Orf virus causes a scabby skin lesions which decreases productivity in small ruminants. The unknown status of this disease in the eastern region of Peninsular Malaysia warrants a study to determine sero-prevalence of orf with regards to farmers’ compliance level towards the Herd Health Program (HHP) programme.P < 0.05). Notwithstanding, all other variables showed no significant difference (P > 0.05). Farms surveyed usually practised intensive management system, keeping animals in the shade at all time, due to limited availability of suitable land as a free-range grazing area. An interview with small holder farmers revealed a lack of awareness of the main goals of herd health programme. An overall compliance level of 42.7% was observed for all HHP parameters. Among the 14 main components of HHP modules, animal identification had recorded highest compliance level (84.62%) while milking management recorded the least compliance (− 82.69%). That explained why there was a high sporadic prevalence of Orf infection in this region.Out of 504 animals, 115 were positive for Orf-virus antibodies. An overall prevalence rate of 22.8% indicated a high prevalence of orf disease in this region. It was observed that 25.1% (92/367) of goats were positive and 16.8% (23/137) of sheep sero-converted for Orf virus antibody. Several factors were measured for their possible association with prevalence of Orf virus infection. The prevalence was higher in LY farm, JC breed, kid and female animals, and in the presence of disease lesion. Chi-square analysis showed a significant association of three risk factors which are species, age and sex of the animals  contains supplementary material, which is available to authorized users. Small ruminants have important contributions to human kind and sustainability in terms of their meat, milk, and other ornamental products including biological products for studies of diabetes and insulin production –4. HowevThe best method for diagnosis of Orf virus infection is culture of the virus on susceptible cell lines –18 but tAt present some commercial vaccines can control this disease however, anecdotal epizootics of a persistent generalized form of the infection has been reported among goats vaccinated with the first generation orf vaccine prepared for sheep . SimilarMany outbreaks of contagious ecthyma disease in different part of the world including but not limited to Africa, Middle East, Europe, North America and most of the Asean countries including Malaysia , 22, 23 All the 13 farms surveyed have consented and responded to the questionnaire. A total of 504 blood samples were collected from sheep and goats.P-value of less than 0.05  were positive for orf virus antibody based on the ELISA assay. This indicated an overall score of 22.8% among the animals sampled across the 13 farms selected. Goat population had the highest percentage (25.1%) of sero-conversion compared to sheep with 16.8% positive results. The finding is statistically significant with 15 were pThere is no cross-reactivity observed upon testing with other related viruses including bluetongue virus (BTV) and Schemallenburg virus (SBV) positive sera samples obtained from sero-converted infected animals. Both BTV and SBV postive control sera produced an optical density (O.D) value below the cut-off reading. Noteworthy, based on the sensitivity (Se) and specificity (Sp) obtained from ROC-curve analaysis determined using MedCalc sortware, the ELISA presented an Se and Sp of approximately 95.2 and 97.8% respectively showed by the area under the curve (AUC).2) = 17.889; P = 0.1191).Table 2 = 17.093; P = 0.0723).Table 2 = 8.163; P = 0.0428). The association amongst the various age groups and rate of sero-conversion antibody for orf virus infection is significant .The highest prevalence of orf disease was found among kids of less than 3-months old. All of them were sero-converted. This was followed by 29.7% in animals of older than 4 years. Interestingly, animals aged 4–9 months have the lowest positive rate (20%). The association amongst the various age groups and rate of sero-coversion for orf virus infection is significant .From a total of 174 male sheep and goats that were examined, 31 (17.8%) were sero-converted for Orf virus. However, out of a total of 330 female sheep and goats examined, 84 (25.5%) were sero-converted for the virus .The rate of sero-conversion among animals with clinical orf disease (66.7%) was higher than the animal’s who are devoid of obvious clinical manifestations of orf (22.6%). The association between the occurrence of clinical orf disease and sero-conversion is not significant  for HHP. However, the effect of HHP modules on farms BF, MF, LH, LB and LW did not record a significant compliance level (p > 0.05); indicating decrease compliance level by these farms to parameters of HHP modules. The overall effect likelihood ratio of HHP program on various farms compliance levels is strong and significant .Table 2 = 114.77; P < 0.0001). Mosaic plot presentation of the main Components of HHP Modules and compliance level is shown in Fig. Table Figure This investigation reported the current status of orf virus disease in the Terengganu state, eastern coast region of Peninsular Malaysia. The study revealed evidence of the presence of orf virus infection among the sheep and goats’ population as confirmed by ELISA assay which amounted to a prevalence of 16.8% in sheep and 25.1% in goats. Replication of Orf virus occurs in the epithelial cell surfaces of the skin layers and at the mucosa of the mouth, oesophagus as well as hairless parts of the body serving as primary sites of the lesions. Accidental abrasions of the skin due to hard stubble, thistles or any analogous plant promote access of the virus and initiates the replication cycle –31. UponELISA test employed in this study revealed an overall seroprevalence rate of 22.8% based on antibody titre among the sampled population. This indicates a considerably high sero-conversion rate of the disease compared with a similar study in the Selangor, Malaysia with a prevalence rate of 14.4% in goats and 12.2% in sheep . HoweverVarious putative risk factors for the prevalence rate were examined to ascertain the possible risk factors. Identification of relevant risk factors is crucial for the proper disease management and outbreak containments. We identified species, age and sex of the animal to be the most significant risk factors. Goat species recorded highest prevalence of 25.1% compared to 16.8% that of ovine like the report by Jesse et al. . NaturalSimilarly, female gender had the highest prevalence and this was recorded as a significant risk factor in this study. A similar observation was reported by Orgeur et al. . More agAmong the 13 farms, farm LY showed the highest historical orf virus infection. This is strongly associated with the fact that farm LY recorded the highest non-compliance levelof HHP. A strict adherence to HHP modules shall reduce the general risk of exposure to contagious ecthyma disease in animal population , 44. AniOther risk factors examined such as the presence lesion and abrasion, history of clinical orf infection in farms and vaccination practice did not appear to be a significant determinant of orf disease prevalence. The presence of skin lesions usually indicates current infections which are best diagnosed by standard virus isolation and identification , 45. AntThe HHP module analysed in this study contains variables that are pertinent to epidemiology of orf disease as previously described elsewhere , 52, 53.The general objective of HHP is to enhance the herd efficiency through general farming, nourishment administration, vaccination, ecological management and parasite control. It endeavors to arrange all data appropriate to goat crowd wellbeing into a straightforward, usable, and effectively recalled lists. Therefore, a proper record-keeping must be in place to enable and ensure the success of HHP , 44. VacDisease monitoring is concerned with understanding changes in its endemicity and distribution . Proper Among the 14 main components of HHP modules, milking management is observed to have the highest level of non-compliances (82.69%), whereas, animal identification had recorded highest compliance level of 84.62%. The highest compliance level associated with animal identification indicated an appreciable livestock production is in accordance with the standard recommended by the OIE. Herd-health information is pertinent to livestock producers and to the public as well as animal welfare, fully comprehending of types and sources of animal’s heath that farmers will utilized is very vital .Many studies carried on the relationship between dose antibody responses and virus replication have generated controversies –63 The lControl and prevention of Orf disease is important to ensure it does not widely spread in the entire animal population –73. A viInteraction between virus and body immune system is a battle among the parties (virus and host). In primary orf virus disease, the virus replicates in the epithelial cells for a period before the host can mount an effective immune response. This leads to an appearance of IgG or IgM that were incriminated as a signal of orf infection. The overall prevalence was found to be 22.8% (25.1% in goats and 16.8% in sheep). Significant risk factors identified were specie, age, and sex. A higher sero-conversion rate was seen among kids younger than 3-months-old and female gender showed higher antibody titre. Poor implementation of HHP may also be associated with a higher sero-conversion rate of orf virus infection observed. Therefore, it is important to carry out epidemiological surveys , 75 in cDaerah in Malay language. The sampling was strategized to capture 4 out of the 8 eight administrative districts as the representative of this state and a total of 13 farms were selected based on the simple random sampling technique. Consent from all participating farms were obtained through written permission of the owners and witnessed by the Terengganu State division of the Department of Veterinary Service (DVS).All the procedures involving animal subjects were conducted in compliance with the recommendations of the Institutional Animal Care and Use Committee (IACUC) – UPM/IACUC/AUP-U013/2018. Goats and sheep from thirteen (13) farms were selected among the private and government owned farms at 4 districts of the Terengganu state. The sampling farms were selected on the basis for the availability of adequate study animals and diversity in agroecology of the areas. Terengganu State is divided into eight (8) administrative districts called A well-structured questionnaire which contained information on farm management practices, possible risk factors and herd health programme implemented by farm owners were filled via an interview session. The questionnaire was designed to contain three (3) sections, namely; Section A (farm management practice), Section B (farm’s HHP compliance level) and Section C . The questionnaire template was added separately in the Additional file Section A of the questionnaire which relates to informations on the sampled farms was administered. The relevant data sought included; details of the operator, category of farmer, man-power, annual production, type of housing and management system, as well as population. Section B on the other hand (farmer’s compliance level to HHP), contains questions relating to the farmer’s awareness, compliance level, and knowledge of each of the 14 modules of herd health programs, based on the Department of Veterinary Service, Malaysia Table . Lastly,This investigation involved thirteen (13) sheep and goat farms located at the four major districts namely; Kuala Terengganu, Kuala Nerus, Marang and Setiu in the Terengganu State, East Malaysia. The respondents were given the questionnaire; a response to each question is a dichotomous outcome as either “YES” or “NO”. Where “YES” denotes the farmers’ compliance to that segment of HHP module, while “NO” is otherwise.A Thorough physical examination to identify infected animals based on the clinical signs of erythema, papule, vesicle, or