Unlocking the diagnostic power of plasma extracellular vesicle miR-200 family in pancreatic ductal adenocarcinoma.

BACKGROUND: Distinguishing benign from malignant pancreaticobiliary disease is challenging because of the absence of reliable biomarkers. Circulating extracellular vesicles (EVs) have emerged as functional mediators between cells. Their cargos, including microRNAs (miRNAs), are increasingly acknowledged as an important source of potential biomarkers. This multicentric, prospective study aimed to establish a diagnostic plasma EV-derived miRNA signature to discriminate pancreatic ductal adenocarcinoma (PDAC) from benign pancreaticobiliary disease. METHODS: Plasma EVs were isolated using size exclusion chromatography (SEC) and characterised using nanoparticle tracking analysis, electron microscopy and Western blotting. EV-RNAs underwent small RNA sequencing to discover differentially expressed markers for PDAC (n = 10 benign vs. 10 PDAC). Candidate EV-miRNAs were then validated in a cohort of 61 patients (n = 31 benign vs. 30 PDAC) by RT-qPCR. Logistic regression and optimal thresholds (Youden Index) were used to develop an EV-miR-200 family model to detect cancer. This model was tested in an independent cohort of 95 patients (n = 30 benign, 33 PDAC, and 32 cholangiocarcinoma). RESULTS: Small RNA sequencing and RT-qPCR showed that EV-miR-200 family members were significantly overexpressed in PDAC vs. benign disease. Combined expression of the EV-miR-200 family showed an AUC of 0.823. In an independent validation cohort, application of this model showed a sensitivity, specificity and AUC of 100%, 88%, and 0.97, respectively, for diagnosing PDAC. CONCLUSIONS: This is the first study to validate plasma EV-miR-200 members as a clinically-useful diagnostic biomarker for PDAC. Further validation in larger cohorts and clinical trials is essential. These findings also suggest the potential utility in monitoring response and/or recurrence.

MicroRNAs as Bile-based biomarkers in pancreaticobiliary cancers (MIRABILE): a cohort study.

BACKGROUND: Biliary obstruction can be due to both malignant and benign pancreaticobiliary disease. Currently, there are no biomarkers that can accurately help make this distinction. MicroRNAs (miRNAs) are stable molecules in tissue and biofluids that are commonly deregulated in cancer. The MIRABILE study aimed to identify miRNAs in bile that can differentiate malignant from benign pancreaticobiliary disease. MATERIALS AND METHODS: There were 111 patients recruited prospectively at endoscopic retrograde cholangiopancreatography (ERCP) or percutaneous transhepatic cholangiography (PTC) for obstructive jaundice, and bile was aspirated for cell-free RNA (cfRNA) extraction and analysis. In a discovery cohort of 78 patients (27 with pancreatic ductal adenocarcinoma (PDAC), 14 cholangiocarcinoma (CCA), 37 benign disease), cfRNA was subjected to small-RNA sequencing. LASSO regression was used to define bile miRNA signatures, and NormFinder to identify endogenous controls. In a second cohort of 87 patients (34 PDAC, 14 CCA, 39 benign disease), RT-qPCR was used for validation. RESULTS: LASSO regression identified 14 differentially-expressed bile miRNAs of which 6 were selected for validation. When comparing malignant and benign pancreaticobiliary disease, bile miR-340 and miR-182 were validated and significantly differentially expressed (P<0.05 and P<0.001, respectively). This generated an AUC of 0.79 (95%CI 0.70-0.88, sensitivity 65%; specificity 82%) in predicting malignant disease. CONCLUSION: Bile collected during biliary drainage contains miRNAs able to differentiate benign from malignant pancreaticobiliary diseases in patients with obstructive jaundice. These bile miRNAs have the potential to increase diagnostic accuracy.

A bile-based microRNA signature for differentiating malignant from benign pancreaticobiliary disease.

