building up A total of 100 terms have to be computed to form the normal equations directly, see Equation . This reMultiplications per voxel:−Nine multiplications in terms of Equation .−86 multiplications in the 100 terms .Further multiplications:−Computation of the product −84 multiplications of the prefactors with the sums in Equation  for the −40 multiplications for the right hand side (Equation ) outsideFurther additions:−28 additions for the right hand side (Equation ).To evaluate the computational effort, the number of additions and multiplications are counted:Altogether, for the direct computation of The computational effort of the standard method contains building up the Jacobian matrix The following terms are calculated and saved in temporal variables for each line of the matrix For the each line of matrix For each element of Multiplications per voxel:−One multiplication for building up the reduced observation vector −Three multiplications for the terms of Equation  that are−Nine further multiplications for building up the Jacobian matrix of Equation .−105 multiplications (105 elements) due to the matrix multiplication of −14 multiplications (14 elements) due to the matrix-vector multiplication of Additions per voxel:−Two additions for building up the reduced observation vector −105 additions (105 elements) due to the matrix multiplication of −14 additions (14 elements) due to the matrix-vector multiplication of Altogether, for building up https://gcc.gnu.org) and compiled with the optimization option -O3. The loop over the matching points with the computation of the 3D LSM is parallelized with the OpenMP library , and the time measurement only considers this loop. The time measurement includes the whole 3D LSM process so that not only the calculation of the normal matrix The comparison was done on a desktop machine with a AMD Ryzen 7 3800X 8-Cores (16 threads) processor at 3.9 GHz. In the test, 3D LSM was applied to 146,933 cuboids defined in the first two data sets of an in-situ tension test of Lorenzoni et al. . The pathttps://eigen.tuxfamily.org/). For M3, the full matrix of Different calculation modes are tested: Method M1 stands for the direct computation. M2 represents the standard computation with building up the Jacobian matrix The paper presents an optimized algorithm to compute 3D least-squares matching in voxel data sequences including geometric and radiometric parameters. Compared to the standard method of the estimation of the normal equations, the number of multiplications is reduced by 28% and the number of additions is reduced by 17%. In a practical test, the computation time was decreased by 27%.


The multivariate analysis showed an independent significant association of OSAHS with SVD (DWM and PVWM)  and: p = 0.002, OR 95% CI: 104.98 (5.15–2141)). The same analysis showed a moderate association of OSAHS with B-WMH ). Our study demonstrated an independent significant association of OSAHS with SVD and a moderate association of OSAHS with B-WMH.Obstructive sleep apnea hypopnea syndrome (OSAHS) is associated with increased risk of cerebrovascular disease. The aim of the present study was to investigate the association between the presence of the small vessel disease (SVD) of the brain in patients with OSAHS. The study included 24 patients with moderate to severe OSAHS and 34 healthy volunteers. All the subjects underwent magnetic resonance imaging (MRI) of the brain, in order to sought periventricular white matter (PVWM), deep white matter (DWM) and brainstem SVD. Among patients with OSAHS, 79.1% had SVD  in DWM and 91.7% in PVWM while 22.4% had brainstem—white matter hyperintensities (B-WMH). Patients with OSAHS had a much higher degree of SVD in the DWM and PVWM compared to the control group ( Obstructive sleep apnea hypopnea syndrome (OSAHS) affects 5–10% of the population. The severity of the syndrome in determined by the number of apneas and hypopneas taking place during an hour of sleep (apnea-hypopnea index—AHI) as well as by the seriousness of the patient’s symptoms at daytime . The patOSAHS impact on patients with ischemic stroke (IS) or transient ischemic attack is quite higher when compared to the general public. It also seems that OSAHS constitutes a factor that influences the course of an IS and the patient’s functionality, after the stroke ,4. ModerSmall vessel disease (SVD) of the brain result from the damage taking place on penetrating cerebral microvessels, capillaries and veins, that damage the underlying tissue, the white and grey matter of the brain. Penetrating vessels play a crucial role in retaining the functionality of the brain and especially of nuclei with high metabolic needs, as well as the complex networks of the white matter . The detThe objective of the present study was to investigate the association between the presence of SVD in patients with OSAHS in a Greek population. Furthermore, we aim to evaluate the impact of OSAHS in brainstem—white matter hyperintensities (B-WMH) since little is known regarding B-WMH topography.A total of 109 adults were approached at their appointment at the Department of Respiratory Sleep Disorders of the University of Thessaly (Greece) during the period from 4 November 2019 to 31 March 2021 . All patPatients and controls provided written informed consent. The study was approved by the Independent Ethical Committee of the University of Thessaly, Medical School (No. 13—12/02/2019).The participants of the study underwent polysomnography. During this procedure, electroencephalography, electrooculography, electromyography, electrocardiography and oronasalthermistor were used. Bonds were attached to the thoracic cage and the abdomen in order to monitor the respiratory motions. Oxygen was monitored with a pulse oximeter. Apnea was defined as a ≥90% reduction in airflow lasting at least 