Immunoglobulin A as a Key Immunological Molecular Signature of Post-COVID-19 Conditions.

COVID-19 has infected humans worldwide, causing millions of deaths or prolonged symptoms in survivors. The transient or persistent symptoms after SARS-CoV-2 infection have been defined as post-COVID-19 conditions (PCC). We conducted a study of 151 Brazilian PCC patients to analyze symptoms and immunoglobulin profiles, taking into account sex, vaccination, hospitalization, and age. Fatigue and myalgia were the most common symptoms, and lack of vaccination, hospitalization, and neuropsychiatric and metabolic comorbidities were relevant to the development of PCC. Analysis of serological immunoglobulins showed that IgA was higher in PCC patients, especially in the adult and elderly groups. Also, non-hospitalized and hospitalized PCC patients produced high and similar levels of IgA. Our results indicated that the detection of IgA antibodies against SARS-CoV-2 during the course of the disease could be associated with the development of PCC and may be an immunological signature to predict prolonged symptoms in COVID-19 patients.

Providing baseline data for conservation-Heart rate monitoring in captive scimitar-horned oryx.

Heart rate biologging has been successfully used to study wildlife responses to natural and human-caused stressors (e.g., hunting, landscape of fear). Although rarely deployed to inform conservation, heart rate biologging may be particularly valuable for assessing success in wildlife reintroductions. We conducted a case study for testing and validating the use of subcutaneous heart rate monitors in eight captive scimitar-horned oryx (Oryx dammah), a once-extinct species that is currently being restored to the wild. We evaluated biologger safety and accuracy while collecting long-term baseline data and assessing factors explaining variation in heart rate. None of the biologgers were rejected after implantation, with successful data capture for 16-21 months. Heart rate detection accuracy was high (83%-99%) for six of the individuals with left lateral placement of the biologgers. We excluded data from two individuals with a right lateral placement because accuracies were below 60%. Average heart rate for the six scimitar-horned oryx was 60.3 ± 12.7 bpm, and varied by about 12 bpm between individuals, with a minimum of 31 bpm and a maximum of 188 bpm across individuals. Scimitar-horned oryx displayed distinct circadian rhythms in heart rate and activity. Heart rate and activity were low early in the morning and peaked near dusk. Circadian rhythm in heart rate and activity were relatively unchanged across season, but hourly averages for heart rate and activity were higher in spring and summer, respectively. Variation in hourly heart rate averages was best explained by a combination of activity, hour, astronomical season, ambient temperature, and an interaction term for hour and season. Increases in activity appeared to result in the largest changes in heart rate. We concluded that biologgers are safe and accurate and can be deployed in free-ranging and reintroduced scimitar-horned oryx. In addition to current monitoring practices of reintroduced scimitar-horned oryx, the resulting biologging data could significantly aid in 1) evaluating care and management action prior to release, 2) characterizing different animal personalities and how these might affect reintroduction outcomes for individual animals, and 3) identifying stressors after release to determine their timing, duration, and impact on released animals. Heart rate monitoring in released scimitar-horned oryx may also aid in advancing our knowledge about how desert ungulates adapt to extreme environmental variation in their habitats (e.g., heat, drought).

Comparison of the Clinical-Functional Vulnerability Index and the frailty phenotype for the identification of falls in older individuals: A cross-sectional study.

BACKGROUND: Frailty increases the risk of falls, disability and death in older adults. The Cardiovascular Health Study identified a frailty phenotype (the Fried Phenotype) that was primarily based on physical domains. Instruments that incorporate additional domains (e.g., cognitive, disability or mood) may more accurately identify falls. OBJECTIVES: The study aimed i) to evaluate the association between falls and the number of phenotypes identified by the Fried Phenotype and CFVI-20 scores and ii) to compare the strength of the association between falls and each frailty instrument. METHODS: This study used the CFVI-20 and the Fried Phenotype and reported falls during the last twelve months. Logistic regression models, odds ratios (ORs), and ROC curves were used to identify associations and perform comparisons (p<0.05). The reporting of the study followed the Strobe guidelines. RESULTS: This study included 1,826 individuals (mean 70.9 (SD 7.3) years old). Prevalence of pre-frailty and low vulnerability was high (72% and 69%) and comparable between frailty instruments. The number of Fried phenotypes increased the odds of having fallen in the past 12 months (OR: 1.5 to 29.5) and the CFVI-20 scores (11% increase/unit change). The CFVI-20 identified falls more accurately than the Fried Phenotype (AUC: 0.68 vs. 0.60, p < 0.001). CONCLUSIONS: The number of phenotypes and the CFVI-20 scores were associated with falls; continuous scores identified falls more accurately than categorical classifications. The CFVI-20 was more strongly associated with falls in community-dwelling older adults than the Fried Phenotype.