Differentiating between pancreatic ductal adenocarcinoma (PDAC) and cholangiocarcinoma (CCA) is crucial for the appropriate course of treatment, especially with advancements in the role of neoadjuvant chemotherapies for PDAC, compared to CCA. Furthermore, benign pancreaticobiliary diseases can mimic malignant disease, and indeterminate lesions may require repeated investigations to achieve a diagnosis. As bile flows in close proximity to these lesions, we aimed to establish a bile-based microRNA (miRNA) signature to discriminate between malignant and benign pancreaticobiliary diseases. We performed miRNA discovery by global profiling of 800 miRNAs using the NanoString nCounter platform in prospectively collected bile samples from malignant (n = 43) and benign (n = 14) pancreaticobiliary disease. Differentially expressed miRNAs were validated by RT-qPCR and further assessed in an independent validation cohort of bile from malignant (n = 37) and benign (n = 38) pancreaticobiliary disease. MiR-148a-3p was identified as a discriminatory marker that effectively distinguished malignant from benign pancreaticobiliary disease in the discovery cohort (AUC = 0.797 [95% CI 0.68-0.92]), the validation cohort (AUC = 0.772 [95% CI 0.66-0.88]), and in the combined cohorts (AUC = 0.752 [95% CI 0.67-0.84]). We also established a two-miRNA signature (miR-125b-5p and miR-194-5p) that distinguished PDAC from CCA (validation: AUC = 0.815 [95% CI 0.67-0.96]; and combined cohorts: AUC = 0.814 [95% CI 0.70-0.93]). Our research stands as the largest, multicentric, global profiling study of miRNAs in the bile from patients with pancreaticobiliary disease. We demonstrated their potential as clinically useful diagnostic tools for the detection and differentiation of malignant pancreaticobiliary disease. These bile miRNA biomarkers could be developed to complement current approaches for diagnosing pancreaticobiliary cancers.

Quantifying the relationship between climatic indicators and leptospirosis incidence in Fiji: A modelling study.

Leptospirosis, a global zoonotic disease, is prevalent in tropical and subtropical regions, including Fiji where it's endemic with year-round cases and sporadic outbreaks coinciding with heavy rainfall. However, the relationship between climate and leptospirosis has not yet been well characterised in the South Pacific. In this study, we quantify the effects of different climatic indicators on leptospirosis incidence in Fiji, using a time series of weekly case data between 2006 and 2017. We used a Bayesian hierarchical mixed-model framework to explore the impact of different precipitation, temperature, and El Niño Southern Oscillation (ENSO) indicators on leptospirosis cases over a 12-year period. We found that total precipitation from the previous six weeks (lagged by one week) was the best precipitation indicator, with increased total precipitation leading to increased leptospirosis incidence (0.24 [95% CrI 0.15-0.33]). Negative values of the Niño 3.4 index (indicative of La Niña conditions) lagged by four weeks were associated with increased leptospirosis risk (-0.2 [95% CrI -0.29 --0.11]). Finally, minimum temperature (lagged by one week) when included with the other variables was positively associated with leptospirosis risk (0.15 [95% CrI 0.01-0.30]). We found that the final model was better able to capture the outbreak peaks compared with the baseline model (which included seasonal and inter-annual random effects), particularly in the Western and Northern division, with climate indicators improving predictions 58.1% of the time. This study identified key climatic factors influencing leptospirosis risk in Fiji. Combining these results with demographic and spatial factors can support a precision public health framework allowing for more effective public health preparedness and response which targets interventions to the right population, place, and time. This study further highlights the need for enhanced surveillance data and is a necessary first step towards the development of a climate-based early warning system.

Towards a leptospirosis early warning system in northeastern Argentina.

