Development and validation of a method to determine the unbound paclitaxel fraction in human plasma

Paclitaxel is pharmaceutically formulated in a mixture of Cremophor EL and ethanol (1:1, v/v). The unbound fraction of the anticancer drug paclitaxel in plasma is dependent on both plasma protein binding and entrapment in Cremophor EL micelles. We have developed a simple and reproducible method for the quantification of the unbound paclitaxel fraction in human plasma. Human plasma was spiked with [3H]paclitaxel and [14C]glucose (unbound reference) and incubated at 37 degrees C for 30 min. Plasma ultrafiltrate was prepared by a micropartition system (MPS-1) and collected in a sample cup containing 100 microl of plasma to prevent the loss of paclitaxel due to adsorption. The radionuclides were separated after combustion of the biological samples using a sample oxidizer and the radioactivity was determined by liquid scintillation counting. The unbound fraction of paclitaxel was calculated by dividing the ratios of 3H and 14C in plasma ultrafiltrate and in plasma. The method was thoroughly validated using human plasma spiked with pharmacologically relevant concentrations of paclitaxel (10-1000 ng/ml) and Cremophor EL (0.25-2.0%). The method was precise, with a within-day precision ranging from 3.9 to 11.0% and a between-day precision ranging from 5.8 to 13.1%. In patient plasma with low serum albumin values containing 1% of Cremophor EL, the unbound fraction appeared to be significantly higher than that in plasma with normal albumin values. The determination of the unbound fraction of paclitaxel proved to be stable during a 10-week storage at -20 degrees C. Furthermore, the assay was applicable in patient samples. This assay can be used to determine the unbound fraction of paclitaxel in plasma. Moreover, its design should allow the determination of the unbound concentrations of other hydrophobic drugs.     

The long-term effect of current and new interventions on the new case detection of leprosy: a modeling study

Background

Although the number of newly detected leprosy cases has decreased globally, a quarter of a million new cases are detected annually and eradication remains far away. Current options for leprosy prevention are contact tracing and BCG vaccination of infants. Future options may include chemoprophylaxis and early diagnosis of subclinical infections. This study compared the predicted trends in leprosy case detection of future intervention strategies.

Methods

Seven leprosy intervention scenarios were investigated with a microsimulation model (SIMCOLEP) to predict future leprosy trends. The baseline scenario consisted of passive case detection, multidrug therapy, contact tracing, and BCG vaccination of infants. The other six scenarios were modifications of the baseline, as follows: no contact tracing; with chemoprophylaxis; with early diagnosis of subclinical infections; replacement of the BCG vaccine with a new tuberculosis vaccine ineffective against Mycobacterium leprae (“no BCG”); no BCG with chemoprophylaxis; and no BCG with early diagnosis.

Findings

Without contact tracing, the model predicted an initial drop in the new case detection rate due to a delay in detecting clinical cases among contacts. Eventually, this scenario would lead to new case detection rates higher than the baseline program. Both chemoprophylaxis and early diagnosis would prevent new cases due to a reduction of the infectious period of subclinical cases by detection and cure of these cases. Also, replacing BCG would increase the new case detection rate of leprosy, but this effect could be offset with either chemoprophylaxis or early diagnosis.

Conclusions

This study showed that the leprosy incidence would be reduced substantially by good BCG vaccine coverage and the combined strategies of contact tracing, early diagnosis, and treatment of infection and/or chemoprophylaxis among household contacts. To effectively interrupt the transmission of M. leprae, it is crucial to continue developing immuno- and chemoprophylaxis strategies and an effective test for diagnosing subclinical infections.     

Poor antibody validation is a challenge in biomedical research: a case study for detection of c-FLIP

Successful translation of findings derived from preclinical studies into effective therapies is critical in biomedical research. Lack of robustness and reproducibility of the preclinical data, due to insufficient number of repeats, inadequate cell-based and mouse models contribute to the poor success rate. Antibodies are widely used in preclinical research, notably to determine the expression of potential therapeutic targets in tissues of interest, including tumors, but also to identify disease and/or treatment response biomarkers. We sought to determine whether the current antibody characterization standards in preclinical research are sufficient to ensure reliability of the data found in peer-reviewed publications. To address this issue, we used detection of the protein c-FLIP, a major factor of resistance to apoptosis, as a proof of concept. Accurate detection of endogenous c-FLIP levels in the preclinical settings is imperative since it is considered as a potential theranostic biomarker. Several sources of c-FLIP antibodies validated by their manufacturer and recommended for western blotting were therefore rigorously tested. We found a wide divergence in immune recognition properties. While these antibodies have been used in many publications, our results show that several of them failed to detect endogenous c-FLIP protein by Western blotting. Our results suggest that antibody validation standards are inadequate, and that systematic use of genetic knockdowns and/or knockouts to establish proof of specificity is critical, even for antibodies previously used in the scientific literature. Because antibodies are fundamental tools in both preclinical and clinical research, ensuring their specificity is crucial.     