Schools reopening and the COVID-19 pandemic: a case study from Macaé, Rio de Janeiro, Brazil.

Since the first reported case of COVID-19 in Brazil, the public and private educational system started to close. Up to November 2020, scientific discussions about the return of schooling activities have been rarely performed by the national scientific community and police-makers. The great delay of school returning in Brazil contrasts with successful international strategies of school reopening worldwide and seems counterintuitive with the reopening of non-essential activities. Here, important issues to be considered before and during school reopening are reviewed and discussed. COVID-19 testing is essential to avoid disease spreading, but high cost of individual RT-qPCRs impairs an extensive testing strategy for school returning. To reduce costs and increase the speed of diagnosis, we tested the efficiency of a pooled-sample PCR strategy in a cohort of the educational staff in the city of Macaé/RJ, finding five asymptomatic individuals (0,66%) among the 754 people tested. Thus, a polled-sample PCR testing strategy of the educational staff might prevent infection spreading in schools at a reasonable cost. We discuss how our test strategy could be coupled with internationally recognized safety rules to allow for a safe school return and how countries from different world regions are dealing with educational activities during COVID-19 pandemic.

Association between Domains of the Clinical-Functional Vulnerability Index and Falls History in Older Adults: A Cross-Sectional Study.

Objectives: The study aimed to determine which domains, sets, and isolated or combined questions of the Clinical-Functional Vulnerability Index (CFVI-20) are associated with falls history in older adults. Methods: Instruments used were the CFVI-20 assessment and reported falls during the last year. The receiver operating characteristics (ROC) curves identified the performance of the CFVI-20 domains and questions in identifying older adults with and without falls history, while logistic regression identified relevant questions to identify fall history. Results: This study included 1725 individuals (71.9 ± 7.3 years). The area under the curve (AUC) between the CFVI-20 and fall history was 0.69. The mobility domain presented the largest AUC (0.71; p < 0.01), and most isolated domains showed low AUCs (0.51 to 0.58). Isolated questions were limited to identifying fallers. The regression analysis identified 7 questions of the CFVI-20 with falls. Conclusions: The CFVI-20 general score identified older adults with a fall history. When considered in isolation, most domains were limited to identifying falls, except for the mobility domain. Combining the CFVI-20 questions enabled identification of fallers.

Systemic cytokines, chemokines and growth factors reveal specific and shared immunological characteristics in infectious cardiomyopathies.

Heart disease is a major cause of death worldwide. Chronic Chagas cardiomyopathy (CCC) caused by infection with Trypanosoma cruzi leading to high mortality in adults, and rheumatic heart disease (RHD), resulting from infection by Streptococcus pyogenes affecting mainly children and young adults, are amongst the deadliest heart diseases in low-middle income countries. Despite distinct etiology, the pathology associated with both diseases is a consequence of inflammation. Here we compare systemic immune profile in patients with these cardiopathies, to identify particular and common characteristics in these infectious heart diseases. We evaluated the expression of 27 soluble factors, employing single and multivariate analysis combined with machine-learning approaches. We observed that, while RHD and CCC display higher levels of circulating mediators than healthy individuals, CCC is associated with stronger immune activation as compared to RHD. Despite distinct etiologies, univariate analysis showed that expression of TNF, IL-17, IFN-gamma, IL-4, CCL4, CCL3, CXCL8, CCL11, CCL2, PDGF-BB were similar between CCC and RHD, consistent with their inflammatory nature. Network analysis revealed common inflammatory pathways between CCC and RHD, while highlighting the broader reach of the inflammatory response in CCC. The final multivariate model showed a 100% discrimination power for the combination of the cytokines IL-12p70, IL-1Ra, IL-4, and IL-7 between CCC and RHD groups. Thus, while clear immunological distinctions were identified between CCC and RHD, similarities indicate shared inflammatory pathways in these infectious heart diseases. These results contribute to understanding the pathogenesis of CCC and RHD and may impact the design of immune-based therapies for these and other inflammatory cardiopathies that may also share immunological characteristics.

Thirty years of recombinant BCG: new trends for a centenary vaccine.