pustule around the lip, gums, mouth and tongue and the general body part was conducted. Relevant demographic data from each animal was also recorded in the data sheet prior to sampling. A total of 504 sheep and goat’s samples were collected using simple random sampling method after calculating the sample size according to the standard formula , 77. TheZ = Standard normal distribution at 95% confidence interval = 1.96p = Prevalence in similar workq = 1 – pL = Allowable error, taken as 5% = 0.05P = 36.7% In this study, Whole blood samples were collected from all the 504 animals in the sampling farms as. A method of collection via jugular venepuncture using a 21 gauge vacutainer needle was applied and collected into a plain serum collection tubes. The tubes containing the whole blood samples were then stored in a cooler box and transported to the laboratory for analysis. The blood samples were left to clot and centrifuged at 3,000 revolution per minute (rpm) for 5 min to separate the serum from the blood. The serum was then pipetted into a 1.5 ml microcentrifuge tubes and stored at -20 °C until required for assay.Hyperimmune serum (HIS) against the UPM1/14 Orf virus isolate was prepared according to the method described by , 78–80. The serological screening was done using an in-house developed antigen-coated ELISA. The virus antigen used for the coating of ELISA plate was a local orf virus isolate (UPM1/14) obtained from the Virology Laboratory, Faculty of Veterinary Medicine of the University Putra Malaysia from previous outbreak cases of contagious ecthyma . This viThe working concentrations for Orf antigen, conjugate and antibodies were optimized using standard protocol of chequerboard titration adopted from Babiuk et al., ; Bhanupr3) buffer (pH 9.6) to give a final concentration of 10 μg/ml antigen protein, and 50 μl of this was used for coating of the plate and then incubated at 4 °C overnight. Following overnight incubation, the plate was washed three times with Phosphate Buffered Saline Tween-20 (PBST), the washing was carried out manually by filling each of the well with 200 μl of the washing buffer, with the aid of a multi-channel micropipette, and then allowed to stand for a few seconds before discarding. This wash procedure was repeated 3 times. After washing 50 μl of 2% of Fraction V (Bovine Serum Albumin (BSA))  was added and the plate was incubated at 45 °C for 2 h, in order to block any unspecific unbound antigens. Upon completion of the incubation, the plate was again washed three times with PBST-tween-20. A two-fold dilutions of the test sera were added to the plate and incubated at 37 °C for 1 h before the plate was washed 3 times using the same wash buffer. A pre-diluted rabbit anti-goat (for goat serum) and anti-sheep (for sheep serum) peroxidase conjugated immunoglobulin G  was added and allowed to react with the antigen bound goat/sheep antibodies by incubation at 37 °C for 1 h. The plate was then washed 3 times and the substrate 2-2l – Azino-bis (3-ethylbenzthioline − 6-sulfonic acid) ABTS diluted in citrate phosphate buffer (containing 0.01 30% hydrogen peroxide (H2O2)) was added and the plate was incubated at room temperature for 30 min. At the end of the incubation period, the optical density was read at 450 nm in an ELISA reader (TECAN Infinite M200).An enzyme linked immunosorbent assay (ELISA) was developed in-house as described by Azmi and Field  with minThe value for ELISA cut-off threshold for tested samples at an optimized dilution of all the reagents was determined by taking the mean absorbance (O.D) reading of pre-immune negative sera plus three standard deviations , 82  and Schemallenburg (SBV) viruses were tested using this developed ELISA. This positive BTV and SBV sera samples were available in the virology laboratory of the Universiti Putra Malaysia which were initially collected from confirmed disease cases occurring among some farms.http://www.medcalc.org; 2018).The sensitivity and specificity of the ELISA employed in this study was determined based on the true positive and false negative value subjected to analysis and interepreted from the optimal cut-off point described elsewhere , 82, 83.Information procured for both Sections A and B of the questionnaires was incorporated into Microsoft Office Excel and analysed using JMP software. The likelihood ratio and regression model were used to evaluate the farmer’s compliance level towards HHP. Responses to each sub-unit questions of the 14 major components modules of HHP Table  were expAdditional file 1:Questionnaire. (DOCX 59 kb)


Influenza pandemics considerably burden affected health systems due to surges in inpatient admissions and associated costs. Previous studies underestimate or overestimate 2009/2010 influenza A/H1N1 pandemic hospital admissions and costs. We robustly estimate overall and age‐specific weekly H1N1 admissions and costs between June 2009 and March 2011 across 170 English hospitals. We calculate H1N1 admissions and costs as the difference between our administrative data of all influenza‐like‐illness patients  and a counterfactual of expected weekly seasonal influenza admissions and costs established using time‐series models on prepandemic (2004–2008) data.We find two waves of H1N1 admissions: one pandemic wave (June 2009–March 2010) with 10,348 admissions costing £20.5 million and one postpandemic wave (November 2010–March 2011) with 11,775 admissions costing £24.8 million. Patients aged 0–4 years old have the highest H1N1 admission rate, and 25‐ to 44‐ and 65+‐year‐olds have the highest costs. Our estimates are up to 4.3 times higher than previous reports, suggesting that the pandemic's burden on hospitals was formerly underassessed. Our findings can help hospitals manage unexpected surges in admissions and resource use due to pandemics. We employ time‐series methodologies on data sets of all influenza admissions  models to analyze the universe of patient‐level admissions and costs from 170 public hospitals in England from 2004 to 2011. To our knowledge, this is the first application of time‐series methods to estimate total and age‐specific excess ILI hospital admissions and costs attributable to H1N1 using administrative data, allowing us to substantially improve over existing studies' data and methods. Our study is the first to estimate unit costs based on patient‐level data using all public hospitals within a health system. We compare our findings with other countries' studies to determine the magnitude of bias in their estimates. Our results will help decision makers improve their admission surge response plans for future influenza pandemics.The remainder of this paper is organized as follows: Section 2n = 132,532,270), including age and primary and secondary diagnoses coded according to the International Statistical Classification of Diseases and Related Health Problems, 10th revision (ICD‐10), across all NHS hospitals in England between April 2004 and March 2011. HES is preferred over laboratory‐confirmed data as the latter may not capture a significant proportion of H1N1 patients. Physicians may not have tested for H1N1 because of time constraints, certainty of accurate diagnosis, or they deemed it redundant since H1N1 would warrant similar treatment as other severe influenza. We extract all ILI inpatients with laboratory‐confirmed and unconfirmed H1N1, seasonal and other influenzas, as their primary or secondary diagnosis  following Pebody et al. , developed by the Department of Health Payment by Results (PbR) Team . Every HM = 0.54, SD = 0.003) and unmatched  admissions  are not statistically significant. Matched admissions are significantly younger  than unmatched , t = 4.78, p < 0.001, and have shorter lengths of stay  than unmatched admissions , t = 13.15, p < 0.001. Although this is a small portion of our data set, our cost estimates may be conservative. The final costing data set includes 28,626 admission–tariff matches across NHS hospitals in England between April 2004 and March 2011 to produce a time series of 416 observations of weekly costs of ILI admissions.Tariffs are first matched with patients according to HRG and admission type. Remaining unmatched admissions are matched only by HRG using an unweighted average of each HRG's tariffs across admission types. Of the 29,403 eligible HES admissions, 2.6% of ILI admissions could not be matched to tariffs in any manner and are excluded from our analysis. Differences in gender between matched  as our baseline to establish our counterfactual nonpandemic estimates (November 2008 to March 2011). The counterfactual enabled us to determine when the pandemic (June 2009 to March 2010) and postpandemic (November 2010 to March 2011) periods occurred within this time frame. We use SARIMA models to disentangle the weekly number and cost of excess ILI admissions due to the pandemic from the number and costs we would see in a typical, nonpandemic year. SARIMA, a multiplicative extension of autoregressive integrated moving average (ARIMA), models are used because our data exhibit seasonal winter peaks in ILI admissions. The general ARIMA  model is as follows:Y^t is a forecasted observation of Y, B is the backshift operator  operator is q.We identify excess ILI admissions and costs by fitting multiplicative SARIMA models S = 52 for our weekly data and generate a time series of the same week across prepandemic years to predict typical ILI admissions and costs that should occur in a nonpandemic year. We use an augmented Dickey–Fuller test for stationarity such that our time‐series' mean, variance, and autocorrelation are constant over time. We then determine AR and MA lags using autocorrelation function and partial autocorrelation function plots, respectively, and select the most parsimonious lags combination that derives the minimum Akaike information criterion and Bayesian information criterion. Fourth, goodness‐of‐fit is tested on the SARIMA regression's standardized residuals using the Ljung–Box portmanteau (Q) test of autocorrelation and the Bartlett test of variances to ensure independence and constant mean and variance over time.Following Box et al.'s  model buPandemic weeks are defined as two or more consecutive weeks during the pandemic and postpandemic periods when actual ILI admissions or costs exceed the upper bound of the 95% prediction interval around the SARIMA counterfactual. The residual balance of ILI admissions and costs, calculated as the difference between actual and counterfactual values, is considered excess beyond nonpandemic influenza and is attributed to H1N1.N) SARIMA models as follows:Y^Nt is the counterfactual of ILI admissions in week t.Overall, we built and ran 28 SARIMA models . For eacC) SARIMA models are specified as follows:Y^Ct is the counterfactual of costs for ILI admissions in week t.The costs for ILI admissions  and smallest (8%) groups of total ILI hospital admissions, respectively  with an During the postpandemic period, patients across all age groups saw increases in their excess ILI admission rates compared with the pandemic except 5‐ to 14‐year‐olds, who had the lowest excess admission rate of 0.53/100,000 Table . This riExcess unit costs for ILI admissions are also higher across all age groups during the postpandemic period compared with the pandemic. Those aged 25–44 years old exhibit the highest excess unit cost of £7,422, which is 3.39 times their pandemic‐estimated unit cost. Furthermore, 0‐ to 4‐year‐olds have the second highest excess unit cost, £5,848.76, which is 2.8 times higher compared with the pandemic. Young adults used fewer antiviral medications at this time, which, in combination with an evolved H1N1 virus, could have resulted in more severe and expensive complications for otherwise healthy adults during the postpandemic period  m models and Ljung‐Box and Bartlett test statistics for overall and age‐split excess hospital admissions attributed to H1N1Table S4 Parameter estimates of SARIMA  m models and Ljung‐Box and Bartlett test statistics for overall and age group specific excess costs attributed to H1N1Table S5 Parameter estimates of SARIMA  Click here for additional data file.


