Human papillomavirus (HPV) infection is thought to be strongly associated with the precarcinomatous state cervical intraepithelial neoplasia (CIN) and cervical carcinoma. To accurately assess the correlation between HPV detection profiles and CIN, the uniplex E6/E7 polymerase chain reaction (PCR) method was used. We detected HPV (37 genotypes) in 267 CIN cases. The detection of a single high-risk HPV genotype occurred in 69.7% of CIN1 and worse than CIN1 (CIN1+) cases whereas other types were detected in 11.6% of cases. Codetection of high-risk HPV genotypes occurred in 4.9% of CIN1+ cases. The high-risk genotype HPV16 was the most frequently detected genotype in CIN1+ lesions; the genotype HPV34 (not a high-risk type) was detected in some CIN3 cases. Furthermore, HPV codetection may not be associated with CIN grades. These results suggest that various HPV genotypes are associated with CIN across all analyzed cases. Human papillomavirus (HPV) is associated with various oncologic diseases, including benign tumors and carcinomas ,2. HPV mCurrently, HPV is phylogenetically classified into over 100 genotypes . Some, sSome previous studies have reported CIN/ICC cells in which no HPV could be detected ,16. ThisSpecimens were obtained from colposcopy-directed punch biopsies collected from 398 patients at the Genki Plaza Medical Center for Health Care, Tokyo, Japan, between 2014 and 2018. Colposcopy-directed punch biopsies contained cervical lesions (except for condyloma) from women with abnormal cytology findings. Cervical tissue biopsies were performed if subjects showed abnormal findings on cervical cytology after medical examination. Biopsy specimens were fixed in neutral buffered formalin and embedded in paraffin. Serial sections were cut from each formalin-fixed paraffin-embedded (FFPE) tissue block. The first section was stained with hematoxylin-eosin for histopathological evaluation; additional sections were used for the preparation of DNA and HPV genotyping. Biopsies were grouped according to morphological diagnosis. All case diagnoses were reviewed and reconfirmed by one pathologist. The mean age of the patients was 39.1 years . No data regarding HPV vaccination history was collected from patients.DNA isolation from CIN1+ FFPE tissue was performed using the hot sodium hydroxide and Tris (HotSHOT) method . BrieflySamples were collected after written informed consent was obtained from subjects. The study protocol was approved by the Ethics Committee on Human Research of Kyorin University (H28–27). The protocol was implemented in accordance with approved guidelines.® 360 buffer, 2 mM MgCl2, 0.025 U/µL AmpliTaq Gold 360 DNA Polymerase , 1 μL DNA, and 0.5 pM primers in a total volume of 25 μL. PCR amplification was performed using a thermal cycler, with 35 cycles of denaturation at 95 °C (30 s), annealing at 60 °C (30 s), and extension at 72 °C (30 s), including an initial denaturation step of 10 min and a final extension step of 5 min. As the PCR method occasionally provides false-positive results, each PCR experiment included DNase-free water as a negative control to eliminate the possibility of DNA contamination. HPV types were classified into two groups in the present study: high-risk genotypes [A new HPV-typing assay method, uniplex E6/E7 PCR, previously established by Okodo et al.  was usedenotypes  . Differences between groups were examined using residual analysis, based on results of Chi-squared test and one-way analysis of variance and according to the characteristics of the data distribution. In all cases, a p = 0.049) and CIN3 (p = 0.007). In 89.3% (267/299) of CIN1+ tissues, amplifiable quality DNA was obtained based on β-globin positive results. Uniplex E6/E7 PCR detected evidence of HPV in 100% of CIN1+ samples. In total, of 39 possible HPV genotypes, 37 genotypes were detected, with only HPV26 and HPV85 not being detected. The amplicon sequence for each HPV genotype was matched completely with the original sequence obtained from the GenBank database (data not shown).Pathological examination suggested that 75.1% (299/398) of cases were CIN positive. These were further classified as 132 CIN1 lesions, 82 CIN2 lesions, and 85 CIN3 and worse than CIN3 (CIN3+) lesions . Ninety-nine cases were benign lesions . Mean ages for the CIN1, CIN2, and CIN3+ groups were 36.6 years , 39.2 years , and 40.4 years , respectively. Patients with CIN1 were significantly younger than those with CIN2 (p < 0.05). In contrast, HPV39, HPV56, and HPV66 were at significantly higher frequency in CIN1 than CIN2 and CIN3 .We summarized the relationships between detected HPV genotypes, codetection rates, and CIN grade. The correlation between the distribution of high-risk and other HPV genotypes and CIN grade was analyzed . GenotypNext, a single high-risk HPV genotype was detected in 69.7% (186/267) of cases, whereas a single other genotype was detected in 11.6% (31/267) of cases . HPV16 wIn the present study, 37 of a possible 39 HPV genotypes were detected in 267 CIN cases, although notably the test did not cover all HPV types. HPV was detected in all present cases; however, HPV codetection may not be associated with CIN grade. The high-risk genotype HPV16 was the most frequently detected genotype in CIN1+ cases, while HPV34 (not a high-risk type) was significantly detected in CIN3. In general, HPV detection by the PCR method may be performed liquid-based cytology . HoweverMost previous reports have focused on correlations between high-risk HPV infection and cervical lesions ,18. For In the study by Xiao et al. , a greatp < 0.05). HPV34 is classified into the alpha-papillomavirus group 11, a group which may be highly oncogenic, although it is not as high risk as HPV16 [In the present study, HPV34 was shown to be significantly detected in CIN3+ cases (as HPV16 . Jacobs as HPV16  attempteas HPV16 . In the as HPV16 . To the The present study has certain limitations. We detected HPV genotypes from CIN+ samples using uniplex E6/E7 PCR, which is able to identify 39 HPV genotypes. However, the present study has not demonstrated any correlation between probable high-risk HPV and CIN grade. It is difficult to clearly define chronological changes in CIN because of the need for histology and HPV genotyping in the short-term. As another limitation, a limited number of cases were included in this study, with a smaller number of CIN2 and CIN3 cases compared to CIN1. However, although the number of cases was limited, HPV was detected in all CIN cases in this study. Therefore, extensive HPV typing is important to clarify the relationship between genotype, superinfection, and CIN. In order to investigate the progression of CIN to cancer, it is necessary to include a greater number of cases and analyze CIN2+ cases in greater detail. Moreover, we did not introduce the primary HPV screening in this study . HoweverIn conclusion, we demonstrated by a classical PCR assay that all CINs are caused by HPV with equal sensitivity using a type-specific primer targeting the E6 or E7 genes. To precisely assess the correlation between HPV infection profiles and cervical lesions, including CIN and ICC, the comprehensive detection of HPV genomes from lesions using such sensitive methods is essential. Since the number of cases is small in this study, further and larger studies are necessary to target 13 HPV genotypes, including HPV34, when introducing primary screening for cervical cancer by HPV testing in Japan.