Active case detection methods for crusted scabies and leprosy: A systematic review

BackgroundCrusted scabies is endemic in some remote Aboriginal communities in the Northern Territory (NT) of Australia and carries a high mortality risk. Improvement in active case detection (ACD) for crusted scabies is hampered by a lack of evidence about best practice. We therefore conducted a systematic review of ACD methods for leprosy, a condition with similar ACD requirements, to consider how findings could be informative to crusted scabies detection.Methods and principle findingsWe conducted systematic searches in MEDLINE, CINAHL, Scopus and the Cochrane Database for Systematic Reviews for studies published since 1999 that reported at least one comparison rate (detection or prevalence rate) against which the yield of the ACD method could be assessed. The search yielded 15 eligible studies from 511. Study heterogeneity precluded meta-analysis. Contact tracing and community screening of marginalised ethnic groups yielded the highest new case detection rates. Rapid community screening campaigns, and those using less experienced screening personnel, were associated with lower suspect confirmation rates. There is insufficient data to assess whether ACD campaigns improve treatment outcomes or disease control.ConclusionThis review demonstrates the importance of ACD campaigns in communities facing the highest barriers to healthcare access and within neighbourhoods of index cases. The potential benefit of ACD for crusted scabies is not quantified, however, lessons from leprosy suggest value in follow-up with previously identified cases and their close contacts to support for scabies control and to reduce the likelihood of reinfection in the crusted scabies case. Skilled screening personnel and appropriate community engagement strategies are needed to maximise screening uptake. More research is needed to assess ACD cost effectiveness, impact on disease control, and to explore ACD methods capable of capturing the homeless and highly mobile who may be missed in household centric models.     

Individual and community factors determining delayed leprosy case detection: A systematic review

BackgroundThe number of new leprosy cases is declining globally, but the disability caused by leprosy remains an important disease burden. The chance of disability is increased by delayed case detection. This review focusses on the individual and community determinants of delayed leprosy case detection.MethodsThis study was conducted according to the PRISMA guidelines (Preferred Reporting Items for Systematic Reviews and Meta-Analysis). The study protocol is registered in PROSPERO (code: CRD42020189274). To identify determinants of delayed detection, data was collected from five electronic databases: Embase.com, Medline All Ovid, Web of Science, Cochrane CENTRAL, and the WHO Global Health Library.ResultsWe included 27 papers from 4315 records assessed. They originated in twelve countries, had been published between January 1, 2000, and January 31, 2021, and described the factors related to delayed leprosy case detection, the duration of the delayed case, and the percentage of Grade 2 Disability (G2D). The median delay in detection ranged from 12 to 36 months, the mean delay ranged from 11.5 to 64.1 months, and the percentage of G2D ranged from 5.6 to 43.2%. Health-service-seeking behavior was the most common factor associated with delayed detection. The most common individual factors were older age, being male, having a lower disease-symptom perception, having multibacillary leprosy, and lack of knowledge. The most common socioeconomic factors were living in a rural area, performing agricultural labor, and being unemployed. Stigma was the most common social and community factor.ConclusionsDelayed leprosy case detection is clearly correlated with increased disability and should therefore be a priority of leprosy programs. Interventions should focus on determinants of delayed case detection such as health-service-seeking behavior, and should consider relevant individual, socioeconomic, and community factors, including stigmatization. Further study is required of the health service-related factors contributing to delay.     

Development and validation of competitive ELISA to determine follicular fluid progesterone concentrations

A simple, direct and reliable heterologous ELISA was developed and validated to determine follicular fluid progesterone concentrations in 10- to 20-mm antral follicles in heifers. A competitive ELISA was developed, using a policlonal antisera raised in New Zealand white rabbits and horseradish peroxidase as label. Standard curve covered a range between 0 and 1 ng per well. The intra- and inter-assay coefficients of variation (CV) were 6.54 and 8.27%, respectively (n = 10). The low detection limit was 1 pg per well with a sensitivity of 50% binding 83.17 pg per well. Comparison of ELISA with RIA showed a correlation coefficient of 0.98. A total of 34 follicles obtained from heifers in the follicular phase of the estrous cycle were tested, and the mean progesterone concentration was 86.72 +/- 20.39 ng/ml. These results were similar to those previously reported for the same species, age, follicular size and stage of cycle.     