Leptospirosis is a zoonotic disease with a high burden in Latin America, including northeastern Argentina, where flooding events linked to El Niño are associated with leptospirosis outbreaks. The aim of this study was to evaluate the value of using hydrometeorological indicators to predict leptospirosis outbreaks in this region. We quantified the effects of El Niño, precipitation, and river height on leptospirosis risk in Santa Fe and Entre Ríos provinces between 2009 and 2020, using a Bayesian modelling framework. Based on several goodness of fit statistics, we selected candidate models using a long-lead El Niño 3.4 index and shorter lead local climate variables. We then tested predictive performance to detect leptospirosis outbreaks using a two-stage early warning approach. Three-month lagged Niño 3.4 index and one-month lagged precipitation and river height were positively associated with an increase in leptospirosis cases in both provinces. El Niño models correctly detected 89% of outbreaks, while short-lead local models gave similar detection rates with a lower number of false positives. Our results show that climatic events are strong drivers of leptospirosis incidence in northeastern Argentina. Therefore, a leptospirosis outbreak prediction tool driven by hydrometeorological indicators could form part of an early warning and response system in the region.

Estimating the duration of antibody positivity and likely time of Leptospira infection using data from a cross-sectional serological study in Fiji.

BACKGROUND: Leptospirosis is a zoonotic disease prevalent throughout the world, but with particularly high burden in Oceania (including the Pacific Island Countries and Territories). Leptospirosis is endemic in Fiji, with outbreaks often occurring following heavy rainfall and flooding. As a result of non-specific clinical manifestation and diagnostic challenges, cases are often misdiagnosed or under-ascertained. Furthermore, little is known about the duration of persistence of antibodies to leptospirosis, which has important clinical and epidemiological implications. METHODOLOGY AND PRINCIPAL FINDINGS: Using the results from a serosurvey conducted in Fiji in 2013, we fitted serocatalytic models to estimate the duration of antibody positivity and the force of infection (FOI, the rate at which susceptible individuals acquire infection or seroconversion), whilst accounting for seroreversion. Additionally, we estimated the most likely timing of infection. Using the reverse catalytic model, we estimated the duration of antibody persistence to be 8.33 years (4.76-12.50; assuming constant FOI) and 7.25 years (3.36-11.36; assuming time-varying FOI), which is longer than previous estimates. Using population age-structured seroprevalence data alone, we were not able to distinguish between these two models. However, by bringing in additional longitudinal data on antibody kinetics we were able to estimate the most likely time of infection, lending support to the time-varying FOI model. We found that most individuals who were antibody-positive in the 2013 serosurvey were likely to have been infected within the previous two years, and this finding is consistent with surveillance data showing high numbers of cases reported in 2012 and 2013. CONCLUSIONS: This is the first study to use serocatalytic models to estimate the FOI and seroreversion rate for Leptospira infection. As well as providing an estimate for the duration of antibody positivity, we also present a novel method to estimate the most likely time of infection from seroprevalence data. These approaches can allow for richer, longitudinal information to be inferred from cross-sectional studies, and could be applied to other endemic diseases where antibody waning occurs.

Transmission modelling of environmentally persistent zoonotic diseases: a systematic review.

Transmission of many infectious diseases depends on interactions between humans, animals, and the environment. Incorporating these complex processes in transmission dynamic models can help inform policy and disease control interventions. We identified 20 diseases involving environmentally persistent pathogens (ie, pathogens that survive for more than 48 h in the environment and can cause subsequent human infections), of which indirect transmission can occur from animals to humans via the environment. Using a systematic approach, we critically appraised dynamic transmission models for environmentally persistent zoonotic diseases to quantify traits of models across diseases. 210 transmission modelling studies were identified and most studies considered diseases of domestic animals or high-income settings, or both. We found that less than half of studies validated their models to real-world data, and environmental data on pathogen persistence was rarely incorporated. Model structures varied, with few studies considering the animal-human-environment interface of transmission in the context of a One Health framework. This Review highlights the need for more data-driven modelling of these diseases and a holistic One Health approach to model these pathogens to inform disease prevention and control strategies.