Stomoxys calcitrans is considered as a major pest of livestock worldwide. Insecticides have been extensively used to control this pest but resistance to these chemical compounds is now reported in many countries. Therefore, a more sustainable and efficient control is needed. New blue polyethylene sticky screens have been proved to be very attractant for stable flies. They are produced at a lower price than all blue fabric screens or traps. More than 70% of flies are captured on the lower half (30 to 60 cm above ground) of the screens. In our conditions, very few non-target fauna was captured as very few pollinators were caught by these screens. These results are highlighting the interest of these blue polyethylene screens to control stable flies in cattle farms, in comparison with more expensive blue fabrics.The haematophagous fly Stomoxys calcitrans is considered as a major pest of livestock worldwide. Insecticides have been extensively used to control this pest but resistance to these chemical compounds is now reported in many countries. Therefore, a more sustainable and efficient control is needed. Seven different types of blue screens, with reflectances around 460 nm, were tested during summer 2016 in southwestern France to evaluate their attractiveness and their specificity for stable flies. Height of the screen and orientation (east or west) of a blue screen were also considered. High levels of S. calcitrans captures were recorded during this study  whereas the numbers of tabanids and pollinator insects remained extremely low . No significant difference in attractiveness has been shown between the different types of blue screens. The lower half of the blue screens caught significantly more stable flies (70%) than the higher half (30%). The “east” side of the screen attracted 60% of stable flies but this was not significantly different from the west side. These results are highlighting the interest in these blue polyethylene screens for controlling stable flies in cattle farms, in comparison with more expensive blue fabrics. Stomoxys calcitrans  is a cosmopolitan pest of livestock, which compromises animal and human welfare. Males and females are harassing hematophagous insects, feeding preferentially on ungulates but also on dogs and humans, and cause painful bites  iS. calcitrans captured by blue screens in this study due to the very high number of flies caught and that were tightly adhering onto the sticky film (making them difficult to sample for observation under a binocular microscope). However, in a complementary study in 2017, we observed that the sex ratio was almost 1 to 1  under the same sampling conditions.Several studies have been conducted to understand the relationships between the height of the traps and the efficacy of catching of stable flies. Beresford and Sutcliffe (2008)  found thThe east and the west exposed sides of the blue screen caught slightly different numbers of stable flies without significant difference. The east side caught a little bit more than the west side and this was observed whatever the type of blue screen and the repetition. This might suggest once again that activity of stable flies was more important in the morning (when the east side of the blue screen is facing the sun) than in the afternoon during the period of both experiments. Our results also showed that the specificity of attractiveness was remarkably high as demonstrated by the low numbers of pollinators captured by the different types of blue screens. As sticky traps are expendable products and cannot be recycled or re-used, it is quite difficult to imagine the systematic association between blue screens and sticky films in a sustainable and cost-effective stable fly control program. Therefore, the impregnation of the blue screen with an insecticide in a combination of “specific attraction” and “contact mortality” could be an alternative solution to the topical application of such insecticides on cattle. Nevertheless, the landing time of stable flies on the blue screens and the mortality rates following short contacts between the screens and the flies should be carefully checked.S. calcitrans caught during both experiments whatever the type of blue screens or the repetition. In most instances, more than a thousand stable flies were caught by one blue screen in only 8 h. This is indicative of extremely high population numbers with a uniform distribution and the similarity in attraction and capture among the various traps.The most striking result of this study was the high numbers of The second important finding of this study was that the different devices tested in both experiments showed the same attractiveness for stable flies on the NVST campus. This similarity could have been influenced by the plastic sticky film placed on screen surfaces. But this suggests that polyethylene films could be used instead of fabrics for the control of hematophagous insects using insecticide-impregnated screens, consequently at a much lower price. Beside this, the results obtained in this study showed clearly that most stable flies tend to land on the lower half of the blue screen, i.e., between 30 and 60 cm height above the ground level.


Transient ground displacement (TGD) that is caused by earthquakes can damage underground pipes. This damage is especially critical for the joints, elbows and tees of the pipes which play an important role in the operation of a pipe network. In this study, a scale pipe network with both elbows and tees, as well as some components of the pipe network with only tees or elbows, has been investigated. The response of the nodes of a pipe network, when installed in non-uniform geology, was analyzed using the shaking table test and ABAQUS finite element simulation. This paper has firstly introduced the preparation of the test and the developed finite element model. Then the system response in terms of strain, the friction, the bending deformation, the node deformation amplification coefficient and the pipe-soil relative displacement along the pipe axis of the pipe network and two pipe network components have been analyzed explaining the correlation between these responses. Finally, the influence of elbows and tees on the pipe network was analyzed, and the conclusions that have been reached about how tees and elbows can change the response of a pipe network during an earthquake can provide theoretical support for the seismic design and layout of an underground pipe network. Transient ground displacement (TGD), which is caused by seismic waves, is a type of geological hazard that can be triggered by earthquakes. The damage to the joints of pipe networks and their components, such as elbows and tees, is the main form of damage that can occur as a result of an earthquake . Fig 1 dThe water supply and the gas supply network are an important part of the urban lifeline system. In many earthquake investigations, it was found that damage to an underground network may cut off the supply of drinking water, fire hydrants and gas, and can cause secondary disasters such as urban waterlogging and fire, resulting in a huge loss of life and property . In 1995At present, research on the underground pipe network system is limited to two aspects, involving: 1) component performance, for which the soil-structure interaction under earthquake loading is evaluated, and 2) system performance, for which the integrated behavior of the network is assessed. These two aspects are very different and are governed by issues that are related to the level of detail that can be achieved in the component vs. system characterization, as well as the spatial variability, uncertainties in the properties of the materials and the state of repair of the components, the network’s flow laws, and the operational logic of the system . For exaThe study into the mechanical research methods of an underground pipe during an earthquake includes numerical analysis, theoretical calculation and experimental research currently. For numerical analysis, most scholars use the finite element method –22. One Based on the above analysis, this paper has studied the seismic response of an underground pipe network by means of the shaking table test. The specific research process was as follows:Straight pipes, elbows and tees made from PVC-U (Unplasticized polyvinyl chloride) were used to make the scale pipe network as well as H-shaped and Z-shaped pipe network components. According to the research content, the corresponding sensors were laid. The H-shaped and Z-shaped pipe networks were put into the soil of the shaking table, and the same artificial waves were then applied;Based on the shaking table test, three-dimensional finite element models of scale pipe network, H-shaped and Z-shaped pipe were established;According to the axial