Introduction: Global perception of the potential for Bacille Calmette-Guérin (BCG), and consequently recombinant BCG (rBCG), in a variety of prophylactic and therapeutic applications has been increasing. A century of information on BCG, and three decades of experience with rBCG, has generated solid knowledge in this field.Area covered: Here, we review the current state of knowledge of BCG and rBCG development. Molecular tools have facilitated the expression of a variety of molecules in BCG, with the aim of improving its efficacy as a tuberculosis vaccine, generating polyvalent vaccines against other pathogens, including viruses, bacteria, and parasites, and developing immunotherapy approaches against noninvasive bladder cancer. BCG's recently appraised heterologous effects and prospects for expanding its application to other diseases are also addressed.Expert opinion: There are high expectations for new tuberculosis vaccines currently undergoing advanced clinical trials, which could change the prospects of the field. Systems biology could reveal effective biomarkers of protection, which would greatly support vaccine development. The development of appropriate large-scale production processes would further support implementation of new vaccines and rBCG products. The next few years should consolidate the broader applications of BCG and produce insights into improvements using the recombinant BCG technology.

Inhibition of Aedes aegypti DNA topoisomerase II by etoposide: Impact on survival and morphology of larvae and pupae.

DNA topoisomerase II enzymes maintain DNA stability during vital processes, such as genome replication, transcription and chromosomal segregation during mitosis and meiosis. In the present work, we analyzed functional aspects of the DNA topoisomerase II (AeTopII) enzyme of the mosquito Aedes aegypti. Here, we show that AeTopII mRNA is expressed at all stages of mosquito development. By in situ hybridization, we found that the AeTopII mRNA is concentrated along the ovarian follicular cells as well as in the region of the follicles. The observed expression profiles likely reflect increased topoisomerase II cellular requirements due to the intense ovarian growth and egg production following blood feeding in Ae. aegypti females. The drug etoposide, a classic inhibitor of topoisomerase II, was used for in vivo testing with 2nd stage larvae, in order to investigate the functional importance of this enzyme in Ae. aegypti survival and development. Inhibition of topoisomerase II activity with etoposide concentrations ranging from 10 to 200 µM did not leads to the immediate death of larvae. However, after 10 days of observation, etoposide treatments resulted in 30-40% decrease in survival, in a dose dependent manner, with persisting larvae and pupae presenting incomplete development, as well as morphological abnormalities. Also, approximately 50% of the treated larvae did not reach the pupal stage. Thus, we conclude that AeTopII is a vital enzyme in the development of Ae. aegypti and its sensitivity to inhibitors should be explored for potential chemical agents to be used in vector control.

Rapid local adaptation in both sexual and asexual invasive populations of monkeyflowers (Mimulus spp.).

BACKGROUND AND AIMS: Traditionally, local adaptation has been seen as the outcome of a long evolutionary history, particularly with regard to sexual lineages. By contrast, phenotypic plasticity has been thought to be most important during the initial stages of population establishment and in asexual species. We evaluated the roles of adaptive evolution and phenotypic plasticity in the invasive success of two closely related species of invasive monkeyflowers (Mimulus) in the UK that have contrasting reproductive strategies: M. guttatus combines sexual (seeds) and asexual (clonal growth) reproduction while M. × robertsii is entirely asexual. METHODS: We compared the clonality (number of stolons), floral and vegetative phenotype, and phenotypic plasticity of native (M. guttatus) and invasive (M. guttatus and M. × robertsii) populations grown in controlled environment chambers under the environmental conditions at each latitudinal extreme of the UK. The goal was to discern the roles of temperature and photoperiod on the expression of phenotypic traits. Next, we tested the existence of local adaptation in the two species within the invasive range with a reciprocal transplant experiment at two field sites in the latitudinal extremes of the UK, and analysed which phenotypic traits underlie potential local fitness advantages in each species. KEY RESULTS: Populations of M. guttatus in the UK showed local adaptation through sexual function (fruit production), while M. × robertsii showed local adaptation via asexual function (stolon production). Phenotypic selection analyses revealed that different traits are associated with fitness in each species. Invasive and native populations of M. guttatus had similar phenotypic plasticity and clonality. M. × robertsii presents greater plasticity and clonality than native M. guttatus, but most populations have restricted clonality under the warm conditions of the south of the UK. CONCLUSIONS: This study provides experimental evidence of local adaptation in a strictly asexual invasive species with high clonality and phenotypic plasticity. This indicates that even asexual taxa can rapidly (<200 years) adapt to novel environmental conditions in which alternative strategies may not ensure the persistence of populations.