PurposeIntracranial aneurysms are relatively common epidemiological problems for which the surveillance, treatment, and follow-up are costly. Although multiple studies have evaluated the treatment cost of aneurysms, the follow-up costs are often not examined. In our study, we analyzed how follow-up costs after treatment affected the overall cost of different endovascular techniques for treating aneurysms.Materials and methodsAn institutional database was used to evaluate the upfront and follow-up costs incurred by patients who underwent elective coiling or placement of a pipeline embolization device (PED) for the treatment of unruptured intracranial aneurysms from July 2011 to December 2017.ResultsA total of 114 patients  were included in the study. There was no significant difference among patients in mean age , sex , American Society of Anesthesiologists (ASA) grade (p = 0.5), discharge disposition (p = 0.1), mean length of stay [3.1 days (±5.5) vs. 2.4 days (±2.6); p = 0.2) or follow-up period [22.7 months (±18.5) vs. 18.6 months (±14.9); p = 0.2). There were no differences in costs during admission (p = 0.5) or in follow-up (p = 0.3) between coiling and PED treatments. Initial costs were predominantly related to supplies/implants (56.1% vs. 63.7%) for both treatments. Follow-up costs mostly comprised facility costs (68.2% vs. 67.5%), and there were no differences in costs of subgroups such as supplies/implants (10.5% vs. 9.4%), imaging (17.0% vs. 17.8%), or facilties between coiling and PED.ConclusionThese results suggest that the upfront and follow-up costs are mostly similar for the treatment of intracranial aneurysms irrespective of whether the providers used coiling or PED endovascular techniques. Hence, we conclude that follow-up costs should not be a deciding factor when considering these treatments. Intracranial aneurysms are a common intracranial disease, with a prevalence of 1-8% in the US and an approximate annual rupture risk of 1% , AnalysiVarious endovascular treatment modalities for aneurysms have been comparatively analyzed for efficacy, complication rates, and patient costs ,8,14-17.Patient populationBefore data collection, the Institutional Review Board (IRB) granted approval with a waiver of consent. Patients initially treated from July 2011 to January 2017 were identified from our institutional database. The inclusion criteria were as follows: elective cases without aneurysm rupture, treatment with coiling or PED, availability of clinical variables, and at least 1 follow-up after the treatment. Patients were identified in the database by Common Procedural Terminology (CPT) codes or International Classification of Disease, Tenth Edition (ICD10) codes, primarily the CPT code 61624 . Patients were also cross-referenced to a prospectively maintained database of endovascularly treated aneurysms. Coiling treatments considered included any embolization performed with stent- or balloon-assisted techniques as well as those performed with primary coiling alone. Follow-up imaging and laboratory studies  were done not based on any standardized protocol but left to the discretion of the treating physician. To verify that patients had unruptured, endovascularly treated aneurysms with adequate recorded data, a retrospective chart review of the electronic medical record was performed. Age, sex, aneurysm location, American Society of Anesthesiologists (ASA) physical status classification system, discharge disposition, length of stay, follow-up length, and costs were collected. The follow-up-length metric was further subdivided into two sections: the date of the last clinical encounter and the date of the last imaging evaluation.AnalysisAn institutional database was used to extract direct initial treatment costs as well as follow-up costs of the aneurysm treatment. This database is a tool designed to assess the direct cost of treatment per patient based on hospital costs . Total cThe standard deviations of the mean (±) or absolute numbers with the percentage of the total were reported as appropriate. A probability value (p) of <0.05 was considered statistically significant. All statistical analysis was done using IBM SPSS Statistics .Patient characteristicsA total of 176 patients with elective aneurysms treated by coiling or PEDs were identified. Of these, 114  patients were included in the final analysis that included adequate follow-up information and cost data  of overall patient treatment costs. The cost of PED was not significantly greater than coiling either at initial treatment or during follow-ups. This study supports a cost-neutral approach towards endovascular treatment, favoring neither PED nor coiling, which may better inform how treatment selection can be done in a cost-conscious manner.Follow-up endovascular costsPrevious studies of endovascular-treatment costs have presented conflicting findings because of the diversity of aneurysm characteristics and treatment types. Coil embolization of ruptured aneurysms is associated with lower upfront costs and follow-up complication rates compared with microsurgical clipping in some parts of the world  . HoweverDirect cost comparison of PED and coiling has been attempted in several studies ,25,28,29LimitationsThe strength of this study is that it represents an analysis of the real-world costs of aneurysm embolization procedures in a busy neurointerventional practice using a well-documented cost database. The coiling and PED techniques were both established and familiar techniques for the surgeons in this practice, who chose the clinically appropriate treatment for each patient/aneurysm. This was further demonstrated in our study of the similar sizes of aneurysms treated with coiling and flow diversion. One limitation of the study is that it was a retrospective single-center study and follow-up times were relatively short in terms of lifetime risk that an aneurysm presents. Thus, the results may not be fit to be generalized to a wider population, different US regions, or a different healthcare system that may have another type of cost structure. Also, the relatively small number of patients included in the study (N = 114) could result in a type 2 error because of the study being underpowered. Future studies should seek to include a larger population across multiple centers to accommodate these potential sources of bias as well as a longer follow-up period. Besides, the lack of a uniform follow-up protocol for coiled or PED aneurysms may have accounted for the wide variability  of follow-up costs. Another limitation of cost studies in endovascular treatment is the inability to perform a direct comparison of coiling and PED cases. Our results showed that internal carotid artery aneurysms were predominantly treated by PED while anterior communicating artery locations were primarily treated by coiling. These differences in indications may affect the cost and are difficult to control.This study evaluated the upfront and follow-up costs incurred by 37 patients who underwent coiling of aneurysms and 77 patients who underwent PED treatment. Overall, the results of this study demonstrate that the upfront and initial follow-up costs for the treatment of intracranial aneurysms remain broadly similar irrespective of whether the providers used coiling or PED endovascular techniques. Upfront costs were predominantly accounted for by device costs, while follow-up costs mostly involved facility fees. Although previous studies have aimed to evaluate whether one type of endovascular treatment is more cost-effective than another, follow-up costs have rarely been included in the equation. Hence, for a large array of aneurysms over a similar period of time, follow-up costs were not shown to be a definitive factor impacting treatment costs. Our results suggest that follow-up costs may not necessarily be a deciding factor for treatment decisions when considering an average aneurysm.


Aedes aegypti and Ae. albopictus mosquitoes. During November 2015–September 2017, a total of 588 travel-associated Zika cases were reported in California, including 139 infections in pregnant women, 10 congenital infections, and 8 sexually transmitted infections. Most case-patients reported travel to Mexico and Central America, and many returned during a period when they could have been viremic. By September 2017, Ae. aegypti mosquitoes had spread to 124 locations in California, and Ae. albopictus mosquitoes had spread to 53 locations. Continued human and mosquito surveillance and public health education are valuable tools in preventing and detecting Zika virus infections and local transmission in California.Zika and associated microcephaly among newborns were reported in Brazil during 2015. Zika has since spread across the Americas, and travel-associated cases were reported throughout the United States. We reviewed travel-associated Zika cases in California to assess the potential threat of local Zika virus transmission, given the regional spread of  We also summarized surveillance for Ae. aegypti and Ae. albopictus mosquitoes in California and laboratory testing for Zika virus during this time.In California, patient testing and evaluation focused on assessment of infection in pregnant women and symptomatic patients, and assessment of potential viremia in these patients in relation to proximity to known https://www.cdph.ca.gov/Programs/CID/DCDC/Pages/CalREDIE.aspx) or through paper case report forms. Cases reviewed by CDPH during November 2015–September 2017 were analyzed for type of Zika disease or infection, as defined by the 2016 Council of State and Territorial Epidemiologists (CSTE)/Centers for Disease Control and Prevention  criteria and classified as confirmed or probable . For Zika case-patients with a travel duration of <6 months, we compared duration of time in Zika-affected areas between pregnant and all other case-patients by using the Kruskal-Wallis test for 2 groups  to retrospectively assess time at risk between these 2 groups.Aedes spp. and other mosquito populations. Mosquito surveillance typically includes trapping and identifying mosquitoes. Surveillance might be augmented by submitting mosquito specimens, especially specimens collected in and around residences or workplaces of case-patients, to the Davis Arbovirus Research and Training Laboratory at the University of California  for Zika virus , which is used to generate statewide data for mapping of Aedes mosquito locations. We used a geographic information system  to generate maps that enabled spatial and temporal mapping of Aedes mosquito populations in relation to presumed places of residence of presumed viremic Zika case-patients. We generated latitude and longitude data by using the Gateway Database for mosquitoes and determined case-patient place of residence by using the California Reportable Disease Information Exchange.California has a network of local vector control agencies that monitors distribution and abundance of <7 days before or any time after return from travel to their place of residence.Testing of humans for Zika virus was performed by the CDPH Viral and Rickettsial Disease Laboratory (VRDL), CDC, local public health laboratories, commercial laboratories, and blood banks. Testing for Zika virus infection was completed for appropriate tissue, serum, or urine specimens by using Zika virus nucleic acid or serologic tests. We analyzed symptomology and pregnancy status of those tested, volume of testing at the CDPH VRDL, types of tests conducted, and time from symptom onset to specimen collection date. For purposes of local transmission risk assessment, a potentially viremic patient was defined as a Zika-positive case-patient with symptom onset >1 of the following: fever, rash, arthralgia, or conjunctivitis); 112 had a symptomatic noncongenital Zika infections, 6 had congenital Zika disease with