strain of the scale pipe network in the direction of the seismic wave, the friction force, the bending deformation, the deformation amplification coefficient of the nodes and the pipe-soil relative displacement in the corresponding direction of the pipe were analyzed;Then the test results of the axial strain of the H-shaped and Z-shaped pipes were analyzed, and finally the seismic response relationship between the nodes of the buried pipe network was determined.3, a Poisson’s ratio of 0.3 and a buried depth of 0.3 m. The actual engineering buried depth of the network was 3.0 m through the similarity ratio conversion. By inserting and applying glue to the elbow of the pipes in order to simulate the welding of the iron pipe’s elbows and tees, the displacement of the pipe network’s nodes could be determined by the pulling distance of the elbow or tee. For the construction of the pipe network, the elbows and tees were combined to form a scale pipe network to study the axial response of the pipe. As there are both tees and elbows in the pipe network, a single type of pipe joint was established to study the seismic response. In this study, H-shaped and Z-shaped basic node forms were constructed to analyze the influence of tees and elbows on the pipe network during an earthquake. The pipe’s size and its position in the shear box are shown in PVC-U is a type of hard-plastic; based on the influence of the scale of the model, the seismic waves could not destroy the pipe since it was in its elastic stage. Therefore, the internal forces and the deformation tendency of the PVC-U pipes were the same as that of iron pipes. Therefore, because of their similar elastic modulus relationship, PVC-U pipes could be used to simulate iron pipes. ρ = 1692 kg/m3, water content ω = 0.18%, specific gravity Gs = 2.62, cohesion c = 0 and the internal friction angle φ = 32°. The corresponding values for the clay were: density ρ = 1430 kg/m3, water content ω = 10.65%, specific gravity Gs = 2.45, cohesion c = 5 kPa and the angle of internal friction φ = 20°.In the test, the sand and clay were packed into the laminar shear box on either side of the pipe and tamped down layer by layer. 2cm thick rubber was embedded in the inner wall of the shear box in order to prevent soil leakage and reduce the impact of the shear box on the pipe’s response. The shear box was as shown in A LVDT  displacement sensor was used in this test; it has an accuracy of 0.1mm. As this kind of displacement sensor can only measure the linear displacement in a single direction on a solid surface, it was located on the outer surface of the shear box, and the arrangement position was buried in the same horizontal plane as the pipe network, which is represented by DS. As the soil was bounded by the soil box, the displacement of the soil was approximately the same as that of the shear box at the same height. Therefore, the displacement sensor DS could be used to measure the displacement of the soil around the pipe network, as shown in Uniaxial strain gauges were used to measure the strain, and the effect of temperature was eliminated by using temperature compensated strain gauges; the accuracy of the gauges was 0.5με. An IMC data acquisition system was used to measure the strain and the displacement measurement at the same time. The strain gauges were placed on the outer surface of the pipes, and special glue was applied to the outer surface of the strain gauges to prevent the strain gauges from being soaked by the water in the soil, as the pipe was buried in the soil. The location of the strain gauges in the pipe network has been shown in The arrangement of the sensors for the pipe network’s tee and elbow components has been shown in The expression of the uniform similarity law was the same as that of the dimensional analysis method used to determine the similarity relationship. However, the equivalent density, after considering the additional part of the artificial mass, was needed to determine the mass density similarity ratio, which is also called the lacking artificial mass model. This model only adds part of the artificial mass and ignores part of the acceleration due to gravity, so there is only a certain degree of the gravity distortion effect. In the shaking table test for seismic simulation, and considering the dynamic response of the pipe network nodes, it was very difficult to ensure that the PVC-U pipe system and the prototype iron pipeline fully met the similarity relationship, which was often impossible to achieve under the existing conditions . In thisg. The actual peak value was 0.4g through the similarity ratio conversion, which was equivalent to a ground motion with an actual intensity of 9 degrees [Due to the limitation of the shaking table test, the seismic wave load only acted in a one-dimensional horizontal motion; the most unfavorable seismic wave input was considered in the test design. An artificial wave was selected as the input prototype wave, as shown in  degrees .In this study, ABAQUS, a three-dimensional finite element program, was used to compare with the test results, and the responses of pipe network nodes, tee and elbow under seismic action were analyzed. A three-dimensional finite element was developed considering the geometry and configuration of the experimental setup. The finite element model of a scaled pipe network sample, i.e., pipe network, sand, clay and viscoelastic boundaries, as shown in The constitutive relations of soil and pipe were mainly based on Mohr-Coulomb model and elastic-plastic constitutive models. The material properties and stress-strain curve for the API 5LB was applied to the pipe model . The cong. Each condition for the implicit method used in the dynamic analysis was determined as follows.The boundary of the model was considered to be viscoelastic . Pipe waA contact condition has various contact models, and of them, the shear friction model was the representative contact model that has been used herein. Normal contact was set to be hard-like contact and the tangential contact surface could only transmit tangential stress , as shown in The maximum value of the soil displacement sensor DS in the shear box of the shaking table was 65.95 mm, and the corresponding time of the maximum displacement was 13.837 s, as shown in εT-n is the measured strain values of S1(T)~S6(T), and εD-n is the measured strain values of S1(D)~S6(D).For a selection time of 13.837 s, the strain value measured by the strain sensor at the measuring point of the pipe network NP has been shown in From The value of the strain at 13.837 s was used to analyze the deformation and stress characteristics of the pipe network NP. The force diagram of the pipe element has been shown in Nf is the axial friction of the pipe element; Np is the axial force at the end x1 of the element; Np is the axial force at the end x2 of the element; E is the elastic modulus of the pipe; A is the cross sectional area of the pipe; ε is the axial strain at the end x1 of the element; ε is the axial strain at the end x2 of the element; L is the length of the pipe element; f is the friction on per unit length of pipe.The friction force per unit length of pipe is:From 2 = 0.9714, this value was greater than 0.8, therefore this has shown that the strain fitting curve met the engineering requirements:S is the axial strain of the pipe in με; x is the position coordinate of the pipe in x direction, in m.The fitted curve was as follows in x direction to calculate the friction acting on the pipe. The direction and magnitude of the friction have been shown in x direction. D is the diameter of the pipe with a value of 0.11 m. The bending deformation of the pipe network is also an important index that can be used to study the response of a pipe network during an earthquake. The bending curvature was obtained from αc of the pipe network nodes. This was used to analyze the deformation characteristics of each type of joint of the pipe network during an earthquake. Ns of the tee element NP-2 and the larger bending moment at the tee joint, the deformation of the tee in sand was larger than that in clay. In contrast, for elbows, the deformation amplification coefficient of the elbows in clay was larger than that in sand, as shown in Ns of a straight pipe that was perpendicular to the seismic wave in clay was larger than that in sand. The bending moment at the elbow in clay was larger than that in sand, so the deformation of the elbow in clay was larger than that in sand.The relationship between node deformation and non-node deformation is able to reflect the seismic response of both the node and the pipe. The deformation of the elbow and the tee is the key to the deformation of the pipe network’s nodes. In this study, the ratio of the maximum strain and the maximum axial