A comparison of the value of two machine learning predictive models to support bovine tuberculosis disease control in England.

Nearly a decade into Defra's current eradication strategy, bovine tuberculosis (bTB) remains a serious animal health problem in England, with c.30,000 cattle slaughtered annually in the fight against this insidious disease. There is an urgent need to improve our understanding of bTB risk in order to enhance the current disease control policy. Machine learning approaches applied to big datasets offer a potential way to do this. Regularized regression and random forest machine learning methodologies were implemented using 2016 herd-level data to generate the best possible predictive models for a bTB incident in England and its three surveillance risk areas (High-risk area [HRA], Edge area [EA] and Low-risk area [LRA]). Their predictive performance was compared and the best models in each area were used to characterize herds according to risk. While all models provided excellent discrimination, random forest models achieved the highest balanced accuracy (i.e. average of sensitivity and specificity) in England, HRA and LRA, whereas the regularized regression LASSO model did so in the EA. The time since the last confirmed incident was resolved was the only variable in the top-ten ranking in all areas according to both types of models, which highlights the importance of bTB history as a predictor of a new incident. Risk categorisation based on Receiver Operating Characteristic (ROC) analysis was carried out using the best predictive models in each area setting a 99 % threshold value for sensitivity and specificity (97 % in the LRA). Thirteen percent of herds in the whole of England as well as in its HRA, 14 % in its EA and 31 % in its LRA were classified as high-risk. These could be selected for the deployment of additional disease control measures at national or area level. In this way, low-risk herds within the area considered would not be penalised unnecessarily by blanket control measures and limited resources be used more efficiently. The methodology presented in this paper demonstrates a way to accurately identify high-risk farms to inform a targeted disease control and prevention strategy in England that supplements existing population strategies.

Neuroendocrine Neoplasms: Identification of Novel Metabolic Circuits of Potential Diagnostic Utility.

The incidence of neuroendocrine neoplasms (NEN) is increasing, but established biomarkers have poor diagnostic and prognostic accuracy. Here, we aim to define the systemic metabolic consequences of NEN and to establish the diagnostic utility of proton nuclear magnetic resonance spectroscopy (1H-NMR) for NEN in a prospective cohort of patients through a single-centre, prospective controlled observational study. Urine samples of 34 treatment-naïve NEN patients (median age: 59.3 years, range: 36-85): 18 had pancreatic (Pan) NEN, of which seven were functioning; 16 had small bowel (SB) NEN; 20 age- and sex-matched healthy control individuals were analysed using a 600 MHz Bruker 1H-NMR spectrometer. Orthogonal partial-least-squares-discriminant analysis models were able to discriminate both PanNEN and SBNEN patients from healthy control (Healthy vs. PanNEN: AUC = 0.90, Healthy vs. SBNEN: AUC = 0.90). Secondary metabolites of tryptophan, such as trigonelline and a niacin-related metabolite were also identified to be universally decreased in NEN patients, while upstream metabolites, such as kynurenine, were elevated in SBNEN. Hippurate, a gut-derived metabolite, was reduced in all patients, whereas other gut microbial co-metabolites, trimethylamine-N-oxide, 4-hydroxyphenylacetate and phenylacetylglutamine, were elevated in those with SBNEN. These findings suggest the existence of a new systems-based neuroendocrine circuit, regulated in part by cancer metabolism, neuroendocrine signalling molecules and gut microbial co-metabolism. Metabonomic profiling of NEN has diagnostic potential and could be used for discovering biomarkers for these tumours. These preliminary data require confirmation in a larger cohort.

Estimating the duration of seropositivity of human seasonal coronaviruses using seroprevalence studies.