Thirty years of recombinant BCG: new trends for a centenary vaccine

Introduction: Global perception of the potential for Bacille Calmette-Guérin (BCG), and consequently recombinant BCG (rBCG), in a variety of prophylactic and therapeutic applications has been increasing. A century of information on BCG, and three decades of experience with rBCG, has generated solid knowledge in this field. Area covered: Here, we review the current state of knowledge of BCG and rBCG development. Molecular tools have facilitated the expression of a variety of molecules in BCG, with the aim of improving its efficacy as a tuberculosis vaccine, generating polyvalent vaccines against other pathogens, including viruses, bacteria, and parasites, and developing immunotherapy approaches against noninvasive bladder cancer. BCG’s recently appraised heterologous effects and prospects for expanding its application to other diseases are also addressed. Expert opinion: There are high expectations for new tuberculosis vaccines currently undergoing advanced clinical trials, which could change the prospects of the field. Systems biology could reveal effective biomarkers of protection, which would greatly support vaccine development. The development of appropriate large-scale production processes would further support implementation of new vaccines and rBCG products. The next few years should consolidate the broader applications of BCG and produce insights into improvements using the recombinant BCG technology.

Subarachnoid neurocysticercosis and an intracranial infectious aneurysm: case report.

Infectious intracranial aneurysms (IIAs) represent 2%-6% of all intracranial aneurysms and, classically, have been associated with bacterial or fungal agents. The authors report the case of a 42-year-old woman who presented with a typical history of subarachnoid hemorrhage. Digital subtraction angiography (DSA) showed an aneurysmal dilatation on the frontal M2 segment of the left middle cerebral artery (MCA). The patient was treated surgically, and multiple cysts were found in the left carotid and sylvian cisterns, associated with a dense inflammatory exudate that involved the MCA. The cysts were removed, and a fusiform aneurysmal dilatation was identified. The lesion was not amenable to direct clipping, so the authors wrapped it. Histopathological analysis of the removed cysts revealed the typical pattern of subarachnoid neurocysticercosis. The patient received cysticidal therapy with albendazole and corticosteroids, and she recovered uneventfully. Follow-up DSA performed 6 months after surgery showed complete resolution of the aneurysm. The authors performed a review of the literature and believe that there is sufficient evidence to affirm that the subarachnoid form of neurocysticercosis may lead to the development of an IIA and that Taenia solium should be listed among the possible etiological agents of IIAs, along with bacterial and fungal agents.

Exopolysaccharides from Aspergillus terreus: Production, chemical elucidation and immunoactivity.

Aspergillus terreus, a fungus commonly used in pharmaceutical industry to produce lovastatin and other secondary metabolites, has been reported to have beneficial biological properties. In this study the exopolysaccharides (AT-EPS) produced by A. terreus were evaluated as potential modulators of certain functions of macrophages. The production parameters for EPS obtained from the liquid culture broth of the studied fungus were optimized using response surface methodology (RSM) and indicated good correlation between the experimental and predicted values. The optimum conditions for AT-EPS extraction included fermentation at 28 °C, pH 8.79, under 98 rpm of agitation, using 2.39% glucose (carbon source) and 0.957% ammonium nitrate (nitrogen source). Under these optimized conditions, AT-EPS production was 1.34 g/L medium. The chemical analyses showed that AT-EPS was composed by mannose (Man; 40.5 mol%), galactose (Gal; 35.2 mol%), and glucose (Glc; 24.3 mol%), and the spectroscopic (FTIR; NMR) and methylation analyses indicated the presence of galactomannans, ß-1,3-glucans, and glycogen-like glucans. AT-EPS was tested on murine macrophages to verify its immunoactivity and the treated cells were able to produce nitric oxide, superoxide anion, TNF-α and interleukin 6 similarly to the positive control cells. Furthermore, the macrophages treated with AT-EPS showed activated-like morphological alterations.

Statistical prediction of interactions between low concentrations of inhibitors on yeast cells responses added to the SD-medium at low pH values.

BACKGROUND: In the present work, the main inhibitors of the yeast cells (vanillin, furfural, formic, and levulinic acid) were generated by pretreatments or hydrolysis (sulfuric acid or enzymes) to convert reducing sugars into ethanol. Inhibitors were added at increasing concentrations to the SD-medium containing yeast extract while negative effects on yeast cells were observed. Statistical analyses were applied to predict and interpret results related to biomass production. RESULTS: Inhibitors affected productivities and yields of biomass and ethanol when added to SD-medium. Based on the 23 full-central-composite design, "predicted" and "observed" values of ethanol and biomass were obtained in presence of the major inhibitors, which were acetic acid, formic acid, and levulinic acids. Increases in biomass and ethanol production are described in the Response surface graphs (RSM graphs) that resulted from multiple interactions between inhibitors. Positive interactions between the inhibitors occurred at low concentrations and pH values. The results were experimentally validated. CONCLUSIONS: Statistical analysis is an extremely useful tool for predicting data during process monitoring, while re-adjustments of conditions can be performed, whenever necessary. In addition, the development of new strains of yeast with high tolerance to biomass inhibitors will have a major impact on the production of second-generation ethanol. Increases in fermentation activity of the yeast Saccharomyces cerevisiae in a mixture containing low concentrations of inhibitors were observed.