strain of the same cross-section measuring point was used as the deformation amplification coefficient λ, to be:The above analysis has shown that the deformation amplification coefficient of a tee was between 2.14 and 8.00, and that of an elbow was between 1.32 and 1.97, which is related to the properties of the soil. Liu and Hou  calculatε = 65.95/1900 = 0.03471 = 34710 με > > 100 με as shown in The displacement of the shear box was 65.95 mm at 13.837 s; due to the inertia the displacement of the shear box was regarded as deformation of the soil, which was compression deformation. The strain in the soil around the pipe was x direction in order to obtain the deformation of the pipe network NP, which was the relative displacement from the elbow point NP-1 to the elbow point NP-5, as shown in Dp of the NP in x direction was 74.6% of the deformation DG of the soil in x direction, and the relative displacement was 25.4% of the deformation of the soil. In other words, about 25% of the deformation energy of the soil was transferred into the internal force and deformation of the pipe during the earthquake and that the larger the ratio of the relative displacement was, the more unfavorable the deformation of the pipe was.The fitting curve of the strain at the measuring point of the pipe network NP was integrated in the g to Gg was on the upper side of the pipe displacement from Ap to Gp. This figure is also a response to the pipe’s friction, that is to say, the friction gradually decreased from Ap to B, and the friction was close to 0 from B to C, which was the midpoint of NP-2. Dp was near the tee, E to F was near NP-4, F to Gp was the right half of NP-4 with increased friction.According to the above data, the graph in ξ in the following Eqs :Dp:Dpξ in For the H-shaped pipe, the maximum value of the displacement of the soil DS in the shear box of the shaking table was 61.307 mm, and the time that corresponded to the maximum displacement was 15.042 s; therefore the selection time that corresponded to the strain was 15.042 s. For the Z-shaped pipe, the maximum value of the displacement of the soil DS in the shear box of the shaking table was 61.088 mm, and the time that corresponded to the maximum displacement of the soil was 8.451 s; therefore the selection time that corresponded to the strain was 8.451 s. The measured strain result, numerical simulation result and the calculated axial strain value of each test point have been shown in t and R are the wall thickness and radius of the pipe, respectively. Therefore, the theoretical local buckling strain in the tee NP-3 was about 8000 με, while the compressive strain of the tee in the test was 40 με; therefore this is far from this degree of buckling. However, during the test, the connection between NP-2 and NP-3 slipped.x direction in order to obtain the relative displacement of the test points at the ends of the H-shaped and Z-shaped pipes, as shown in The fitting curves of the strain at the test points of the H-shaped and Z-shaped pipes were then integrated in the When an underground pipe branches at an intersection, a tee will be added to the straight pipe section of the pipe network. When a straight road becomes a curved road, elbows will be added to the pipe network underneath the road. The seismic strain response after the addition of the tee will make two extreme points appear on either sides of the tee, so that the strain of the pipe will not increase infinitely; this is conducive to the safety of straight pipes in the pipe network. Moreover, the tees will also change the friction direction between the soil and the pipes, so that the friction at the pipe network will tend to reduce. The existence of a tee will make the deformation of straight pipes on both sides of a tee display extreme points in the pipe network, and thus improve the safety of straight pipes. However, due to the existence of a tee in the pipe network, this will aggravate the degree of uneven deformation of the tee, while an elbow set in the pipe network will not aggravate the degree of uneven deformation of an elbow in the case of deformation of a single pipe.An increase in the numbers of tees or elbows in a pipe network will reduce the relative displacement between the pipe network and the soil. Although from the surface, with the decrease in the relative displacement, the seismic response of the pipe network will also decrease, due to the mutual restriction of the deformation among the pipes in the pipe network, and the response of each pipe in the pipe network was often more severe than that of a single type of pipe in the soil, and the damage that resulted at the nodes was also serious.With the increase of the number of elbows or tee joints in the pipe network, the discoordination of the deformation of the pipes will increase, and the force between the pipes and the nodes will increase; this means that the connections between the nodes will be more easily damaged. The results from the above research can provide guidance for the layout and design of an underground pipe network.In this paper, through the use of shaking table tests for a scale pipe network, H-shaped and Z-shaped pipe network components, the response of the pipe network during an earthquake and the influence of tees and elbows on the response of pipe network have been obtained. The following conclusions could be drawn:For the pipe network, the extreme values of strain are produced between a tee and an elbow, where the pipe-soil friction is the smallest and the friction at the tee is the largest. The response of sand to axial strain was greater than that of clay; inhomogeneous geology will also change the direction of the friction.The bending deformation of a tee in the pipe network was larger than that of an elbow, and the deformation amplification coefficient of an elbow in sand was larger than that in clay. For the relative displacement between the pipe network and the soil, most of the displacement of the soil was transferred into the displacement of the pipe, and the remaining small part was transferred into the deformation energy of the pipe network. The relative displacement between the pipe network and the soil was also a response to the friction between the pipe and the soil.The strain and deformation of the straight pipes on both sides of the tee in pipe network have displayed extreme points, which resulted in the straight pipes in the pipe network tending to be safe. Moreover, the pipe network will aggravate the degree of uneven deformation of the tee. An increase in the number of elbows or tees in a pipe network will reduce the relative displacement between the pipe network and the soil, but this will lead to an increase in the discoordination of the deformation of the nodes of the pipe network. The tee and elbow joints are more likely to be destroyed.S1 Table(XLSX)Click here for additional data file.S2 Table(XLSX)Click here for additional data file.S3 Table(XLSX)Click here for additional data file.


IP and oncocytic papillomas  were more often high‐risk (HR) HPV‐positive (15.5% and 16.7%) than fungiform papilloma . CIS and SCC ex‐IP displayed higher HPV rates than benign IP (83.3% vs. 38.8%), especially higher rates of HR‐HPV . Data from this study endorse the hypothesis that recurrence of sinonasal papillomas is promoted by LR‐HPV infection and that malignant progression of IP is promoted by HR‐HPV infection.Sinonasal papillomas are characterized by their potential for frequent recurrences and malignant progression. Currently, the role of human papillomavirus (HPV) infection in sinonasal papillomas is unclear. A study was conducted to elucidate the impact of HPV infection on recurrence and malignant progression of sinonasal papillomas. One hundred and seven patients with 151 tumors could be examined. One hundred and one patients suffered from benign papilloma, mostly inverted papillomas (IP); six patients suffered from carcinomas in situ and squamous cell carcinomas (SCC) ex‐IP. Recurrent IP were more often HPV‐positive than non‐recurrent tumors (38.8% vs. 60%–65%). Low‐risk (LR) HPV infection  increased the risk of tumor recurrences . The authors do not have any conflict of interest and therefore nothing to disclose.This is a completely anonymized cohort study. All patients with clinical or histological/molecular biological data included in this study had to undergo the procedure regardless the scientific evaluation of the histological and molecular biological findings and gave their informed consent prior to the examinations and publication of results.All procedures performed in this examination involving human participants were in accordance with the ethical standards and with the Declaration of Helsinki and its later amendments.1Papillomas in the sinonasal tract are usually arising from the