Quantifying cultural macro-evolution: a case study of the hinoeuma fertility drop

Understanding patterns and underlying processes of human cultural diversity has been a major challenge in evolutionary anthropology. Recent developments in the study of cultural macro-evolution have illuminated various novel aspects of cultural phenomena at the population level. However, limitations in data availability have constrained previous analyses to use simplest models ignoring factors that potentially affect cultural evolutionary dynamics. Here, we focus on two such factors: accumulated effects of cultural transmission between populations over time and variation in social influence among populations. As a test case, we analyze data on the hinoeuma fertility drop, the Japanese nation-wide drastic decline in the number of births caused by a culturally-transmitted superstition recurring every sixty years, to show that these factors do play significant roles. Specifically, our results suggest that transmission of the superstition in a short timescale has tended to occur among neighboring populations, while transmission in a long timescale is likely to have occurred between populations culturally close to each other, with the cultural closeness being measured by similarity in dialects. The results also indicate a special role played by a population occupying a center in a language–distance network (the cultural center) in the spread of the superstition.     

The quick delay questionnaire: a measure of delay aversion and discounting in adults

Individuals with ADHD often display an altered response to delay. To date, assessment of this has typically involved neuropsychological testing-however, such tests are designed specifically for children and may not be suitable for adults. They are also relatively time-consuming and expensive. In the current paper, we describe the initial validation of a short questionnaire to assess delay-related behaviour in adults. The Quick Delay Questionnaire (QDQ) is a 10-item scale. The questionnaire was administered to 575 participants from the normal population (ranging in age from 18 to 77 years). Forty of the original sample were selected at random and tested 1 week later. Data on ADHD, anxiety and depression were also collected. There were two five-item scales-(1) delay aversion; and (2) delay discounting. These had internal consistency and had good reliability. Subscales were differentially associated with ADHD, anxiety and depression. The QDQ is a potentially valuable way of assessing response to delay in adults. Further work is required to validate the scale against direct observation and neuropsychological assessment in clinically ascertained samples.     

Development and field validation of a regional,management‐scale habitat model: A koala Phascolarctos cinereus case study

Species distribution models have great potential to efficiently guide management for threatened species, especially for those that are rare or cryptic. We used MaxEnt to develop a regional‐scale model for the koala Phascolarctos cinereus at a resolution (250 m) that could be used to guide management. To ensure the model was fit for purpose, we placed emphasis on validating the model using independently‐collected field data. We reduced substantial spatial clustering of records in coastal urban areas using a 2‐km spatial filter and by modeling separately two subregions separated by the 500‐m elevational contour. A bias file was prepared that accounted for variable survey effort. Frequency of wildfire, soil type, floristics and elevation had the highest relative contribution to the model, while a number of other variables made minor contributions. The model was effective in discriminating different habitat suitability classes when compared with koala records not used in modeling. We validated the MaxEnt model at 65 ground‐truth sites using independent data on koala occupancy (acoustic sampling) and habitat quality (browse tree availability). Koala bellows (n = 276) were analyzed in an occupancy modeling framework, while site habitat quality was indexed based on browse trees. Field validation demonstrated a linear increase in koala occupancy with higher modeled habitat suitability at ground‐truth sites. Similarly, a site habitat quality index at ground‐truth sites was correlated positively with modeled habitat suitability. The MaxEnt model provided a better fit to estimated koala occupancy than the site‐based habitat quality index, probably because many variables were considered simultaneously by the model rather than just browse species. The positive relationship of the model with both site occupancy and habitat quality indicates that the model is fit for application at relevant management scales. Field‐validated models of similar resolution would assist in guiding management of conservation‐dependent species.     

Development and validation of a multiplex real-time qPCR assay using GMP-grade reagents for leprosy diagnosis

Leprosy is a chronic dermato-neurological disease caused by Mycobacterium leprae, an obligate intracellular bacterium. Timely detection is a challenge in leprosy diagnosis, relying on clinical examination and trained health professionals. Furthermore, adequate care and transmission control depend on early and reliable pathogen detection. Here, we describe a qPCR test for routine diagnosis of leprosy-suspected patients. The reaction simultaneously amplifies two specific Mycobacterium leprae targets (16S rRNA and RLEP), and the human 18S rRNA gene as internal control. The limit of detection was estimated to be 2.29 copies of the M. leprae genome. Analytical specificity was evaluated using a panel of 20 other skin pathogenic microorganisms and Mycobacteria, showing no cross-reactivity. Intra- and inter-operator C_p variation was evaluated using dilution curves of M. leprae DNA or a synthetic gene, and no significant difference was observed between three operators in two different laboratories. The multiplex assay was evaluated using 97 patient samples with clinical and histopathological leprosy confirmation, displaying high diagnostic sensitivity (91%) and specificity (100%). Validation tests in an independent panel of 50 samples confirmed sensitivity and specificity of 97% and 98%, respectively. Importantly, assay performance remained stable for at least five months. Our results show that the newly developed multiplex qPCR effectively and specifically detects M. leprae DNA in skin samples, contributing to an efficient diagnosis that expedites the appropriate treatment.     