Zika-associated birth defects ; and 4 had congenital Zika infections with no Zika-associated birth defects  of case-patients were female; median age of case-patients was 35 years (range <1–89 years). Of persons with reported ethnicity, 69% (306/443) were Latino/Latina. For the 139 women pregnant at the time of diagnosis, median age was 27 years (range 14–44 years), and 78% (87/111) of those with reported ethnicity were Latina.Of 570 case-patients who contracted Zika virus while traveling outside California, most case-patients reported travel to Mexico (36.4%), Central America (34.3%), or the Caribbean (13.1%). The top 10 countries and territories for travel were Mexico (36.4%), Nicaragua (9.6%), Guatemala (8.4%), El Salvador (7.0%), Dominican Republic (4.4%), Puerto Rico (4.4%), Honduras (3.9%), Costa Rica (3.7%), Jamaica (2.5%), and Colombia (1.8%). The timeline for travel-associated Zika cases reported in California mirrored the spread of the outbreak across the Americas ; the num>6 months before coming to California, where they were subsequently tested for Zika virus. When we excluded these 79 persons, women who were pregnant at the time of Zika diagnosis had a significantly (p = 0.03) longer travel duration (median 14 days [range 1–153 days]) than all other Zika case-patients (median 11 days [range 1–137 days]).Of 570 case-patients who traveled, 79 (13.9%) lived in their country of exposure for For 466 case-patients with symptoms, rash was the most common , followed by arthralgia , fever , myalgia , and conjunctivitis . A rash without any other symptom was seen in 49 (10.5%) case-patients. For those case-patients with >1 symptom, the most common combination of symptoms, reported by 13% of case-patients, was rash, arthralgia, and fever. Seven case-patients were hospitalized for a median of 3 (range 1–8) days. On the basis of symptom onset date, the number of Zika cases reported in California in 2016 increased from June through August and then decreased through November .Of 139 women who were pregnant at the time of Zika diagnosis, 120 had completed their pregnancies by September 1, 2017: 114 with live births and 6 with fetal losses. Fourteen women were still pregnant, and the status of 5 women was unknown. For live births, 90 (78.9%) infants were tested for Zika virus at or shortly after birth; 84 (73.7% of live births) infants showed negative results for Zika virus by nucleic acid and IgM tests, and 6 (5.3% of live births) showed positive results. Of the remaining 24 live births, 7 infants were not tested, and the testing status of 17 infants was unknown. In addition to the 6 congenitally infected infants that were born to Zika virus–positive mothers, 4 additional infants whose mothers were exposed to Zika virus but showed negative results by nucleic acid or IgM tests were positive for Zika virus.Eight infants were born in California with Zika-associated birth defects. Of these infants, 2 were negative and 6 were positive for Zika virus by PCR and IgM test. Both Zika virus–negative infants had mothers who were positive for Zika virus, and 3 of the Zika virus–positive infants had mothers who had negative results for Zika virus.Ae. aegypti mosquitoes and 25 new locations for Ae. albopictus mosquitoes, for a total of 133 cities or census-designated places for Ae. aegypti mosquitoes and 56 for Ae. albopictus mosquitoes, an increase of 142% for Ae. aegypti mosquitoes and 81% for Ae. albopictus mosquitoes in 20 months. In 2017, Ae. aegypti mosquitoes were detected in 12 counties and Ae. albopictus mosquitoes in 5 counties, including 2 new counties containing Ae. aegypti mosquitoes in the Central Valley  were likely viremic while in California. Of those viremic case-patients, 279 (64.1%) were also residents of California counties where Ae. aegypti or other Aedes spp. mosquitoes have been detected; their co-location was more common in southern California  testing for Zika-positive cases in California, commercial laboratories accounted for 17.5% (103/588), local health departments for 13.1% (77/588), and CDC for 9.5% (56/588). Seven Zika cases reported in California were identified through blood bank screening. Most testing at VRDL was performed for asymptomatic pregnant women . Eighty (1.0%) of these asymptomatic pregnant women were positive for Zika virus by quantitative RT-PCR (1 woman) or IgM test and plaque-reduction neutralization test (PRNT) (79 women).Of the 120 completed pregnancies for women who were infected with Zika virus while pregnant, 45 placental tissues  were sent to CDC for testing. Zika virus was detected by RT-PCR in placental tissues of 8 women. Detection of Zika virus in the placental tissue provided confirmatory testing for 5 of these women .Of the 410 confirmed Zika cases reported in California, 319 (77.8%) case-patients had Zika virus detected by nucleic acid tests in serum, urine, or placental tissue, and the other 91 were confirmed by detection of neutralizing antibodies to Zika virus (and not DENV). For symptomatic case-patients, the median time from illness onset to specimen collection was 5 days (range 1–194 days). For cases confirmed by serum or urine nucleic acid tests, the time to collection was shorter, with a median of 3 days (range 1–33 days), than for PRNT, with a median of 16 days (range 1–194 days).Ae. aegypti and Ae. albopictus mosquitoes in California, prevention of local transmission of Zika virus has been and continues to be a public health priority. In working to identify possible local transmission, CDPH used the data for travel-associated Zika cases described in this article to develop our Zika testing prioritization. Although CDC recommended specific criteria for travel-associated Zika virus testing, different criteria were needed when testing persons without travel history, especially when the number of confirmed Zika cases was increasing in California and local Zika virus transmission was reported in Florida , without an alternative explanation, such as a drug reaction or other infection. Rash was recommended as the primary criterion in this setting because nearly 90% of Zika case-patients had a rash. This allowance for Zika virus testing for persons with no travel or sexual exposure was used in some counties in California and identified several persons suspected of having Zika who were tested, all of whom showed negative results. This testing allowance would not be appropriate in areas that did not contain Aedes spp. mosquitoes and is being reconsidered as the number of Zika cases has decreased.CDPH subsequently provided criteria for local health departments in California to consider in evaluating whether suspected persons without travel history should be considered for Zika virus testing, including factors that could increase risk for local transmission, as well as signs and symptoms most suggestive of Zika. For example, CDPH allowed that, for counties where Ae. aegypti and Ae. albopictus mosquitoes, especially in southern California, as has also been found for dengue and chikungunya  and to limit potential spread.Although California health officials did not identify any episodes of local Zika virus transmission, our data indicate that large numbers of likely viremic travelers returned to areas containing Aedes spp. mosquitoes can be found year-round in California, they are most abundant from June through November, typically peaking in September and October . Third, an estimated 80% of persons infected with Zika virus are asymptomatic (Ae. aegypti and Ae. albopictus mosquitoes into 12 counties in California, especially along the southern border region, increases the risk for local Zika transmission in California. The large percentage of potentially viremic travelers returning to areas that contain Aedes spp. mosquitoes, in addition to an unknown number of returned travelers who are asymptomatically infected but not detected, makes the risk for local transmission a continuing threat, albeit low, in California. Zika has complicated disease manifestations and transmission dynamics, such as sexual and congenital transmission, which are not typically observed for other arboviruses. It is vital that we apply the public health lessons learned during the Zika outbreak to prepare for complexities that might arise during future epidemics of emerging and reemerging arboviruses.Although Zika virus transmission and Zika case numbers have decreased across the Americas, we expect to see continued, limited, local transmission in some affected countries. Thus, there is still a risk for pregnant women and all those who travel to these countries, and it is necessary that prevention messaging remains targeted and operative. Healthcare providers should continue to be suspicious of returning travelers with rash, fever, conjunctivitis, or arthralgia, particularly when other diagnoses have been ruled out. The expansion of


Terpolymer raspberry vesicles contain domains of different chemical affinities. They are potential candidates as multi-compartment cargo carriers. Their efficacy depends on their stability and load capacity. Using a model star terpolymer system in an aqueous solution, a dissipative particle dynamic (DPD) simulation is employed to investigate how equilibrium aggregate structures are affected by polymer concentration and pairwise interaction energy in a solution. It is shown that a critical mass of polymer is necessary for vesicle formation. The free energy of the equilibrium aggregates are calculated and the results show that the transition from micelles to vesicles is governed by the interactions between the longest solvophobic block and the solvent. In addition, the ability of vesicles to encapsulate solvent is assessed. It is found that reducing the interaction energy favours solvent encapsulation, although solvent molecules can permeate through the vesicle’s shell when repulsive interactions among monomers are low. Thus, one can optimize the loading capacity and the release rate of the vesicles by turning pairwise interaction energies of the polymer and the solvent. The ability to predict and control these aspects of the vesicles is an essential step towards designing vesicles for specific purposes. They are important because they can be used as nanoreactors or carriers ,2,3,4. W(S/tBMA) , [poly(3(S/tBMA) , and [po(S/tBMA)  have sucThe variety and properties of aggregates formed by ABC terpolymers are related to their structures. Star ABC terpolymers are block copolymers composed of blocks connected at one common point. The blocks are mutually immiscible and have different solvophobicities. The strong repulsion among the blocks causes ABC terpolymers to self-assemble into aggregates with segregated A, B, and C nanodomains. These aggregates have a variety of structures, many of which cannot be observed in solutions of linear copolymers. Polygonal bilayers , nanostrThe equilibrium structures formed by star ABC terpolymers in selective solvents depend on concentration ,15,16, iThe length of individual blocks of block copolymer determines the morphology and stability of the equilibrium aggregates ,17,22,2312B6C2 as the model polymer. The chosen length ratio corresponds to a solvophilic weight fraction of 35% (±10%) and is known to promote vesicle formation [It has been shown that miktoarm star ABC terpolymer with one longer solvophobic arms but shorter solvophilic arms can self-assemble into spherical vesicles in a wide range of solvent conditions . The sysvesicles ,22,23. Sormation . As a reUsing our model system, the equilibrium structure of the aggregates is explored using dissipative particle dynamics (DPD) simulations. Effects of the interactions among beads and solvents and that of polymer concentration are investigated. Surface morphologies and structures of aggregates are examined. The thermodynamics that governs the equilibrium aggregate structures