10 s. Hypopnea was defined as a ≥30% reduction in airflow lasting at least 10 s and at least a 3% drop  in oxygen saturation. The AHI score was calculated as the average number of apneas and hypopneas per hour of sleep. According to the AASM [MRI was performed in all participants using a standard protocol. A 3-T Magnetic Resonance Scanner  was used and the protocol included whole-brain T2-weighted, T1-weighted, and T2*-weighted gradient-recalled echo, FLAIR, as well as diffusion sequences. The assessment of the MRI images was performed by a neuroradiologist that was blinded to the participants’ clinical data.White matter lesions were defined as follows: Any signal abnormality despite of size that is located in the white matter displaying hyperintensity on T2-weighted images like fluid-attenuated inversion recovery, that does not present cavitation .We used the Fazekas score (FS) to evaluate WMH on axial T2-weighted and FLAIR sequences . PatientAdditionally, we assessed for B-WMH. We defined B-WMH as areas of hyperintensity without the presence of distinct borders on FLAIR images, presenting with minor or absent corresponding hypointensity on T1 -weighted images in mesencephalon, pons, and/or medulla. The lesions were classified by a neuro -radiologist (SM) with a dichotomized classification scheme as absent or present. Lesions with clear demarcated hyperintensity located in the brainstem that were associated with a corresponding DWI hyperintensity or clear T1-weighted hypointensity were considered infarcts and were not included in the analysis. Since the WMH areas located in the brainstem are too small, we cannot adequately discriminate B-WMH according to volume by the semiautomated volumetric image analysis as we performed to PV (periventricular) and D (deep) WMH. The clinical data were not available to the readers of the MRI images. We used the Microbleed Anatomical Rating Scale to identify deep microbleeds .t-Test was performed for continuous data since there were no deviation from normal distribution  and violation of assumption of homogeneity of variance (Levene’s test). Categorical variables were compared using the χ2test.Multivariate analysis was performed in the form of binary and multinomial logistic regression. In multivariate analysis, factors with a p value < 0.10 in univariate analysis included. For all the analyses a 5% significance level was set. Analyses were performed using SPSS version 22 for Windows® .Continuous variables are expressed as means ± standard deviations and categorical variables as frequencies and proportions. Student’s 2 level was 76.5 ± 12.8%. Sleep parameters and characteristics of OSAHS patients are presented in The patients with OSAHS consisted of 24 patients  with a mean age of 58 ± 11.9 years. Of the 24 patients with OSAHS, 16 had severe syndrome and 8 had moderate. The mean AHI was 40.4 ± 21.2 events/h. Mean apnea index (AI) was 16.3 ± 17.3 events/h and mean hypopnea index (HI) was 24.1 ± 13.4 events/h. Oxygen desaturation index (ODI) was 41.2 ± 23.2 events/h, time with hemoglobin saturation < 90% (T < 90%) was 36.8 ± 65.6 min and that of the minimum SaOp < 0.001) (p < 0.001) (OR: 33 (3.87–281.70))  in DWM and 91.7% in PVWM while 22.4% had B-WMH. Patients with OSAHS had a much higher degree of SVD in the DWM, PVWM compared to the control group with statistically significant difference (< 0.001) . When co281.70)) .p = 0.097), coronary artery disease (p < 0.001), hypertension (p < 0.001) and diabetes (p = 0.039). The duration of the disorder (OSAHS) is also an important factor that affects the presence of SVD (p < 0.001)  ).p = 0.035), coronary artery disease (p = 0.017) and hypertension (p = 0.002). The duration of the disorder (OSAHS) is also an important factor that affects the presence of SVD (p < 0.001)  .p = 0.033, OR 95% CI: 8.66 (1.19–63.08) and p = 0.002, OR 95% CI: 104.98 (5.15–2141), respectively) )  (ctively) . The sam234.65)) .Lobar microbleeds were not associated with OSAHS. We did not find any other significant association between several sleep parameters and OSAHS characteristics with SVD.To our knowledge, there are no studies in the Greek population that describe the neuroimaging characteristics of SVD associated with OSAHS. SVD, which is generally asymptomatic and commonly associated with cardiovascular risk factors, is a prevalent disease in older persons ,19,20. SThese findings agree with previous studies ,23,24 shWe observed that patients with OSAHS had increased prevalence of cardiovascular risk factors when compared with controls. Wang et al.  have preSimilar results seem to emerge from the study of Chokesuwattanaskul et al., who observed an association between OSAHS and CSVD MRI findings of WMH and asymptomatic lacunar infarction (ALI) when compared to patients without OSAHS . A recenThe association between OSAHS and SVD may be bidirectional . There aOur study is not without limitations. First, we acknowledge that we included a relatively small number of participants. Additionally, we did not assess intra- and interobserver reproducibility for brain imaging. Future longitudinal studies including larger cohort, may allow us to better clarify the potential association and causality—between SVD and OSAHS, preferably by including both histopathology samples and biomarkers. We acknowledge that the use of visual scales is useful when one aims to assess SVD on MRI. Early identification of SVD lesions could permit the application of possible preventive measures. Thus, we believe it is mandatory that all specialists that are involved in the care of OSAHS should be able to identify and classify SVD quantitatively.In conclusion, our study demonstrated an independent significant association of OSAHS with SVD. In addition, the results confirmed for the first time the moderate association impact of OSAHS on B-WMH.