Schneiderian membrane and are therefore called “Schneiderian papillomas”. According to their histopathological characteristics, three different papilloma groups can be distinguished: (a) inverted papilloma (IP), which is the most frequent group preferentially affecting patients between 40 through 70 years of age, (b) fungiform (exophytic) papilloma (FP), which affects patients younger in average (20 through 50 years of age), and (c) oncocytic papilloma (OCP), which is very rare and usually affects patients older than 50 years.Human papillomaviruses (HPV) are DNA viruses infecting mucous membrane and skin cells. Oncogenically, they express a variety of viral antigens or oncoproteins that can lead to uncontrolled cell proliferation in infected tissue.The present paper describes several HPV genotypes identified in sinonasal papillomas and discusses whether such an infection might be regarded as a risk factor for tumor recurrences and malignant progression.22.1For this retrospective study, the operative specimens of 107 patients sinonasal papilloma were examined, who were admitted to the Department of Otorhinolaryngology, Head and Neck Surgery of a tertiary referral teaching hospital. The specimens were collected in a 10‐year period (January 2000 through December 2010). If patients presented with recurrent papillomas within this time , or if patients suffered from recurrence after 2010, samples of their tumors were also studied. In the end, the earliest tumor dated back to 1996, the latest tumor recurrence was treated in May 2017. The clinical follow‐up was 3.1 years in the average.To conduct the retrospective analysis, patients were identified by the hospital information system searching for ICD code D14.0. Finally, 107 patients with histologically confirmed sinonasal papillomas or carcinoma ex‐papilloma could be included in this study.Some patients underwent functional endonasal sinus surgery for non‐suspicious chronic rhino‐sinusitis with nasal or sinus polyps. The diagnosis of a papilloma was then made histopathologically by investigating the specimens. In these cases, a control CT scan of the paranasal sinuses was performed about 3 months following initial surgery, assuming that wound healing is completed by then. If suspicious masses were found in this CT scan or endoscopically during patient examination, patients underwent revision surgery under oncologic conditions. For this study, these cases were counted as persisting papillomas, not as real tumor recurrences.The specimens were obtained during surgery, were fixed in formalin, and then embedded in paraffin. 5‐µm slices were cut from the paraffin‐embedded tissues, dewaxed, and stained with hematoxylin and eosin (HE) for histopathological examination. According to histological results, patients were divided into four groups: (a) patients with inverted papilloma (IP), (b) fungiform papilloma (FP), (c) oncocytic papilloma (OCP), and (d) carcinoma in situ ex‐papilloma (CIS ex‐IP) or squamous cell carcinoma (SCC ex‐IP).This patient cohort was examined extensively for their clinical risk factors of papilloma recurrence previously.2.2® DNA FFPE Tissue Kit  as reported in detail previously.Tumor tissue was identified microscopically in HE‐stained specimens and harvested. DNA was extracted using the spin‐column‐based QiAamp2.3® polymerase , and an appropriate amount of primers (Table The PCR procedure for detection of HPV DNA was performed as reported previously.rs Table . Homogen2.4The HPV Type 3.5 LCD‐array kit (Chipron GmbH) was used for HPV genotyping as reported previously.2.5All descriptive statistics were performed using the GraphPad Prism 5 software. To analyze the impact of the different HPV genotypes on recurrence of sinonasal papillomas, data were compared by univariate logistic regression using the R project for statistical computing. Fisher's exact test was used when investigating the risk of HPV infection and malignant progression of inverted papillomas (GraphPad Prism 5 software). Statistical significance was assumed at p‐values of less than 0.05.33.1p = 0.6161).One hundred and seven patients with 166 tumors were included in this study. Six of the 107 patients (5.3%) displayed 12 tumors with malignant criteria (7.2%), such as carcinoma in situ (CIS) and squamous cell carcinoma (SCC), 101 patients suffered from benign papillomas. In regard to benign papillomas, most of the patients presented with inverted papilloma (IP) (91 patients = 85%), followed by fungiform papilloma (FP) (7 patients = 6.5%), and oncocytic papilloma (OCP) (2 patients = 1.9%).All malignant tumors arose from inverted papillomas. Median age of patients with benign IP at the time of initial diagnosis was 59 years (range 26 through 83 years).Most papillomas recurred within the first 5 years (roughly 74%); however, there were also late recurrences/secondary papillomas after more than 15 years.3.2p = 0.4362). Overall, IP reappeared quite often: 20 patients suffered from one tumor recurrence (22%), five patients from two (5.5%), three patients from three (3.3%), and one patient from more than three tumor recurrences (1.1%). Not all specimens could be investigated for HPV infection, because sometimes patients were referred to our hospital only after tumors recurred and tissue samples from the initial surgery could not be gathered. A total of 80 initially diagnosed IP, 21 first recurrent tumors, and five second recurrent tumors could be examined for HPV infection. Recurrent tumors were more often HPV‐positive than non‐recurrent IP (p = 0.0504): at initial diagnosis, 38.8% of inverted papillomas were HPV‐positive (31 of 80 IP); 65% of first recurrent tumors were HPV‐positive (13 of 21 IP), and 60% of second recurrent tumors (3 of 5 IP). Especially, the rate of HPV 11, 16, 42, and 58 increased in recurrent papillomas; however, none of these HPV types individually revealed a statistically significant higher detection rate in recurrent IP (p = 0.3079 up to 0.8128)  .Twenty‐nine of the 91 patients with inverted papilloma suffered from at least one tumor recurrence (31.9%). Recurrence rates for men and women did not differ significantly (p = 0.4667).Two of the seven patients with fungiform papilloma suffered from tumor recurrence (28.6%). Both patients were male. One of them presented with one recurrent FP, while FP reappeared three times in the other patient. Recurrence rates for IP and FP did not differ significantly. All primary and recurrent FP analyzed in this study were HPV‐positive . HPV 6 and 11 were detected in recurrent FP . Additionally, both of the recurrent FP presented low‐ to moderate‐grade dysplasia on the basis of histological examinations. Two of the seven patients presented with co‐infection of two different HPV genotypes at the time of initial diagnosis. Both patients did not develop tumor recurrences. Therefore, as described for IP patients, co‐infections with multiple HPV types did not increase the risk for recurrence of FP in this cohort of patients . One was HPV‐negative, the other one was HPV‐positive 1 of 2 OCP, 50%) , five patients with squamous cell carcinoma (SCC). All of these malignant tumors developed from IP (none from FP or OCP). Two women and four men were affected. Except for one patient, all of them presented with advanced stage tumors, affecting at least three paranasal sinuses. Two patients (33.3%) presented with non‐resectable tumors. Five of the six patients with malignant tumors ex‐papilloma were initially diagnosed in our hospital. Therefore, the tissue samples of the primary tumors could be analyzed for HPV DNA. Two of them were positive for high‐risk HPV 16 (40%) at the time of initial diagnosis; three were HPV‐negative (60%) at that point.When looking at the course of the disease of the six patients with malignant tumors , five of the six patients displayed HPV positivity during their oncologic course (83.3%). Four cases presented with high‐risk HPV infection . The last HPV‐positive patient showed a co‐infection with HPV 90, for which the risk classification is still under discussion, and HPV 6 Figure .There were four patients with malignant tumors, for whom different regions of the same tumor could be investigated for HPV infection. That means, that tissue was harvested separately from the benign part of the tumor (sometimes displaying dysplasia but not yet carcinoma) and from the malignant part. These benign and malignant parts of the same tumor were then compared with regard to HPV infection. Three of the four cases were HPV‐positive in the benign part of the tumor, but no virus could be detected in the SCC part. For one patient, it was the other way round.All of the six patients displayed malignancy at the time of initial diagnosis but for one exception: one patient presented with benign papilloma initially and returned after