is discussed.As vesicles formed with star terpolymers can be efficient carriers, it is necessary to identify conditions under which they are the equilibrium structures and to study how their encapsulation efficiency and properties vary with their morphological details and with the environment. A state diagram showing the equilibrium aggregate structure is constructed to ascertain the optimal conditions for vesicle formation. The ability of the aggregates to encapsulate solvent is explored. This paper sheds light on how polymer concentration and interaction energies among various blocks of a terpolymer and with the solvent govern the equilibrium aggregate structures and their properties. The ability to predict and control these aspects of the vesicles is an essential step towards designing vesicles for specific purposes .The remainder of the paper will present the methodology applied and the major findings of this study. Note, figures, and tables denoted by the letter “S” can be found in the accompanying Dissipative particle dynamics (DPD) simulations are used to study the spontaneous self-assembly of ABC star terpolymers in a solvent S. DPD is a particle-based mesoscopic simulation technique first introduced by Hoogerbrugge and Koelman . The terIn our simulations,  to be 3  and tempd Warren , the weiThis restricts the interaction among beads only with their neighbours. The dissipative force as 100.0 , and theThe integration of the equation of motion (Equation (1)) is carried out using the Velocity-Verlet algorithm  with a tThe interactions among blocks are varied with a coupling parameter λ:The coupling parameter The volumes of the repeat unit of PEE, PEO, and PFPO are 122.5, 65.2, and 149 12B6C2 star terpolymer in solutions made of S beads. Each polymer chain has 20 beads which consists of 12 A beads, six B beads, and two C beads. Unless explicitly stated, results presented are from simulations consist of 192,000 DPD beads in a simulation box of 40The present DPD simulations focus on the effect of polymer concentration on the spontaneous vesicle formation of AFree energies are evaluated to identify the factors driving the aggregation of the terpolymers into micelles and vesicles. The free energy The integrand The majority of the analysis carried out in this work concerns properties of surfaces of aggregates, which we refer to as “skin”. Take a vesicle as an example: a vesicle is made of a membrane, with a cavity inside. The membrane is made of three parts; the inner shell, which is adjacent to the cavity; the outer shell, which is the surface exposed to the surroundings; and the inner leaflet, which separates the two shells see . The outThe aggregate size 12B6C2 star terpolymers form fragmented aggregates at low The equilibrium aggregates formed at various values of h φp see . In thesh φp see  reside oAt Similar structures are observed for intermediate values of The equilibrium structures at high λ have been observed both in experiments  and simuterature . The netTo illustrate the effect of interactions among various beads in the star terpolymer and the solvent, equilibrium structures for various At very low λ, where interactions among all beads and the solvent are equal or relatively small, the solvent and the terpolymer mix well  of aggregates can be characterized by their principal moments of inertia The relative total free energy maps see . Note thThe dominant contribution to the free energy for all clusters is Further insight into the contributions to the free energy can be gained from Various combinations of Information about aggregate sizes (excluding hydrated layers), in the form of average radii r chains . Howeverth λ see . Note, aComposition profiles, showing how fractions of various beads Vesicles contain S beads in their cavities. Their inner and outer shells have properties generally similar to shells of micelles  of test chains for vesicles formed at iffusion  can be oIn this work, interactions energies between B-B beads, B-S beads, and S-S beads are similar see . Hence, The degree of B-bead solvation can be examined by the length of the solvated B-bead segments extending from the outer skin. The solvation of B-beads of the inner skin is excluded in this analysis because they might be affected by a confinement effect. The average solvation length and its distribution under various conditions are shown in long see . As φp dVesicles are frequently cited as potential cargo carriers, so their ability to encapsulate target chemicals at intended domains is important. In this work, S-beads are encapsulated at the cavity of the vesicles. The amount of S-beads encapsulated normalized by the number of terpolymer chains making up the vesicle is presented in Assuming that the vesicles are spherical for ores see . Since tin films . Hence, The presence of S-beads in the inner A-bead leaflet of the membrane with low Increasing 12B6C2 star terpolymers in solutions made of S beads are the model systems. The effects of concentration and strength of interactions are investigated. When the concentration of polymer In this work, the equilibrium structures of an ABC star terpolymer are examined with DPD simulations. AIn terms of encapsulation of S-beads, vesicles made of terpolymers having lower interaction energies with solvents are more efficient in terms of solvent encapsulation capability. This is because their membranes are swollen by the solvent. As a result, these membranes can be stretched more easily. The thinner membrane for these vesicles means they have more encapsulation volume per terpolymer. With more S-beads encapsulated, these vesicles are also slightly larger and have thicker hydration layers. The high efficiency for S-bead encapsulation of these vesicles comes with a price. The fact that solvent can swell the membrane of these vesicles relatively easily means encapsulated molecules can permeate through the membrane. Hence, the repulsion energy among beads should be tuned carefully to design vesicles of desired encapsulation efficiency and release rate.


We therefore demonstrate the potential of the inexpensive and efficient hydrogel electrode to replace conventional gold-cup electrodes for the recording of VEPs and possibly other recordings from the human cortex.The aim of this study was to investigate the use of inexpensive and easy-to-use hydrogel “marble” electrodes for the recording of electrical potentials of the human visual cortex using visual evoked potentials (VEPs) as example. Top hat-shaped holders for the marble electrodes were developed with an electrode cap to acquire the signals. In 12 healthy volunteers, we compared the VEPs obtained with conventional gold-cup electrodes to those obtained with marble electrodes. Checkerboards of two check sizes—0.8° and 0.25°—were presented. Despite the higher impedance of the marble electrodes, the line noise could be completely removed by averaging 64 single traces, and VEPs could be recorded. Linear mixed-effect models using electrode type, stimulus, and recording duration revealed a statistically significant effect of the electrode type on only VEP N75 peak latency (mean ± SEM: 1.0 ± 1.2 ms) and amplitude (mean ± SEM: 0.8 ± 0.9 µV) The mean amplitudes of the delta, theta, alpha, beta, and gamma frequency bands of marble electrodes were statistically significantly different and, on average, 25% higher than those of gold-cup electrodes. However, the mean amplitudes showed a statistically significant strong correlation (Pearson’s  Visual evoked potentials (VEPs) are changes of the electrical potential elicited by visual stimuli and recorded using electrodes mounted on the forehead and the scalp above the inion. VEPs are part of electroencephalogram (EEG) and are extracted by stimulus correlation and averaging. VEPs are used to measure the functional integrity of the visual pathways from retina via the optic nerves to the visual cortex . A typicRecording of visual evoked potentials is usually carried out using gold-cup electrodes. To obtain a good signal-to-noise ratio , extensiThe aim of this study was to explore the use of “water beads”, a commercially available hydrogel formulation that is also known as marble electrodes, for the recording of VEPs. The hydrogel, made of an acrylic sodium salt of cross-linked polyacrylic acid, is superabsorbent and inexpensive, and its use as an electrode avoids the lengthy preparation normally involved in VEP recordings. A comprehensive review of superabsorbent polymers, their properties, and their applications has been given by several authors -]n), a hydrogel, which is able to absorb water up to 500 times its weight. After swelling, the beads consist of up to 99.9% water  and therefore similar to that of tap water. Water beads are commonly used for watering plants or for decoration purposes and are available in home improvement stores and garden centers as well as online for about €2 per 1000 pieces. The marble electrodes are always damp but, in contrast to a sponge, they do not loose water when squeezed. The stiffness of a marble electrode is about κ = 0.4 N/mm.Water beads are made of an acrylic sodium salt of cross-linked polyacrylic acid (PAA)  in a workshop of the University Eye Hospital, Tuebingen. The inner diameter of the top hat corresponds to the diameter of the marble electrodes, and its height is about half the diameter of the marble electrode. A conventional gold-cup electrode clipped into the upper end of the hat (opposite to the brim) connects the leads with the marble electrode b. These The holder and the skin electrode can be reused, while the marble electrode can be disposed of after use. The marble electrodes were soaked in pure water for about six hours, until they were swollen to their maximal size of about 1 cm in diameter.Twelve healthy volunteers  with best-corrected visual acuity and no history of eye or neurological diseases were recruited from the staff of the Centre for Ophthalmology of the University of Tuebingen. All volunteers gave informed consent. The study followed the tenets of the Declaration of Helsinki and was approved by the Institutional Review Board of the Faculty of Medicine, University of Tuebingen.The VEP recordings were performed monocularly with one eye covered with an eye patch. The checkerboard stimulus was presented using a 21” CRT monitor   at a disOz, reference electrode at Fz, and ground electrode at Cz.Electrodes were mounted according to the International 10–20 system : active In the first session, marble electrodes were used as active, reference, and ground electrodes. No skin abrasion was performed. In the subsequent second session, the skin was cleaned using abrasive paste, and gold-cup electrodes were applied using conductive paste.2  with a sampling frequency of 1000 Hz and digitally band-pass filtered (1.25–100 Hz). No notch filter was used. Post-trigger time was 300 ms. Automated baseline correction was applied by averaging and subtracting a 20 ms pretrigger period. Three averages, consisting of 64 single sweeps, were recorded for each check size , t(125) = 0.9845, p = 0.3268; peak time: mean (marble) = 75.49 ms, mean (gold-cup) = 74.52 ms, SEM = 1.23, difference = 0.97 ms, 95% CI: , t(125) = 0.9675, p = 0.3352).Post hoc comparisons of least square means of the N75 amplitudes and peak times using the Student’s Visual evoked potentials were successfully recorded in 12 volunteers for two check sizes using marble electrodes and conventional gold-cup electrodes. The marble electrodes had about 10 times higher impedance compared to the gold-cup electrodes.