Background: The duration of immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is still uncertain, but it is of key clinical and epidemiological importance. Seasonal human coronaviruses (HCoV) have been circulating for longer and, therefore, may offer insights into the long-term dynamics of reinfection for such viruses. Methods: Combining historical seroprevalence data from five studies covering the four circulating HCoVs with an age-structured reverse catalytic model, we estimated the likely duration of seropositivity following seroconversion. Results: We estimated that antibody persistence lasted between 0.9 (95% Credible interval: 0.6 - 1.6) and 3.8 (95% CrI: 2.0 - 7.4) years. Furthermore, we found the force of infection in older children and adults (those over 8.5 [95% CrI: 7.5 - 9.9] years) to be higher compared with young children in the majority of studies. Conclusions: These estimates of endemic HCoV dynamics could provide an indication of the future long-term infection and reinfection patterns of SARS-CoV-2.

The Role of Circular RNAs in Pancreatic Ductal Adenocarcinoma and Biliary-Tract Cancers.

Pancreatic Ductal Adenocarcinoma (PDAC) and biliary-tract cancers (BTC) often present at a late stage, and consequently patients have poor survival-outcomes. Circular RNAs (circRNAs) are non-coding RNA molecules whose role in tumourigenesis has recently been realised. They are stable, conserved and abundant, with tissue-specific expression profiles. Therefore, significant interest has arisen in their use as potential biomarkers for PDAC and BTC. High-throughput methods and more advanced bioinformatic techniques have enabled better profiling and progressed our understanding of how circRNAs may function in the competing endogenous RNA (ceRNA) network to influence the transcriptome in these cancers. Therefore, the aim of this systematic review was to describe the roles of circRNAs in PDAC and BTC, their potential as biomarkers, and their function in the wider ceRNA network in regulating microRNAs and the transcriptome. Medline, Embase, Scopus and PubMed were systematically reviewed to identify all the studies addressing circRNAs in PDAC and BTC. A total of 32 articles were included: 22 considering PDAC, 7 for Cholangiocarcinoma (CCA) and 3 for Gallbladder Cancer (GBC). There were no studies investigating Ampullary Cancer. Dysregulated circRNA expression was associated with features of malignancy in vitro, in vivo, and ex vivo. Overall, there have been very few PDAC and BTC tissues profiled for circRNA signatures. Therefore, whilst the current studies have demonstrated some of their functions in these cancers, further work is required to elucidate their potential role as cancer biomarkers in tissue, biofluids and biopsies.

Size-Exclusion Chromatography as a Technique for the Investigation of Novel Extracellular Vesicles in Cancer.

Cancer cells release extracellular vesicles, which are a rich target for biomarker discovery and provide a promising mechanism for liquid biopsy. Size-exclusion chromatography (SEC) is an increasingly popular technique, which has been rediscovered for the purposes of extracellular vesicle (EV) isolation and purification from diverse biofluids. A systematic review was undertaken to identify all papers that described size exclusion as their primary EV isolation method in cancer research. In all, 37 papers were identified and discussed, which showcases the breadth of applications in which EVs can be utilised, from proteomics, to RNA, and through to functionality. A range of different methods are highlighted, with Sepharose-based techniques predominating. EVs isolated using SEC are able to identify cancer cells, highlight active pathways in tumourigenesis, clinically distinguish cohorts, and remain functionally active for further experiments.

COVID-19 length of hospital stay: a systematic review and data synthesis.