Development of a validation protocol to determine the ability of screw-top containers to be watertight and to withstand impact damage from accidental dropping

Polypropylene screw-top containers are used to collect, transport and store a variety of tissues within tissue banks. These containers are validated for use by tissue banks but no standard validation protocol is used. We present here a protocol for testing screw-top containers for leakage, evaporation and the ability to withstand accidental impact damage. Three different containers were tested, MedFor S072, MedFor S277 and MacoPharma PROT0483. The validation can detect differences between different manufacturer’s containers and this protocol will be used in future validations of screw-top containers within National Blood Service Tissue Services.     

End-of-life practices in Danish ICUs: development and validation of a questionnaire

ABSTRACT: BACKGROUND: Practices for withholding or withdrawing therapy vary according to professional, cultural and religious differences. No Danish-validated questionnaire examining withholding and withdrawing practices exists, thus the aim of this study was to develop and validate a questionnaire for surveying the views of intensive care nurses, intensivists, and primary physicians regarding collaboration and other aspects of withholding and withdrawing therapy in the ICU. METHODS: A questionnaire was developed on the basis of literature, focus group interviews with intensive care nurses and intensivists, and individual interviews with primary physicians. The questionnaire was validated in the following 3 phases: a qualitative test with 17 participants; a quantitative pilot test with 60 participants; and a survey with 776 participants. The validation process included tests for face and content validity (by interviewing participants in the qualitative part of the pilot study), reliability (by assessing the distribution of responses within the individual response categories), agreement (by conducting a test-retest, evaluated by paired analyses), known groups' validity (as a surrogate test for responsiveness, by comparing two ICUs with a known difference in end-of-life practices), floor and ceiling effect, and missing data. RESULTS: Face and content validity were assessed as good by the participants in the qualitative pilot test; all considered the questions relevant and none of the participants found areas lacking. Almost all response categories were used by the participants, thus demonstrating the questionnaires ability to distinguish between different respondents, agreement was fair (the average test-retest agreement for the Likert scale responses was 0.54 (weighted kappa; range, 0.25-0.73), and known groups' validity was proved by finding significant differences in level of satisfaction with interdisciplinary collaboration and in experiences of withdrawal decisions being unnecessary postponed. Floor and ceiling effect was in accordance with other questionnaires, and missing data was limited to a range of 0-7% for all questions. CONCLUSIONS: The validation showed good and fair areas of validity of the questionnaire. The questionnaire is considered a useful tool to assess the perceptions of collaboration and other aspects of withholding and withdrawing therapy practices in Danish ICUs amongst nurses, intensivists, and primary physicians.     

On the relationship between interindividual cultural transmission and population-level cultural diversity: a case study of weaving in Iranian tribal populations

It is often assumed that parent-to-child cultural transmission leads to similarities and differences among groups evolving through descent with modification (“phylogenesis”). Similarly, cultural transmission between peers, and between adults and children who are not their offspring, is widely believed to result in groups exchanging cultural traits (“ethnogenesis”). However, neither of these assumptions has been examined empirically. Here, we test them using ethnographic data on craft learning in Iranian tribal populations and the cladistic method of phylogenetic analysis. We find that parent-to-child transmission dominates learning during childhood, but the other two forms of interindividual transmission become more important in later periods. The latter do not, however, appear to have resulted in extensive exchange of cultural traits among tribes. Instead we find that most of the variation among the tribes' craft assemblages can be explained by descent with modification. This can be accounted for by the fact that weavers usually only share their knowledge with members of their own tribe and are prevented from interacting with women from other groups by social norms. These findings demonstrate that the relationship between processes of cultural evolution at the level of the individual and processes of cultural evolution at the level of the group is more complex than is usually acknowledged, and highlight the need for more integrated studies of the processes operating at both scales.     