+ and Cl− ions. However, in the saline solution, the swelling ability was drastically reduced. This effect was also shown by Horkay et al. [To reduce the impedance, we tried using saline solution instead of water for soaking the hydrogel in order to increase its electrical conductivity by adding Nay et al. , who inv2 system used in this study has an input impedance of one gigaohm. A high impedance of the electrode may cause only problems when using old amplifiers with an input impedance of less than 100 megaohms. This is in line with the findings of Ferree et al. [Even though ISCEV recommends keeping the electrode impedance lower than 5 kΩ ,33, thise et al.  and Kappe et al. , who fouFz compared to Oz is probably the result of a worse contact at Oz, caused by dense hair [Oz may have been lower than at Fz, resulting in a smaller contact area of the marble electrode, which is related to an increase in noise [The statistically significant lower impedance of the marble electrodes mounted on nse hair . Combingnse hair . Furtherin noise .Fz and Oz is likely the reason for picking up of line noise, which caused a contamination of the single sweeps with a 50 Hz signal. Line noise usually results from electrodes with high impedance, as is the case with marble electrodes, in combination with a differential amplifier [The difference in the impedance between mplifier .The increased noise caused by the higher impedance of the marble electrodes may have also led to the statistically significant higher mean amplitudes in the Fourier analysis of the different frequency bands of the recordings compared to those of conventional gold-cup electrodes. On average, the mean amplitudes recorded using gold-cup electrodes were between 67% and 82% lower than for marble electrodes. However, the mean amplitudes of the different frequency amplitudes obtained with marble electrodes showed a statistically significant high correlation to those recorded with conventional gold-cup electrodes.Even though the single traces were strongly contaminated with line noise, these artifacts could be removed almost completely using averaging and therefore only had a small effect on the measured amplitudes and peak times of the VEP. Linear mixed-effect models revealed a good agreement between the cursors obtained from recordings of the marble electrodes and the conventional gold-cup electrodes. A statistically significant difference was found for the N75 amplitude and the peak time between recordings using marble electrodes and conventional gold-cup electrodes. For the P100 peak time and amplitude, the interaction between check size and electrode type showed a statistically significant effect. A statistically significant effect of the recording duration was only found as an interaction with the check size and with the electrode type for the P100 peak time. All effects, except check size, reached statistical significance only because the alpha level was raised to 0.5. Furthermore, the mean differences in amplitudes and peak times between the conventional gold-cup electrode and the marble electrode were in the range of the intrasubject variability published by several groups ,37,38,39This study demonstrated the potential of marble electrodes to replace conventional gold-cup electrodes for the recording of visual evoked potentials. Using modern differential amplifiers and averaging, high-quality VEP recordings were obtained without scalp abrasion. The differences in the amplitudes and the peak times between conventional gold-cup and marble electrodes were within the range of intrasubject variability and therefore of no clinical relevance. The ease of use and their low cost may render marble electrodes useful for other application domains, such as simultaneous recordings of the electroencephalogram during functional magnetic resonance imaging, brain–computer interfaces, or transcorneal electrical stimulation. However, further studies are needed to evaluate such applications, especially with regard to the differences in the amplitudes of the Fourier spectrum in the different frequency bands.In a previous study, we demonstrated the application of marble electrodes for the recording of electroretinograms in small animals . Additional uses for the application of marble electrodes may be brain–computer interfaces. This is because, in contrast to currently used electrodes, hydrogel-based electrodes, such as marble electrodes, provide a higher wearing comfort and are better tolerated, as Pinegger et al. investigated [As marble electrodes consist of up to 99% water, they do not dry out, rendering them useful for long-time recordings, e.g., for brain–computer interfaces or during functional magnetic resonance imaging.Avoiding the need for cleansing and abrasion of the skin increases patient comfort and significantly reduces the time for preparation. Additionally, it eliminates the risk of infection during abrasion of the skin . BecauseMarble electrodes may also be used for transcorneal electrical stimulation . SeveralT.S., T.P., and E.Z. hold a patent on using this type of hydrogel electrodes for electrophysiological applications .


The economic burden of ulcerative colitis (UC), specifically related to indirect costs, is not extensively documented. Understanding and quantifying it is required by health care decision makers.To assess the impact of indirect costs of UC in observation studies.A systematic literature search was conducted in MEDLINE®, Embase® and Cochrane Library to capture all relevant publications reporting outcomes on absenteeism, presenteeism and productivity losses in moderate to severe UC. Eligibility criteria for inclusion into the review were established using a predefined PICOS scheme. All costs were adjusted to 2017 currency values .In total, 18 studies reporting data on indirect costs were included in the analysis. Absenteeism costs were classified into three categories: sick leave, short-term and long-term disability. Most of the studies captured absenteeism costs related specifically to sick leave, which was experienced on average by 10 to 24% patients with UC. Only three studies captured presenteeism costs, as these are difficult to measure, however costs ranged from 1602 $ to 2947 $ per patient year. The proportion of indirect costs accounted for 35% of total UC costs .A limited number of studies were identified describing the indirect costs in patients with moderate to severe UC. Insufficient data on different components of costs allowed a limited analysis on the impact of indirect costs in patients with UC. Further studies are needed to gain an understanding of the influence of UC on patients’ functional abilities. Ulcerative colitis (UC) is a chronic disease that is characterized by diffuse mucosal inflammation limited to the colon , 2. In aThe management of UC has changed with the approval of new biological therapies such as infliximab, which was approved by the FDA 2005) and by the EMA 2006) for the treatment of moderate to severe UC [ and by t for the Apart from the clinical repercussions associated with this disease, UC also has a societal burden on patients and their caregivers. On a global scale, this disorder accounts for a quarter million physician visits, 30,000 hospitalizations, and more than a million workdays missed annually . The dirvConclusions drawn from clinical trials are not always sufficient for decision makers, as they assess the value of a specific drug in a controlled setting. However, real word studies that collect data beyond Phase III controlled trials  allow decision makers to better manage and understand uncertainties, specifically related to epidemiology, compliance, adherence and cost insights . The aimAs previously defined in the publication by Kawalec et al.  indirectThis review was conducted to identify studies that report indirect costs in ulcerative colitis. The protocol for this review was not registered.The electronic databases Embase®, MEDLINE® and Cochrane Library were searched on the 22nd of May 2017 to capture studies reporting outcomes on absenteeism, presenteeism and productivity losses. Search terms included the following medical subject headings ‘ulcerative colitis’/exp. OR ‘ulcerative colitis’ OR ‘inflammatory bowel disease’/exp. OR ‘inflammatory bowel disease’ OR ‘ibd’ as well as cost subject headings such as ‘indirect’ OR ‘productivity’ OR ‘economic’ OR ‘cost’ OR ‘loss’ OR ‘burden’ OR ‘human capital’. Additional hand searches were performed to identify studies published in important medical societies such as United European Gastroenterology Week (UEGW), European Crohn’s and Colitis Organization (ECCO) and Digestive Disease Week (DDW).I)Population: Adult patients with a confirmed diagnosis of activeII)Intervention/ Comparators: No restrictions were applied to these two parameters.III)Outcomes: indirect costs or productivity losses associated with absenteeism and/or presenteeism were considered of interest.IV)Study type: Real Word Evidence (RWE) data, meaning observational studies, systematic literature reviews, cost estimation studies and cost effectiveness studies written in English and published between 1st of January 2006 (availability of anti-tumor necrosis factor drugs) and 22nd of May 2017 were included.For inclusion in this review, studies needed to fulfill specific criteria in accordance with a predefined PICOSA publication was excluded if it did not fulfil the abovementioned inclusion criteria, meaning if the study did not report any kind of indirect costs or if the patient population did not have confirmed moderate to severe active UC. Randomized controlled trials, , long-term extensions of clinical trials and studies with rules of protocol violation, as well as crossover trials and interventional cohort studies beyond the scope of public health interventions were excluded from this review. Publications that were not written in English were also excluded.To assess the eligibility of a study, two authors independently examined titles and abstracts identified from the search strategy. Articles, which have been identified as potentially relevant based on title and abstract, were then reviewed in full text and selected according to the list of pre-specified inclusion/exclusion criteria. All discrepancies were solved by discussion. If no agreement was found, a third reviewer was involved in the discussion and final decision making.As most of the studies extend across a wide time frame, all costs were converted to 2017 American dollars ($), using country specific consumer price index from the worldwide inflation data source.This review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement to ensure that all records were well tracked .Data from the eligible studies were collected, including publication details, specifications of the study question , methodology used, main indirect costing results , as well as limitations associated to the study. Results were then tabulated and analyzed using descriptive statistics. Data extraction was carried out by three researchers and quality control has been done for at least 20% of extracted data, as defined in the study protocol.In order to achieve comparability of the results across publications, we attempted to break down indirect costs into the same categories (e.g. sick leave or short- and long-term disability). We also reported cost data per patient per year and assumed that costs are stable throughout the year.Descriptive statistics were reported for the study outcomes for all patients to describe the basic features of the data in the identified studies. Univariate, bivariate and multivariate analyses were not conducted to assess the association of baseline characteristics with cohorts nor patient characteristics with indirect costs.After having applied a search strategy for indirect costs in UC, 18 studies were incorporated in the narrative review, as outlined in Fig. Data on indirect costs were collected from web-based questionnaire or through database analysis 17 studies), one study collected data through patient cost diaries. The majority of studies had large sample sizes, ranging from 53 to 6900 patients, as reported