years with recurrent SCC ex‐papilloma. In this patient, HPV 16 DNA was detected in the primary tumor (IP) but not later in the SCC.In regard to the other risk factors, two patients with malignant tumors ex‐IP were active smokers . In comparison to our earlier study, 33.3% of male benign IP patients and 38.5% of female IP patients were active smokers or had smoked before.4p = 0.6161). Additionally, we could not confirm some of the previously described possible risk factors, such as higher age and smokingSchneiderian papillomas are rare and usually benign tumors that arise from the sinonasal Schneiderian membrane. In the retrospective study presented in this article, we focused on the possible impact of HPV infection on tumor recurrence and on malignant progression of sinonasal papillomas. A cohort of 107 patients with 166 tumors was investigated. In decreasing number of frequency, patients were diagnosed with inverted papilloma (IP) (91 patients = 85%), fungiform papilloma (FP) (7 patients = 6.5%), and oncocytic papilloma (OCP) (2 patients = 1.9%). For one patient, papilloma entity  could not be clearly determined. Malignant progression is a rare but severe complication of Schneiderian papillomas; therefore, risk factors and predictors for carcinogenesis have been subject to controversial discussions. The study presented here includes six patients with malignant tumors ex‐papilloma, that is, one patient with carcinoma in situ (CIS) and five patients with squamous cell carcinoma (SCC). Tissue samples from 12 malignant tumors could be gathered from these six patients. All malignant tumors arose from IP (none from FP or OCP). Gender did not seem to have an important impact on malignant progression of Schneiderian papillomasOur previous analysis found that two patients showed dysplasia in FP (not yet CIS). Both suffered from recurrent tumors.Human papillomaviruses (HPV) are DNA viruses that infect differentiating skin and mucous membrane cells. Through expression of viral oncogenes they can induce uncontrolled cell proliferation and even malignant progression.p = 0.0504): at initial diagnosis, 38.8% of inverted papillomas were HPV‐positive (31 of 80 IP), first recurrent tumors were 65% HPV‐positive (13 of 21 IP), and second recurrent tumors were 60% HPV‐positive (3 of 5 IP). Especially the rate of HPV 11, 16, 42, and 58 increased in recurrent papillomas, although none of these types individually showed significantly higher detection rates in recurrent IP (p = 0.3079 up to 0.8128) .The role of HPV infection as a risk or prognostic factor for recurrence rates or rates of malignant progression of sinonasal papillomas is still under controversial discussion. Some authors have observed that recurrence occurs more often in patients infected with HPV,ositive 3 of 80 IPp = 0.0780). Additionally, patients with malignant tumors ex‐IP displayed higher rates of HR‐HPV than patients with benign inverted papillomas . Detected HR‐HPV genotypes comprised HPV 16 , HPV 45 and 66 . Concerning malignant potential, both IP and OCP (but not FP) have been associated with malignant conditions, especially squamous cell carcinoma.p = 0.3333) . In the head and neck area, many reports have been published on the probable role of HPV in oropharyngeal cancer: Ndiaye et al. published an extensive review with evidence that roughly 42% of oropharyngeal cancers are related to HPV in central Europe.Returning to the data gathered in sinonasal papillomas in our study: there were four carcinoma ex‐IP patients, for whom different regions of the same tumor could be investigated for HPV infection: when comparing tissue derived from the benign part of the tumor (sometimes displaying dysplasia but not yet carcinoma) to parts of IP with proven malignancy, three of four patients were HPV‐positive in the benign part of the papilloma, but no virus could be detected in the SCC part of the same tumor. For one patient, it was the other way round. Furthermore, one patient initially presented with benign papilloma and returned after years with recurrent SCC ex‐papilloma. In this patient, HPV 16 DNA was detected in the primary tumor but not later in the SCC. Lawson et al. hypothesize that HPV can initiate development of papillomas but is not important for them to persist.To provide a wider picture, in regard to oncogenic HPV infections, two opposite modes of carcinogenic potential have long been discussed. On the one hand, there are authors providing evidence for a hit and run mode of HPV‐associated carcinogenesis.In conclusion, our data suggest that HPV infection, particularly infection with LR‐HPV, such as HPV 6, can be regarded as a risk factor for tumor recurrences of sinonasal papillomas. Furthermore, malignant progression of inverted papillomas seems to be a multi‐step process that may be supported (but not exclusively caused) by high‐risk HPV genotypes. Further studies are required to elucidate the potential impact of HPV on the disease course of inverted papillomas, which could help to improve therapeutic strategy and post‐therapeutic management for the patients.


The prevalence of HPV16/18 infection significantly decreased from 1.3% in 2014 to 0% in 2017 (p = 0.02). The three most prevalent types were HPV52, 16 and 56 in 2014, and HPV52, 58 and 56 in 2015–2017, respectively. HPV16 and 33 infection rates decreased. On the other hand, the HPV58 infection rate was obviously increased after OHPV from 0.3% to 2.1%. Our study demonstrates that the prevalence of HPV16/18 infection dramatically decreased and the profile of type-specific HPV infection was changed after OHPV.Organized human papillomavirus vaccination (OHPV) in Japan was introduced in 2010 for girls aged 12–16 years who were born in 1994 or later. The rate of OHPV coverage was 70–80%. However, after suspension of the government vaccination recommendation, the coverage dramatically decreased. We aim to investigate the change in prevalence of HPV infection after the initiation of HPV vaccination. We recruited females aged 20–21 years attending public cervical cancer screening from 2014 to 2017 fiscal years (April 2014 to March 2018). Residual Pap test specimens were collected for HPV testing. We compared the prevalence of HPV type-specific infection between women registered in 2014  and registered in 2015–2017 . We collected 2379 specimens. The vaccination coverage figures were 30.7%, 86.6%, 88.4% and 93.7% ( In Japan, there are over 10,000 cases of cervical cancer annually; nearly 3000 women die from the disease . For cerTM  for pooled infection with one or more from 13 high-risk HPV genotypes: HPV types 16, 18, 31,33, 35, 39, 45, 51, 52, 56, 58, 59 and 68. Only those samples positive for the HC2TM test underwent HPV genotyping with the MEBGENTM HPV kit  [We recruited 2379 women aged 20–21 years attending a public cervical cancer screening held in Niigata, Japan, from fiscal years (FY) 2014 to 2017 (FY2014–2017: from April 2014 to March 2018). We performed an HPV test, HCII, for screening and used the MEBGEN kit for HPV genotyping. Residual liquid-based cytology  specimens from the cervical screening were used for HPV testing. All samples were tested with Hybrid Capture (HC) 2The target group in this study is shown in p value of <0.05 was considered to indicate statistical significance.All experiments were performed in accordance with the relevant guidelines and regulations, and written informed consent was obtained from all participants. The present study protocol was approved by the institutional review board of Niigata University Graduate School of Medical and Dental Science and registered at the UMIN Clinical Trials Registry, trial number UMIN000026757. Data were analyzed using EZR , which is a graphical user interface for R . A two-sided The background of participants in this study is shown in p for trend <0.01). However, the prevalence of high-risk HPV infection was slightly increased from 10.0% in 2014 to 11.3% in 2015, 11.2% in 2016 and 11.6% in 2017 (p for trend = 0.54). On the other hand, in regard to vaccine types HPV16/18 infection, the prevalence significantly decreased from 1.3% in 2014 to 0.4% in 2015, 0.4% in 2016 and 0.0% in 2017 (p for trend = 0.02), sequentially  in 2014, because the group included women born in 1994 (OHPV generation). If the rate had been sufficiently low, the profile of type-specific HPV infection might have clearly changed after initiation of HPV vaccination.The prevalence of vaccine types HPV16/18 infection dramatically decreased after the initiation of HPV vaccination. However, the HPV vaccination rate has been nearly 0% in Japan, so Japanese girls are once again at risk for HPV16/18-related cervical cancer. This risk could be mitigated if the Japanese Ministry of Health, Labor and Welfare resumed proactive recommendations for the vaccine.