BACKGROUND: The COVID-19 pandemic has placed an unprecedented strain on health systems, with rapidly increasing demand for healthcare in hospitals and intensive care units (ICUs) worldwide. As the pandemic escalates, determining the resulting needs for healthcare resources (beds, staff, equipment) has become a key priority for many countries. Projecting future demand requires estimates of how long patients with COVID-19 need different levels of hospital care. METHODS: We performed a systematic review of early evidence on length of stay (LoS) of patients with COVID-19 in hospital and in ICU. We subsequently developed a method to generate LoS distributions which combines summary statistics reported in multiple studies, accounting for differences in sample sizes. Applying this approach, we provide distributions for total hospital and ICU LoS from studies in China and elsewhere, for use by the community. RESULTS: We identified 52 studies, the majority from China (46/52). Median hospital LoS ranged from 4 to 53 days within China, and 4 to 21 days outside of China, across 45 studies. ICU LoS was reported by eight studies-four each within and outside China-with median values ranging from 6 to 12 and 4 to 19 days, respectively. Our summary distributions have a median hospital LoS of 14 (IQR 10-19) days for China, compared with 5 (IQR 3-9) days outside of China. For ICU, the summary distributions are more similar (median (IQR) of 8 (5-13) days for China and 7 (4-11) days outside of China). There was a visible difference by discharge status, with patients who were discharged alive having longer LoS than those who died during their admission, but no trend associated with study date. CONCLUSION: Patients with COVID-19 in China appeared to remain in hospital for longer than elsewhere. This may be explained by differences in criteria for admission and discharge between countries, and different timing within the pandemic. In the absence of local data, the combined summary LoS distributions provided here can be used to model bed demands for contingency planning and then updated, with the novel method presented here, as more studies with aggregated statistics emerge outside China.

Tooling-up for infectious disease transmission modelling.

In this introduction to the Special Issue on methods for modelling of infectious disease epidemiology we provide a commentary and overview of the field. We suggest that the field has been through three revolutions that have focussed on specific methodological developments; disease dynamics and heterogeneity, advanced computing and inference, and complexity and application to the real-world. Infectious disease dynamics and heterogeneity dominated until the 1980s where the use of analytical models illustrated fundamental concepts such as herd immunity. The second revolution embraced the integration of data with models and the increased use of computing. From the turn of the century an emergence of novel datasets enabled improved modelling of real-world complexity. The emergence of more complex data that reflect the real-world heterogeneities in transmission resulted in the development of improved inference methods such as particle filtering. Each of these three revolutions have always kept the understanding of infectious disease spread as its motivation but have been developed through the use of new techniques, tools and the availability of data. We conclude by providing a commentary on what the next revoluition in infectious disease modelling may be.

Decision tree machine learning applied to bovine tuberculosis risk factors to aid disease control decision making.

Identifying and understanding the risk factors for endemic bovine tuberculosis (TB) in cattle herds is critical for the control of this disease. Exploratory machine learning techniques can uncover complex non-linear relationships and interactions within disease causation webs, and enhance our knowledge of TB risk factors and how they are interrelated. Classification tree analysis was used to reveal associations between predictors of TB in England and each of the three surveillance risk areas (High Risk, Edge, and Low Risk) in 2016, identifying the highest risk herds. The main classifying predictor for farms in England overall related to the TB prevalence in the 100 nearest cattle herds. In the High Risk and Edge areas it was the number of slaughterhouse destinations and in the Low Risk area it was the number of cattle tested in surveillance tests. How long ago the last confirmed incident was resolved was the most frequent classifier in trees; if within two years, leading to the highest risk group of herds in the High Risk and Low Risk areas. At least two different slaughterhouse destinations led to the highest risk group of herds in England, whereas in the Edge area it was a combination of no contiguous low-risk neighbours (i.e. in a 1 km radius) and a minimum proportion of 6-23 month-old cattle in November. A threshold value of prevalence in 100 nearest neighbours increased the risk in all areas, although the value was specific to each area. Having low-risk contiguous neighbours reduced the risk in the Edge and High Risk areas, whereas high-risk ones increased the risk in England overall and in the Edge area specifically. The best classification tree models informed multivariable binomial logistic regression models in each area, adding statistical inference outputs. These two approaches showed similar predictive performance although there were some disparities regarding what constituted high-risk predictors. Decision tree machine learning approaches can identify risk factors from webs of causation: information which may then be used to inform decision making for disease control purposes.

Using species distribution models to predict potential hot-spots for Rift Valley Fever establishment in the United Kingdom.