in Table 7 studiesOverall, 13 studies out of 18 studies reported data from nine different countries (mainly in Europe), on absenteeism in UC, capturing cost/loss of earning of absenteeism per patient year and/or total average indirect costs of absenteeism per patient year. With regards to presenteeism, we were only able to identify three studies, one conducted in Poland , one in Throughout the identified studies, we noted that absenteeism costs were often classified into categories; either (1) costs associated to sick leave Table , and/or While some publications specifically define the components associated with absenteeism, there were few publications such as Cannon et al.  that didThe proportion of absenteeism in ulcerative colitis patients was reported in two different countries the USA and Spain. The percentage of patients experiencing absenteeism was high, even if data was limited, ranging from 85 to 98% as reported by Gibson et al.  and CoheThe average number of absenteeism days per patient year varied widely, ranging from 1.08 to 60 days per patient year. The low rate of 1.08 days reported in Spain by Aldeguer et al.  is relatThe identified publications also report costs associated to absences from work ranging from 3197 $ to 7794 $ per patient year. Here again, the study by Aldeguer et al.  reportedEight studies out of 18, all conducted in Europe, reported absenteeism costs related specifically to sick leaves in UC, as shown in Table Five , 25, 28 Three studies estimated indirect costs associated to presenteeism, one in Poland , one in UC patients were poorly described in the publication by Kawalec et al.  and in KConsistent results were found when comparing presenteeism data, as all reported that patients with UC experience a 19% job productivity loss per week, with presenteeism costs ranging from 1602 $ to 2947 $ per patient year.Total productivity losses were reported in four studies, one in Poland , one in The chronic nature of UC as well as the way the disease evolve over time makes it a costly condition to manage. Typically, costs of hospitalizations, surgery and the management of its complications are drivers of direct medical cost. Given the disease’s epidemiological characteristics and age distribution, the indirect costs due to productivity losses further contribute to high overall total disease costs. The objective of this review was to understand the impact of UC on indirect costs.n = 34) therapies with patients who underwent surgery, either J-pouch (n = 81) or Ileostomy (n = 48). Results indicated that indirect costs were highest in patients taking anti-TNF therapies (4340€ per patient year and 28 days of sick leave per year) compared to surgical patients (with 1508 $ per patient year and 20 days of sick leave per year for ileostomy and 1932 $ per patient year and 8 days of sick leave per year for pouch patients). However, little is known on the long-term cost trends of these interventions. In fact, surgery has typically been considered as a significant cost driver in UC patients, mainly due to the need for hospitalization as well as the risk for complications after the procedure. A recent review by Lindsay et al. [Understanding and quantifying indirect costs associated to biologics versus surgery was possible through one study by Van Der Valk published in 2015 , that coouch n = 1 or IleoAs healthcare systems vary greatly between countries, it was expected to identify variations in indirect costs between Europe and the USA. Giving that the majorities of identified publications reported indirect costs in Europe, it was difficult to quantify this difference. However, we noted that costs related to absenteeism were greater in the USA compared to Europe. Productivity losses were only reported in three studies. Although limited information was found, it was possible to conclude that, on average, 35% of total UC costs were associated to indirect costs. This proportion remains uncertain, as indirect costs are very difficult to assess, mainly because of difficulties in measurement, especially when talking about presenteeism (i.e. the reduction of workers’ effectiveness due to illness).n = 1728) had significantly (p < 0.0001) higher hospitalization rates (26.5% vs 6.2%) and adjusted total direct  and indirect costs (5666 $ vs 1960 $) [This review allowed us to conclude that costs in UC tend to be highly variable based on the subpopulation to which they refer. In fact, Cohen et al.  reported 1960 $) . Similar 1960 $)  and Hils 1960 $)  who repoThis review has several limitations. No population-based studies with control patients were included in our analysis and our findings may not be applicable to all patients with UC. After completion of our analysis, results from a prospective study was published describing data for patients with UC and CD from the Danish national registry . This poWithin the small number of studies that reported indirect costs in UC, heterogeneity of reporting data was a key limitation in analyzing and quantifying the impact of indirect costs in UC. Analysis was confounded by differences in costing components within the same category and across different cost categories, due to the lack of definition of these components. There remains uncertainty in the cost associated with absenteeism based on the limited information provided in the primary publications. A recent review by Kawalec et al.  concludeCosts were adjusted to the 2017 USD values to allow comparison between countries; however, these may not be comparable across different societies. Other limitations include: heterogeneity in the inclusion criteria .In conclusion, the findings of this review showed that indirect costs in UC are not well documented in the literature. Therefore, additional studies are needed in UC to quantify these costs per disease state and to evaluate the impact of the disease on patients’ functional abilities.


Reoviridae). BTV serotypes 1–24  are transmitted by competent biting Culicoides midges and replicate in mammalian (BSR) and midge (KC) cells. Previously, we showed that genome segment 10 (S10) encoding NS3/NS3a protein is required for virus propagation in midges. BTV serotypes 25–27  do not replicate in KC cells. Several distinct BTV26 genome segments cause this so-called ‘differential virus replication’ in vitro.Transmission of vector-borne virus by insects is a complex mechanism consisting of many different processes; viremia in the host, uptake, infection and dissemination in the vector, and delivery of virus during blood-feeding leading to infection of the susceptible host. Bluetongue virus (BTV) is the prototype vector-borne orbivirus . BTV11 with a small in-frame deletion in S10 infected blood-fed midge midguts but virus release from the midgut into the haemolymph was blocked. BTV11 with S1[VP1] of BTV26 could be adapted to virus growth in KC cells, and contained mutations subdivided into ‘corrections’ of the chimeric genome constellation and mutations associated with adaptation to KC cells. In particular one amino acid mutation in outer shell protein VP2 overcomes differential virus replication in vitro and in vivo.A 100 nl blood meal containing ±10Culicoides midges can strongly differ for very closely related viruses.Small changes in NS3/NS3a or in the outer shell protein VP2 strongly affect virus propagation in midges and thus vector competence. Therefore, spread of disease by competent  Arthropod-borne viruses have a significant social and economic impact on both human and animal health. The majority of emerging and re-emerging infectious diseases are vector-borne and/or zoonotic , 2. VectCulicoides midges (Diptera: Ceratopogonidae). Historically, BT is caused by 24 serotypes of bluetongue virus (BTV)   of atypical BTV26 were evaluated for their ability to replicate in mammalian and Culicoides cell lines and in C. sonorensis midges. Effects of viral genetics on vector competence is discussed.To further investigate differential virus replication Culicoides (KC) cells were grown in modified Schneider’s Drosophila medium with 15% heat inactivated FBS, 100 IU/ml penicillin and 100 μg/ml streptomycin at 28 °C . A virus stock was obtained by one passage on BSR cells at Wageningen Bioveterinary Research (WBVR) and designated BTV26. BTV11 was isolated from the spleen of a white-tailed deer from Texas in 2011, passaged once in embryonated chicken eggs, and four times in BHK21 cells before use in midge feeding/injecting. A virus stock for in vitro experiments was obtained by one passage on BSR cells at WBVR, and designated wtBTV11. All other viruses in this study were generated by reverse genetics , containing the same BTV11 as above (S111) and BTV26  sequences were designed and purchased. Each chimeric S1 contained one of three defined domains of the RdRp of BTV26 (S111/26) 26, S2[VP2]11, S4[VP4]11, and S8[NS2]11. Subsequent virus passages on BSR cells (p6) resulted in three aa mutations; one in S1[VP1]26, two in S8[NS2]11, and one incomplete aa mutation in S4[VP4]11, as well as one incomplete silent mutation in S2[VP2]11. These five nucleotide changes were also found after ‘rescue’ of BTV11(S126) in KC cells (p3r) but four aa mutations were not completely changed immediately after ‘rescue’ in BTV11(S126)kc(r). Apparently, these aa mutations were rapidly selected in both cell types and are likely associated with chimeric corrections in BTV11(S126). In addition, one incomplete E321G aa mutation in VP211 was found. After three subsequent virus passages (p3r+3p), BTV11(S126)kc contained three additional incomplete mutations, whereas the incomplete E321G aa change in VP211 was complete (Table 26 was complete after three subsequent virus passages (p3r+p6), whereas two other incomplete changes remained with mixed nucleotide mutations. The respective regions of BTV11kc and BTV11bsr were also sequenced, but no mutations were found, except for aa mutation E321G in S2[VP2]11 in BTV11kc. This indicated that the aa mutation E321G in S2[VP2]11 is associated with virus growth in KC cells.BTV11(S126) in KC cells. Seven out of nine mutations resulted in aa mutations. Five point mutations (four aa mutations and one silent mutation) are associated with corrections of chimeric interactions, since in BTV11(S126)bsr contained the same mutation. Four additional point mutations (three aa mutations and one silent mutation) seem to be associated with adaptation to KC cells, although the selection pressure was not very high since two aa mutations were still incomplete after six virus passages in KC cells. The most obvious and strongly selected change is aa mutation E321G in VP211. Even more, rgBTV11 also contained this E321G mutation after virus passages in KC cells.In summary, a total of nine mutations were found in several genome segments after rescue and passages of BTV11 or at 10 dpf to distinguish between infection, replication and dissemination of virus [5.1 TCID50/ml, M: 106.3 TCID50/ml, H: 108.2 TCID50/ml. Fully engorged midges were selected and processed at 0 dpf to confirm uptake, and at 10 dpf to study virus replication and dissemination  - is roughly calculated to a blood meal titer of ±2 × 105 TCID50/ml for wtBTV11. Thus, one MAID50 corresponds to ±20 TCID50 wtBTV11 in a blood meal estimated to be 100 nl for fully engorged competent C. sonorensis midges.Bodies of all groups were PCR-positive at 0 dpf, except for one in group L. The difference in virus dose in blood meals was clearly observed. Heads were negative at 0 dpf, except for one in groups L and M and two in group H. Likely, contamination by decapitation, virus on mouth parts, or incomplete swallowing of the blood meal can occur. At 10 dpf, positive results of bodies and heads indicated infection, replication and dissemination of wtBTV11. Bodies and heads in all three groups clearly segregated in PCR-positives and negatives. The percentage