Five-year cumulative incidence of recurrence was 46 %, and was not statistically related to HPV presence (p = 0.22) or genotype (p = 0.44). p16 sensitivity, specificity, PPV, and NPV in predicting HPV infection were 27.3%, 36.4%, 46.2%, and 20.0%, respectively. In our experience, beta HPV was more frequent than alpha HPV in MESCC. Neither HPV presence nor HPV genotypes relate to DSS or cumulative incidence of recurrence. p16 expression was not predictive for HPV infection in MESCC. The role of HPV infection in oncogenesis, maintenance, and prognosis of MESCC seems to be different from that in oropharynx and skin cancer.Middle ear squamous cell carcinoma (MESCC) is rare. Human Papilloma Virus (HPV) infection has been found in a significant number of cases of MESCC. Despite the emerging role of HPV in oncogenesis, its role in the pathogenesis and prognosis of MESCC is not known. This study aims to identify the prognostic impact of alpha and beta HPV in MESCC and its correlation with p16 protein. We retrospectively investigated 33 patients with MESCC surgically treated between 2004 and 2016. HPV DNA was ascertained by polymerase chain reaction (PCR) and P16INK4a detection was performed. Disease-specific survival (DSS) and cumulative incidence of recurrence were calculated in relation to HPV presence and genotype. p16 sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) in predicting HPV infection were calculated. HPV was detected in 66.7% of patients . Five-year DSS was 55.0% and was not statistically related to HPV presence ( Primary middle ear squamous cell carcinoma (MESCC) has an annual incidence of 0.8–1 per million inhabitants and it is the most common histology in this site . In partThe carcinogenic role of Human Papilloma Virus (HPV) has emerged in last decades. Alpha HPVs can be categorized as high-risk (HR) or low-risk (LR) according to their carcinogenic potential , whereasBeta HPVs have been detected also in oral cancer and in middle ear tumors, but their role in carcinogenesis has not been fully investigated ,8,9.HPV has been found both in inflammatory middle ear disease and in cancer, but its role as co-pathogen is not fully understood yet . FurtherThe prognostic role of HPV infection, using p16 as a diagnostic biomarker, has been demonstrated in oropharyngeal SCC (OPSCC) ; conversThe objective of the present study was to evaluate the association between HPV infection and MESCC, its prognostic role, and possible association with p16 expression.Patients diagnosed with MESCC who underwent surgery between 2004 and 2016 were selected from our institutional oncologic database to be retrospectively reviewed. The study was conducted according to the principles stated in the Declaration of Helsinki and approved by the local Ethics Committee (# RS852010). Patients gave their written informed consent for the use of their data. Inclusion criteria were: primary tumors of the middle ear with SCC histology, surgical resectability, and at least a 24-months follow-up. Exclusion criteria were: patients with recurrent tumors, skin tumors extended to the middle ear, previous radiotherapy (RT), and/or chemotherapy.Tumor staging was determined using ear and neck computed tomography (CT) and/or magnetic resonance imaging (MRI) with contrast, and a chest CT. Tumors were classified according to the Arriaga staging system . SurgicaIndications for adjuvant intensity-modulated radiotherapy (IMRT) were Arriaga stage ≥III or close/positive resection margins at histology . IMRT wa® p16INK4a, Roche Diagnostics, Basel, Switzerland) and was defined by a strong and diffuse nuclear and cytoplasmic staining in ≥70% of the tumor cells. The presence of HPV and EBV (to exclude contamination from the nasopharynx) DNA was retrospectively ascertained on histological samples. Surgical specimens were embedded in formalin and a nested polymerase chain reaction (PCR) with consensus primers was carried out followed by direct sequencing according to previously reported standard procedures . P16INK4Follow-up clinical evaluations were scheduled every three months for the first two years and then twice a year for a total of at least five years. Head and neck CT scan or MRI were performed every 6 months for the first two years and thereafter once a year for the following years to identify loco-regional recurrence. 18-fluorodeoxyglucose (FDG) positron emission tomography (PET) CT scan was performed once a year to detect loco-regional and/or distant metastasis.Categorical variables were reported as absolute frequencies and percentages. Continuous variables were summarized as mean ± standard deviation and range. Disease-specific survival (DSS) rate was defined as the interval between the date of surgery and the date of the death by the specified cancer, with censoring at the last examination date for patients who were alive. The median follow-up was computed for censored patients, excluding patients with events of interest (reverse Kaplan–Meier method). DSS was estimated using the Kaplan–Meier approach and differences of survival between groups  were assessed through the Log-rank test.Disease-free survival (DFS) rate was defined as the interval between the date of surgery and the date of local (T) and/or regional (N) recurrence, with censoring on the last examination date for patients who were disease-free. The cumulative incidence function (CIF) was used to describe recurrence trends, taking into account the presence of competing risks  since most patients died precluding the occurrence of relapse. Cumulative incidences of recurrence between groups  were compared with the Gray test. CIF was calculated with the R package cmprsk . The performance of p16 to predict HPV infection calculating sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) was evaluated for all types of HPV and for alpha versus beta HPV with 95% CI.A total of 33 patients met the inclusion criteria and were recruited for the present study. Mean age was 75 ± 11 . Information on patient demographics, tumor characteristics, and treatment type are summarized in HPV DNA was found in 66.7% of MESCC, of which 36.4% were alpha HPV and 63.6% were beta HPV. Eighteen patients died from MESCC, 14 died from other causes, while 1 patient was alive at the time of follow-up closure. p = 0.55) (p = 0.87) (Median follow-up was 9.62 years (IQR 4.50–11.14), , and the median survival was 6.06 years. Five-year DSS was 55.0% (CI 95%: 35.2–71.0%); DSS was not statistically associated to HPV presence ( = 0.55) a nor to  = 0.87) b.p = 0.22) (p = 0.44) (The 5-year cumulative incidence of MESCC recurrence was 46.0% (CI 95%: 28–62%) with a median time of MESCC recurrence at 5.30 years. Among the 19 patients who experienced MESCC recurrence, 10 (52.6%) had T recurrence, and 9 (47.4%) had N recurrence. HPV-negative patients experienced a higher incidence of recurrence compared to HPV-positive patients (0.64% versus 0.36%), but the difference was not statistically significant ( = 0.22) a. Recurr = 0.44) b.Sensitivity, specificity, PPV, and NPV of p16 in predicting HPV infection were 27.3% (CI 95%: 10.7–50.2%), 36.4% (CI 95%: 10.9–69.2%), 46.2% (CI 95%: 19.2–74.9%), and 20.0% (CI 95%: 5.7–43.7%), respectively.Sensitivity, specificity, PPV, and NPV of p16 in predicting alpha HPV infection were 37.5% (CI 95%: 8.5–75.5%), 36.4% (CI 95%: 10.9–69.2%), 30.0% (CI 95%: 6.7–65.2%), and 44.4% (CI 95%: 13.7–78.8%), respectively.Sensitivity, specificity, PPV, and NPV of p16 in predicting beta HPV infection were 21.4% (CI 95%: 4.7–50.8%), 36.4% (CI 95%: 10.9–69.2%), 30.0% (CI 95%: 6.7–65.2%), and 26.7% (CI 95%: 7.8–55.1%), respectively.This study focused on the prognostic impact of HPV in MESCC, and the relationship between p16 presence and HPV infection status in a homogeneous group of patients in terms of staging, treatment, and follow-up. The presence of alpha and beta HPV is reported in middle ear mucosa but their role in carcinogenesis, and the relationship with middle ear disease, still needs to be established . Alpha HThe difference between HPV-positive MESCC in our cohort (66.7%) compared to that reported by Surono et al. (29%) may be due to the fact that HPV was detected by immunohistochemistry, a low-sensitive method, in the latter . PreviouThe prognostic impact of HPV is well known in pharyngeal SCC. Specifically, HPV-positive OPSCC are characterized by a better progression-free survival and overall survival ,25, and Previous studies reported the presence of p16 in 38–64% of oropharyngeal, 7–28% of hypopharyngeal, and 9–80% of nasopharyngeal mucosal cancers . In our Alpha HPV can induce up-regulation of p16 and concordance between p16 and HPV was the basis for the current use of p16 as a surrogate immune-histochemical (IHC) marker for HPV infection in OPSCC . ConversHPV can be present in MESCC. In our sample, MESCC prognosis was not influenced by the presence nor by the HPV genotype; moreover, p16 positivity was not associated with HPV presence. A high percentage of MESCC patients presented with otorrhea. Future studies should investigate on the role of chronic infection in MESCC pathogenesis. Considering the low incidence of this cancer, multicenter studies are advised in order to verify this hypothesis.