How to reduce the impact of artificial lighting on moths: A case study on cultural heritage sites in Slovenia

In an ever more artificially illuminated world, common moth behaviour, flight-to-light, causes declines in their abundance and diversity that can have severe impacts on ecosystems. To test if it is possible to reduce the number of moths attracted to artificially illuminated objects, the original lighting of 15 cultural heritage buildings in Slovenia was substituted with blue or yellow lighting. These three illumination types differed in the amount of luminance, percentage of UV and short-wavelength light which are known to affect flight-to-light of moths. During our three-year field study approximately 20% of all known moth species in Slovenia were recorded. The blue and yellow illumination type attracted up to six times less specimens and up to four times less species compared to the original illumination type. This was true for all detected moths as well as within separate moth groups. This gives our study a high conservation value: usage of alternative, environmentally more acceptable illumination can greatly reduce the number of moths attracted to artificially illuminated objects.     

The alpine rural landscape as a cultural reserve: the case study of Teglio in Valtellina

Biodiversity and Conservation - This research examines a portion of the Italian alpine landscape in order to find a comprehension mode and a strategic proposal to safeguard this rural heritage. The...     

Preventing nerve function impairment in leprosy: validation and updating of a prediction rule

Background

To validate and update a prediction rule for estimating the risk of leprosy-related nerve function impairment (NFI).

Methodology/Principal Findings

Prospective cohort using routinely collected data, in which we determined the discriminative ability of a previously published rule and an updated rule with a concordance statistic (c). Additional risk factors were analyzed with a Cox proportional hazards regression model. The population consisted of 1,037 leprosy patients newly diagnosed between 2002 and 2003 in the health care facilities of the Rural Health Program in Nilphamari and Rangpur districts in northwest Bangladesh. The primary outcome was the time until the start of treatment. An NFI event was defined as the decision to treat NFI with corticosteroids after diagnosis. NFI occurred in 115 patients (13%; 95% confidence interval 11%–16%). The original prediction rule had adequate discriminative ability (c = 0.79), but could be improved by substituting one predicting variable: ‘long-standing nerve function impairment at diagnosis’ by ‘anti-PGL-I antibodies’. The adjusted prediction rule was slightly better (c = 0.81) and identified more patients with NFI (80%) than the original prediction rule (72%).

Conclusions/Significance

NFI can well be predicted by using the risk variables ‘leprosy classification’ and ‘anti-PGL-I antibodies’. The use of these two variables that do not include NFI offer the possibility of predicting NFI, even before it occurs for the first time. Surveillance beyond the treatment period can be targeted to those most likely to benefit from preventing permanent disabilities.     

Number of people requiring post-exposure prophylaxis to end leprosy: A modeling study

BackgroundWorldwide, around 210,000 new cases of leprosy are detected annually. To end leprosy, i.e. zero new leprosy cases, preventive interventions such as contact tracing and post-exposure prophylaxis (PEP) are required. This study aims to estimate the number of people requiring PEP to reduce leprosy new case detection (NCD) at national and global level by 50% and 90%.Methodology/Principal findingsThe individual-based model SIMCOLEP was fitted to seven leprosy settings defined by NCD and MB proportion. Using data of all 110 countries with known leprosy patients in 2016, we assigned each country to one of these settings. We predicted the impact of administering PEP to about 25 contacts of leprosy patients on the annual NCD for 25 years and estimated the number of contacts requiring PEP per country for each year. The NCD trends show an increase in NCD in the first year (i.e. backlog cases) followed by a significant decrease thereafter. A reduction of 50% and 90% of new cases would be achieved in most countries in 5 and 22 years if 20.6 and 40.2 million people are treated with PEP over that period, respectively. For India, Brazil, and Indonesia together, a total of 32.9 million people requiring PEP to achieve a 90% reduction in 22 years.Conclusion/SignificanceThe leprosy problem is far greater than the 210,000 new cases reported annually. Our model estimates of the number of people requiring PEP to achieve significant reduction of new leprosy cases can be used by policymakers and program managers to develop long-term strategies to end leprosy.     

广西南丹县生物多样性和文化多样性与生态旅游的关系

南丹县地处广西西北边陲 ,与贵州接壤 ,为多民族聚居地 ,是大西南出海铁路和高速公路通道的咽喉要地。全县总面积 3916ｋｍ2 ,人口 2 7.6万 ,全国第二大水电站———龙滩水电站距县城 76ｋｍ ,是一个具有交通枢纽和商贸物资集散功能的地方。矿产资源丰富 ,目前已查明的有锡、锑、铅、铜、银和铟等 2 0多种有色金属 ,储量超过 10 0 0× 10 4 ｔ,是罕见的富矿区 ,有“有色金属之乡”之称。其中 ,锡的储量 144× 10 4 ｔ,随着云南固旧锡都的衰落 ,南丹县将成为中国新的锡都。该县农业资源也很丰富 ,有许多著名的土特产品 ,例如黄蜡李、瑶鸡、魔…