Thymic stromal lymphopoietin (TSLP) is an innate cytokine, belonging to the group of alarmins, which plays a key pathogenic role in asthma by acting as an upstream activator of cellular and molecular pathways leading to type 2 (T2-high) airway inflammation. Released from airway epithelial cells upon tissue damage induced by several noxious agents including allergens, viruses, bacteria, and airborne pollutants, TSLP activates dendritic cells and group 2 innate lymphoid cells involved in the pathobiology of T2-high asthma. Tezepelumab is a fully human monoclonal antibody that binds to TSLP, thereby preventing its interaction with the TSLP receptor complex. Preliminary results of randomized clinical trials suggest that tezepelumab is characterized by a good safety and efficacy profile in patients with severe, uncontrolled asthma. Asthma is a chronic obstructive respiratory disease, mainly characterized by airflow limitation due to bronchial inflammation and airway remodeling ,2. A halIn addition to chronic inflammation, all asthma endotypes are often characterized by airway structural changes, which span throughout the various layers of the bronchial wall . These rIn T2-high asthma, the development, persistence, and amplification of eosinophilic inflammation are driven and orchestrated by multiple cellular elements including dendritic cells, T helper 2 (Th2) lymphocytes, group 2 innate lymphoid cells (ILC2), mast cells, basophils, and airway epithelial cells ,13,14. W2-adrenergic agonists (LABA) within ICS/LABA fixed combinations [https://clinicaltrials.gov/ website. Poster presentation at the American Academy of Allergy Asthma & Immunology, 26 February–1 March 2021).Most patients with mild or moderate asthma are well controlled by inhaled corticosteroids (ICS), eventually integrated by the addition of long-acting βinations . Furtherinations ,20. Despinations ,24,25,26In addition to the already approved, above-mentioned anti-asthma biologics, other monoclonal antibodies are currently undergoing clinical investigation, which might potentially guarantee a wider coverage of therapeutic advantages . Among tIn light of the above considerations, this article aims to review the role of TSLP in asthma pathophysiology, as well as to discuss the therapeutic properties of tezepelumab as an eventual add-on treatment option for severe asthma.Originally identified in thymic stromal cells, TSLP is an innate pleiotropic cytokine belonging to the four-helix-bundle cytokine family and is distantly related to interleukin-7 (IL-7) . Two TSLTSLP belongs to the family of alarmins. Indeed, this innate cytokine is mostly released by airway epithelial cells as an alarm signal when tissue injury is triggered by several environmental insults such as allergens, cigarette smoke, airborne pollutants, viruses, bacteria, and chemical and physical irritants ,34. GenoOnce secreted from damaged bronchial epithelium, TSLP affects the functions of several immune/inflammatory and structural cells, which co-express TSLPR and IL-7Rα. In particular, ILC2 is included among the most relevant cellular targets of TSLP. Together with other alarmins such as interleukin-25 (IL-25) and interleukin-33 (IL-33), TSLP prolongs ILC2 survival and stimulates these cells to produce large quantities of interleukin-5 (IL-5), interleukin-9 (IL-9), and interleukin-13 (IL-13)  13,15,4,41. IL-5In addition to ILC2, within the context of innate immunity, TSLP also regulates the functions of both eosinophils and eosinophil progenitors, which play a key role in type 2 airway inflammation. In particular, both TSLPR and IL-7Rα are expressed by human eosinophils susceptible to the activating and anti-apoptotic effects of TSLP, which enhances eosinophil survival and potentiates the release of eosinophil-derived chemokines and cytotoxic proteins ,47. Thes2 (PGD2), a pleiotropic eicosanoid that exerts a wide spectrum of pathogenic actions in type 2 asthma [Other cellular targets of TSLP are basophils, whose TSLPR expression can be stimulated by allergens via IgE-dependent mechanisms ,52. TSLP2 asthma ,55. More2 asthma ,57.