Vector borne diseases are a continuing global threat to both human and animal health. The ability of vectors such as mosquitos to cover large distances and cross country borders undetected provide an ever-present threat of pathogen spread. Many diseases can infect multiple vector species, such that even if the climate is not hospitable for an invasive species, indigenous species may be susceptible and capable of transmission such that one incursion event could lead to disease establishment in these species. Here we present a consensus modelling methodology to estimate the habitat suitability for presence of mosquito species in the UK deemed competent for Rift Valley fever virus (RVF) and demonstrate its application in an assessment of the relative risk of establishment of RVF virus in the UK livestock population. The consensus model utilises observed UK mosquito surveillance data, along with climatic and geographic prediction variables, to inform six independent species distribution models; the results of which are combined to produce a single prediction map. As a livestock host is needed to transmit RVF, we then combine the consensus model output with existing maps of sheep and cattle density to predict the areas of the UK where disease is most likely to establish in local mosquito populations. The model results suggest areas of high suitability for RVF competent mosquito species across the length and breadth of the UK. Notable areas of high suitability were the South West of England and coastal areas of Wales, the latter of which was subsequently predicted to be at higher risk for establishment of RVF due to higher livestock densities. This study demonstrates the applicability of outputs of species distribution models to help predict hot-spots for risk of disease establishment. While there is still uncertainty associated with the outputs we believe that the predictions are an improvement on just using the raw presence points from a database alone. The outputs can also be used as part of a multidisciplinary approach to inform risk based disease surveillance activities.

Trend analysis of imported malaria in London; observational study 2000 to 2014.

BACKGROUND: We describe trends of malaria in London (2000-2014) in order to identify preventive opportunities and we estimated the cost of malaria admissions (2009/2010-2014/2015). METHODS: We identified all cases of malaria, resident in London, reported to the reference laboratory and obtained hospital admissions from Hospital Episode Statistics. RESULTS: The rate of malaria decreased (19.4[2001]-9.1[2014] per 100,000). Males were over-represented (62%). Cases in older age groups increased overtime. The rate was highest amongst people of Black African ethnicity followed by Indian, Pakistani, Bangladeshi ethnicities combined (103.3 and 5.5 per 100,000, respectively). The primary reason for travel was visiting friends and relatives (VFR) in their country of origin (69%), mostly sub-Saharan Africa (92%). The proportion of cases in VFRs increased (32%[2000]-50%[2014]) and those taking chemoprophylaxis decreased (36%[2000]-14%[2014]). The overall case fatality rate was 0.3%. We estimated the average healthcare cost of malaria admissions to be just over £1 million per year. CONCLUSION: Our study highlighted that people of Black African ethnicity, travelling to sub-Saharan Africa to visit friends and relatives in their country of origin remain the most affected with also a decline in chemoprophylaxis use. Malaria awareness should focus on this group in order to have the biggest impact but may require new approaches.

Crossing the boundary between the Balpha and Bbeta mating-type loci in Schizophyllum commune.

In this study, genes of the Schizophyllum commune Balpha and Bbeta mating-type loci are shown to be within a few kilobases of each other. The region between the nearest Balpha and Bbeta genes contains many short direct repeats. Predicted amino acid sequences and activity spectra of three pheromones encoded in the Balpha3 mating-type specificity are presented along with a re-evaluation of pheromone activity of many previously reported S. commune lipopeptide pheromones. This analysis showed that S. commune pheromones belong to five subtypes. Several pheromones activate both a Bbeta receptor and a Balpha receptor, a phenomenon previously unrecognized. Clues from mating tests and DNA hybridization led to the cloning of bar8, the gene encoding the Balpha8 pheromone receptor, Bar8. Bar8 is similar in sequence to Bbr1, the Bbeta1 pheromone receptor, and functionally identical to it. These data begin to elucidate the enigmatic recombination patterns previously encountered at the B mating-type complex.