This study was to conduct a predictive model for the prognosis of aneurysmal subarachnoid hemorrhage (aSAH) and validate the clinical data.A total of 235 aSAH patients were enrolled in this study, dividing into the favorable or poor prognosis groups based on Modified Rankin Scale (mRS) at 3 months postoperatively. Multivariate analysis was assessed using binary Logistic regression and Fisher discriminant analysis. The receiver operating characteristic (ROC) curve was used to determine the cut‐off value.Our findings showed that the high Glasgow Coma Scale (GCS) score 24‐hour after surgery reduced the risk of poor prognosis, and the surgical clipping and elevated neutrophil‐lymphocyte ratio (NLR) increased the risk of poor prognosis. The discriminant function was V = 0.881 × GCS score − 0.523 × NLR − 0.422 × therapeutic approach, and V = −0.689 served as a cut‐off value. When V ≥ −0.689, the good prognosis was considered among these patients with aSAH. The correctness for predicting the prognostic outcomes by self‐validation was 85.11%.This predictive model established by a discriminant analysis is a useful tool for predicting the prognostic outcomes of aSAH patients, which may help clinicians identify patients at high risk for poor prognosis and optimize treatment after surgery. The ROC curve was established according to V as the test value and mRS score results as the gold standard  and the critical value of V (cut off) was −0.689. When V ≥ −0.689, the prognosis was considered as good, otherwise as poor All cases were consecutively recruited in the retrospective investigation that were divided into the favorable (n = 201) or poor (n = 34) prognosis groups based on Modified Rankin Scale (mRS) at 3 months postoperatively.Inclusion criteria: (a) 18‐80 years old; (b) clearly diagnosed as aSAH according to guidelines for the diagnosis and treatment of Chinese subarachnoid hemorrhage; (c) CT scan of the head confirmed aSAH; (d) patients with first rupture of aneurysm by CT angiography (CTA) and digital subtraction angiography (DSA); (e) admission within 24 hours of the initial symptom onset; (f) hospitalization ≥3 days in neurosurgery.Excluded criteria: (a) aSAH patients caused by non‐aneurysmal causes such as arteriovenous malformation or arteriovenous fistula; (b) patients with histories of acute or chronic infections, autoimmune diseases, recent cardiovascular and cerebrovascular diseases, and other systemic diseases including malignant neoplasms, uremia, chronic heart disease, and chronic pulmonary disease; (c) previous use of anticoagulant/antiplatelet drugs; (d) patients with histories of acute craniocerebral trauma, brain tumor or stroke with neurological dysfunction; (e) incomplete clinical data; (f) patients with preoperative rebleeding of aneurysms and surgical treatment failure; (g) severe liver and kidney dysfunction; (h) refuse or lost to follow‐up; (i) patients who have taken other experimental drugs or ongoing clinical studies within 1 month.2.2The blood biochemical indexes were tested using the automatic biochemical analyzer. The clinical indicators consisted of high‐density lipoprotein (HDL), low‐density lipoprotein (LDL), heart rate (HR), neutrophil‐lymphocyte ratio (NLR), platelet‐lymphocyte ratio (PLR), white blood cell (WBC) count, platelet (PLT) count, neutrophil count (NEUT), lymphocyte count (LC), glutamic oxaloacetic transaminase (AST), albumin and fasting blood glucose (FBG).2.3Glasgow Coma Scale score mainly includes Motor response (M), Verbal response (V), and eye‐opening (E). The degree of coma is evaluated based on the sum of M, V, and E scores. A score above 14 was considered as normal, a score below 7 was coma, and GCS score ≤3 was indicated brain death or poor prognosis.2.4(x¯±s) and analyzed by ANOVA. Counting data were presented as n (%) with χ2 or Fisher tests. The single‐factor analysis of the prognosis at 3 months postoperatively was carried out by t, Kruskal‐Wallis or χ2 tests. Multivariate analysis was assessed using binary Logistic regression and Fisher discriminant analysis. The receiver operating characteristic (ROC) curve was used to determine the cut‐off value. P < .05 was considered statistically significant.Statistical analysis was performed using SPSS 23.0 . Measuring data were presented as the mean ± standard 33.12χ = 0.04, P = .836), age , BMI , drinking , smoking (P = .378), diabetes (P = .068), hypertension , operation history (P = .610), surgical timing (P = .345), initially ruptured aneurysm bleeding (P = .855), aneurysm numbers , and sizes (P = .153) between favorable and poor prognosis groups. Significant differences in Hunt‐Hess grade  and surgical techniques  were showed between the two groups  and albumin  in the favorable prognosis group was higher than that in the poor prognosis group, with statistical differences. The levels of NLR , PLR , HR , WBC count , NEUT , AST , and FBG  in the poor prognosis group were higher than the favorable group. We also found that there were no statistical differences in rebleeding or death 24 hours after onset (P = .054), HDL , LDL , PLT count , and LC  between the two groups (Table The GCS score (3.3P = .001). The risk of poor prognosis in the surgical clipping was higher than that in the endovascular therapy . In addition, the higher the NLR, the higher the risk of poor prognosis .In Table 3.42χ = 77.36, P < .001), with significant differences.The GCS score 24‐hour after surgery, surgical techniques, and NLR was as independent variables, the prognosis at 3 months postoperatively was as the grouping variable, and Fisher discriminant analysis was carried out. Then, the discriminative equation was as follows: V = 0.881 × GCS score − 0.523 × NLR − 0.422 × surgical techniques. The value of Wilks'Lambda was 0.712  and surgical techniques  were assigned in this study. The V‐value was calculated based on the discriminative equation. The ROC curve (Figure The self‐validation method was used to assess the efficacy of the predictive model in Table 4Aneurysmal subarachnoid hemorrhage is a serious disease that threatens human health. Previous studies reported that >30% of mortality was related to the existence of aSAH, and merely about 30% of aSAH patients could return to independent living.Studies have shown that the discriminant analysis has been clinically used to select significant indicators.Previous studies have demonstrated that NLR was associated with the outcomes and could predict the courses of different medical conditions, such as ischemic stroke,In addition, we also discovered that the risk of poor prognosis in the surgical clipping was higher in comparison with the endovascular therapy. The surgical clipping is a gold standard treatment in recent decades, and Yasargil applied the microsurgical techniques to neurosurgery to improve the surgical approach to intracranial aneurysms.A strength of this study is that we established a predictive model for the prognosis of aSAH, and validated with the internal data. The high GCS score 24‐hour after surgery was a protective factor, and the surgical clipping and NLR were risk factors for the occurrence of poor prognosis in aSAH patients. The establishment of our discriminant function had a good performance for predicting the prognostic outcomes in aSAH patients. There were some limitations in our study. First, the sample size may reduce the statistical power. Second, only a small number of enrolled patients developed poor prognosis, which may affect the statistical outcomes. Third, due to the possible influence of treatments, the longitudinal assessment of laboratory indices over time was not carried out. Additionally, the predictive value of this model was evaluated base on an internal data, lacking of the external validation. These should be cautious in interpreting the results. Hence, future studies should further validate the results of the present study.5We established a predictive model to assess the prognosis of aSAH using a discriminant analysis, and to conduct an internal validation to identify the effectiveness of this model. Our results revealed that the correctness for predicting the favorable prognosis was 85.67%, as well as for predicting the poor prognosis was 76.47%. The accuracy obtained by discriminant analysis was 85.11%, indicating that the effectiveness of this predictive model was relatively reasonable. These findings obtained from our study may help clinicians identify patients at high risk for poor prognosis and optimize treatment after surgery.