of PCR positives at 10 dpf was similar for bodies and heads in each group, and was approximately 40%, 79% and 89% for group L, M and H, respectively Fig. . The meadel) lacking a region in NS3/NS3a encompassing Late Domain, and is called ‘DISA’. Midges were blood-fed containing ±2 × 106 TCID50/ml DISA or rgBTV11. This corresponded to ±200 TCID50 which is ±10 MAID50, and thus sufficient to infect a high percentage of midges. In addition, virus replication was studied after intrathoracic inoculation with the same amount of DISA vaccine virus.BTV1 deficient for NS3/NS3a expression, named Disabled Infectious Single Animal (DISA) vaccine does not propagate in midges after intrathoracic inoculation . Here, wvia the haemolymph into the head.Virus uptake by feeding or inoculation was confirmed by 100% PCR positive bodies at 0 dpf and 0 dpi Fig. . Mean Cq50. Despite of 75% PCR-positive bodies at 10 dpf with DISA, all heads remained PCR negative. Likely, DISA initially infected gut cells but is not released into the haemolymph and was therefore not disseminated to the head. DISA-propagation in competent midges after oral uptake was unsuccessful by lack of Late Domain in NS3/NS3a protein. After intrathoracic inoculation, a small decrease in mean Cq value (more virus) in bodies and heads was observed at 10 dpi. This indicated infection and replication of DISA after intrathoracic inoculation similarly as observed in bodies after blood-feeding. Deletion of Late Domain of NS3/NS3a protein caused differential virus propagation in vivo. We conclude that functional NS3/NS3a is essential for BTV propagation in competent midges after oral uptake. These results confirmed the DISA principle of BTV lacking functional NS3/NS3a expression.At 10 dpf, 65% of the bodies and 44% of the heads of midges fed with rgBTV11 were PCR-positive, indicating infection, replication and dissemination of rgBTV11 bsr, BTV11(S126)kc, and rgBTV11 were fed to midges using a normalized virus titer of ±2 × 106 TCID50/ml bsr. As expected, rgBTV11 efficiently propagated in blood-fed midges, since bodies and heads at 10 dpf were 80% and 88% PCR-positive, respectively kc propagated in midges, as indicated by > 90% infected midges at 10 dpf. In contrast, BTV11(S126)bsr showed 36% and 16% PCR-positive bodies and heads at 10 dpf, respectively. This indicated that infection of midges by mammalian cell-adapted BTV11(S126)bsr was less efficient than by Culicoides-adapted BTV11(S126)kc and rgBTV11. In addition, mean Cq values were higher (less virus) for BTV11(S126)bsr than for BTV11(S126)kc and rgBTV11 kc propagated similar to, or even slightly better than, rgBTV11 and more importantly, much better than BTV11(S126)bsr. We conclude that infection, replication and dissemination of BTV11(S126)kc is more efficient in competent midges than of BTV11(S126)bsr. Furthermore, the increased propagation in vitro and in vitro of BTV11(S126)kc is likely caused by one aa mutation in outer shell protein VP2.BTV11 of reverse genetics derived BTV1 reassortant (corresponding to ±3 MAID50 of wtBTV11) did not infect competent midges, although intrathoracic inoculation was successful [50 but likely the used virus strain is even more important. The midge feeding model was used to study propagation of BTV11 mutants in more detail.Spread of arthropod-borne viruses depends on virus replication in the host and the competent insect vector leading to viremia in the host and virus excretion in insect saliva, respectively. wtBTV11 infects competent on 7 dpf . In thisf midges . A clearccessful . ClearlyC. sonorensis midges after blood-feeding  replicated well in BSR and KC cells  (here named DISA) lacking Late Domain of NS3/NS3a demonstrates similar characteristics  corresponds to the in-frame deletion of 77 aa codons in the experimental DISA vaccine for African Horse Sickness [in vitro characteristics. It is likely that it would also be blocked in virus release and dissemination in blood-fed midges. Taken together, a small deletion in NS3/NS3a protein encompassing Late Domain is sufficient to establish nonvirulence, the DIVA principle (Differentiating Infected from Vaccinated), and the DISA principle  .in vitro for several reassortants of typical BTV1 and atypical BTV26. In this study, we aimed to determine differential virus replication in vivo, and rescued typical BTV11 with S126. Rescue of BTV11(S126) was less efficient than BTV11, and adaptive changes after rescue were assumed to improve virus growth on BSR cells. In agreement with Pullinger et al. [26) initially showed limited virus production in KC cells  to KC cells. BTV11(S126)kc had obvious phenotypical differences as observed by increased virus growth and enlargement of immunostained foci in infected KC cell monolayers kc and rgBTV11kc. BTV11(S126) was independently rescued again, and in addition, an S126 mono-reassortant of BTV1 strain RSArrrr/01 was rescued (BTV1(S126) according to previous results [26 mono-reassortants showed hardly replication in KC cells but could be adapted to KC cells .26)  contained the same point mutation leading to the D309G aa mutation, while independently rescued BTV11(S126) had a point mutation G936A leading to aa mutation D309N. These results demonstrate the importance of mutating Asp on aa position 309. Asp-309 is mapped in the unmodelled region between NTD and PD of RdRp and has been suggested to interact with virus proteins of the replication complex [26 with proteins of BTV1 or BTV11 resulting in increased virus replication in both BSR and KC cells  is a non-conserved aa residue. The VP4, a capping enzyme, is part of the replication complex and catalyses the formation of a cap1 structure at the 5’ termini of core RNA transcripts. VP4 has distinct domains for its different activities [The aa mutation L399S in VP4tivities . The aa 11 of rescued BTV11(S126) contained aa mutations L171F and E221G. NS2 of atypical BTV25-27 also contained F-171, and E-221 in NS2 of atypical BTVs is highly conserved within a variable region. Independently rescued BTV11(S126) also contained L171F but E221G was not found, whereas S81 of rescued BTV1(S126) was not mutated. We speculate that these aa residues of NS2 are involved in interactions with VP1. NS2 recruits viral ssRNA from the cytoplasm, but its interactions with core proteins VP1, 3, 4, 6, and 7 are largely unknown (reviewed in [NS2iewed in ). Our re26, one aa mutation in VP311, and one silent mutation in S126 were found in BTV11(S126)kc  virus. More importantly, no obvious mutations in VP126 were found that are associated to adaptation to virus growth in KC cells. Surprisingly, we found that VP126 is not involved in differential virus replication in vitro.One aa mutation in VP126)kc and BTV11(S126)bsr is nucleotide mutation A981G in S211 resulting in E321G in VP211  contained also mutations in VP211; E401G and I503V, and the S126 chimeric virus based on BTV serotype 1 also contained one N229S mutation in VP21. In a previous study, BTV8/net07/e1 was also passaged in KC cells (BTV8/net07/e1kc3) leading to two nucleotide changes of which one resulted in R400G in VP28 [11 are located in or close to the externally exposed flexible tip domain of VP2 mapped to aa 191–407. A similar region of VP2 (aa residues 278–504) has been shown for AHSV [in vitro virus replication in both cell types, but the corresponding region (aa residues 284–510) in AHSV4-VP2 seems to be involved specifically in virus replication in KC cells, since virus release from KC cells of this AHSV deletion mutant was slightly delayed [in vitro.The most obvious difference between BTV11kc propagated better in competent midges than BTV11(S126)bsr  of wtBTV11 infects 50% of fully engorged midges and corresponds to a complete blood meal of 100 nl containing ±2 × 105 TCID50/ml or 20 TCID50 wtBTV11. Clearly, infection of midges by blood-feeding depends on virus uptake but the used virus strain is even more important. A small 72 amino acid in-frame deletion of NS3/NS3a protein completely blocks virus dissemination in blood-fed midges. Further, detailed knowledge of protein-protein interactions in the virion was generated by analysis of BTV reassortants. In addition, a point mutation in outer shell protein VP2 was identified in Culicoides-adapted BTV that is associated with differential virus replication in vitro and in vivo and thus with vector competence. In conclusion, two examples of small changes in BTV are shown which strongly affect virus infection, replication and dissemination of virus in competent midges. All these processes are part of the key mechanism crucial for vector competence and thus for spread of the bluetongue virus.The midge feeding model, including decapitation and separate testing of individual bodies and heads, is a suitable approach to identify viral factors involved in propagation of virus mutants in more detail. One MAID


Ebolavirus genus has at least five members, four of which are known to cause deadly disease in humans. An ideal therapy or a vaccine would protect against all ebolaviruses, but identifying a common weakness in all of them has remained elusive.The  Ebolavirus genus has at least five members, four of which are known to cause deadly disease in humans. An ideal therapy or a vaccine would protect against all ebolaviruses, but identifying a common weakness in all of them has remained elusive. West et al.  make the exciting discovery of an “Achilles’ heel,” a cryptic and conserved pocket, on the surface antigen glycoprotein (GP) that is nearly identical in all known ebolaviruses. Key to this discovery was their study of antibody ADI-15878, the only isolated human antibody that can block infectivity of all known ebolaviruses. Following tenacious efforts in X-ray crystallography, West et al. report the high-resolution crystal structures of the Ebola virus GP and the Bundibugyo virus GP, each bound to antibody ADI-15878. These structures reveal a highly conserved but partially obscured site on the virus GP, providing a foundation for design of vaccine antigens or antiviral therapies.The  Ebolavirus genus, their surface antigens, called glycoprotein (GP), differ in amino acid sequence by at least 50%, making it difficult to develop a vaccine or treatment that would protect against all of them.The 2013-2016 Ebola virus pandemic, which infected 28,000 people and had 41% lethality, was a stark reminder that for any virus with pandemic potential, vaccines and antiviral therapies need to be developed and ready for immediate deployment. Since this pandemic, three other Ebola virus outbreaks have occurred, and there have also been several outbreaks of the related Bundibugyo virus and Sudan virus, all with similar symptoms and lethality. While all of these viruses are genetically related and belong to the Ebolavirus genus (Identifying the common weakness in all ebolaviruses was not obvious, but researchers at The Scripps Research Institute knew that such a weakness existed; they knew this because a human monoclonal antibody called ADI-15878, isolated from a survivor of the 2013-2016 Ebola virus pandemic, had already been shown to “neutralize” or block infectivity of all known members of the The article by West al.  describeThe implications for this study are that researchers can now develop vaccine antigens and antiviral therapies that target this site. In terms of a vaccine, researchers can now pursue the design of vaccine antigens that better expose this important site of vulnerability on GP and elicit broadly neutralizing antibodies, like ADI-15878, that neutralize all ebolaviruses. Hope is bolstered by their other paper, which describes the structure of a mouse antibody that binds to the same site as ADI-15878 but was elicited by repeated vaccination . People What is not immediately obvious from the West et al. article  is how tIn summary, the article by West et al.  is a cle