+ T lymphocytes into mature Th2 cells [+ T lymphocytes, IL-4 biosynthesis, and Th2 cell differentiation [+ monocytes/interstitial macrophages [+ and CD8+ T lymphocytes. Indeed, TSLP can directly induce the commitment of naïve CD4+ T cells towards the Th2 lineage, as well as the expansion of CD8+ T lymphocytes [In regard to the crosstalk between innate and adaptive immunity underlying the cellular pathophysiology of asthma, TSLP exerts a pivotal function by acting at the level of dendritic cells. Indeed, human myeloid dendritic cells express TSLPR, whose activation by airway epithelium-derived TSLP leads to up-regulation of major histocompatibility complex class II (MHC-II) and several co-stimulatory molecules such as CD40 and CD86 . Moreoveh2 cells . Therefoh2 cells . When puh2 cells . Interesntiation . In allentiation . A furthntiation . Indepenrophages . Althougphocytes ,69. In pphocytes . In partphocytes .+ T cells into Th17 lymphocytes [In addition to being implicated in the pathobiology of type 2 airway inflammation, TSLP appears to be involved in T2-low neutrophilic asthma, characterized by a pivotal pathogenic role played by Th17 lymphocytes . In thisphocytes ,73. It hIn addition to significantly affecting the functions of immune/inflammatory cells, TSLP extends its biologic actions to airway structural cells. In particular, airway smooth muscle cells express TSLPR and respond to TSLP by increasing their production of IL-6 and IL-8 ,75. Airw1), respectively, with respect to allergen-induced FEV1 decreases detected on the same days [1 decrement (methacholine PC20). Moreover, significant decreases in blood/sputum eosinophils, as well as in fractional exhaled nitric oxide (FeNO) levels, were detected in patients treated with tezepelumab [Tezepelumab is a fully human monoclonal IgG2λ antibody that specifically ligates TSLP at the level of its binding site for TSLPR, thereby impeding the human TSLP–TSLPR interaction . Initialame days . Tezepelepelumab . This drepelumab .1 values ranged from 40% to 80%, and a positive response to the reversibility test was shown by a post-bronchodilator FEV1 increase of at least 12% and 200 mL with respect to baseline measures. Subjects with clinically relevant respiratory diseases other than asthma were excluded from the study. Among 918 screened patients, 584 were randomly subdivided into four groups including a placebo arm of 148 subjects, and three subgroups of patients receiving subcutaneous injections of tezepelumab every 4 weeks, at dosages of 70 mg (low dose: 145 patients), 210 mg (medium dose: 145 patients), or 280 mg (high dose: 146 patients).Subsequently, the randomized, double-blind, placebo-controlled, multicenter PATHWAY phase 2b trial was carried out between December 2013 and March 2017 at 108 sites throughout 12 countries . Enrollep < 0.001) decreased AAER by 61%, 71%, and 66% at the dosages of 70, 210, and 280 mg, respectively [1 by 120, 110, and 150 mL in the low-dose, medium-dose, and high-dose groups, respectively [The primary goal of the study was to assess at week 52 the effects of tezepelumab on the annualized asthma exacerbation rate (AAER). In comparison to placebo, tezepelumab significantly  and adolescents (12–17 years) with severe asthma not adequately controlled by medium-to-high dosages of ICS, associated with at least another controller drug . The stuSOURCE is another multicenter, 48-week, double-blind, randomized and placebo-controlled trial, enrolling 150 severe asthmatic patients on treatment with medium-to-high doses of ICS/LABA combinations, associated with an additional chronic OCS therapy . The priOutside of the phase 3 PATHFINDER program, which includes both NAVIGATOR and SOURCE studies, DESTINATION is a further phase 3, 1-year trial recruiting 960 patients who have already completed one of the above two studies . TherefoFinally, CASCADE is a phase 2, double-blind, randomized and placebo-controlled study, focusing on the anti-inflammatory activity of tezepelumab . Here, 1TSLP represents one of the most suitable therapeutic targets for emerging biological treatments of severe asthma ,90,91. I