Hydrated Electron Generation by Excitation of Copper Localized Surface Plasmon Resonance.

Hydrated electrons are important in radiation chemistry and charge-transfer reactions, with applications that include chemical damage of DNA, catalysis, and signaling. Conventionally, hydrated electrons are produced by pulsed radiolysis, sonolysis, two-ultraviolet-photon laser excitation of liquid water, or photodetachment of suitable electron donors. Here we report a method for the generation of hydrated electrons via single-visible-photon excitation of localized surface plasmon resonances (LSPRs) of supported sub-3 nm copper nanoparticles in contact with water. Only excitations at the LSPR maximum resulted in the formation of hydrated electrons, suggesting that plasmon excitation plays a crucial role in promoting electron transfer from the nanoparticle into the solution. The reactivity of the hydrated electrons was confirmed via proton reduction and concomitant H2 evolution in the presence of a Ru/TiO2 catalyst.

Weak whole-plant trait coordination in a seasonally dry South American stressful environment.

A core question involving both plant physiology and community ecology is whether traits from different organs are coordinated across species, beyond pairwise trait correlations. The strength of within-community trait coordination has been hypothesized to increase along gradients of environmental harshness, due to the cost of adopting ecological strategies out of the viable niche space supported by the abiotic conditions. We evaluated the strength of trait relationship and coordination in a stressful environment using 21 leaf and stem traits of 21 deciduous and evergreen woody species from a heath vegetation growing on coastal sandy plain in northeastern South America. The study region faces marked dry season, high soil salinity and acidity, and poor nutritional conditions. Results from multiple factor analyses supported two weak and independent axes of trait coordination, which accounted for 25%-29% of the trait variance using phylogenetically independent contrasts. Trait correlations on the multiple factor analyses main axis fit well with the global plant economic spectrum, with species investing in small leaves and dense stems as opposed to species with softer stems and large leaves. The species' positions on the main functional axis corresponded to the competitor-stress-tolerant side of Grime's CSR triangle of plant strategies. The weak degree of trait coordination displayed by the heath vegetation species contradicted our expectation of high trait coordination in stressful environmental habitats. The distinct biogeographic origins of the species occurring in the study region and the prevalence of a regional environmental filter coupled with local homogeneous conditions could account for prevalence of trait independence we observed.

Randomized controlled study comparing 2 surgical techniques for decompressive craniectomy: with watertight duraplasty and without watertight duraplasty.

OBJECTIVE: Decompressive craniectomy (DC) is a widely used procedure in neurosurgery; however, few studies focus on the best surgical technique for the procedure. The authors' objective was to conduct a prospective randomized controlled trial comparing 2 techniques for performing DC: with watertight duraplasty and without watertight duraplasty (rapid-closure DC). METHODS: The study population comprised patients ranging in age from 18 to 60 years who were admitted to the Neurotrauma Service of the Hospital da Restauração with a clinical indication for unilateral decompressive craniectomy. Patients were randomized by numbered envelopes into 2 groups: with watertight duraplasty (control group) and without watertight duraplasty (test group). After unilateral DC was completed, watertight duraplasty was performed in the control group, while in the test group, no watertight duraplasty was performed and the exposed parenchyma was covered with Surgicel and the remaining dura mater. Patients were then monitored daily from the date of surgery until hospital discharge or death. The primary end point was the incidence of surgical complications (CSF leak, wound infection, brain abscess, or subgaleal fluid collections). The following were analyzed as secondary end points: clinical outcome (analyzed using the Glasgow Outcome Scale [GOS]), surgical time, and hospital costs. RESULTS: Fifty-eight patients were enrolled, 29 in each group. Three patients were excluded, leaving 27 in the test group and 28 in the control group. There were no significant differences between groups regarding age, Glasgow Coma Scale score at the time of surgery, GOS score, and number of postoperative follow-up days. There were 9 surgical complications (5 in the control group and 4 in the test group), with no significant differences between the groups. The mean surgical time in the control group was 132 minutes, while in the test group the average surgical time was 101 minutes, a difference of 31 minutes (p = 0.001). The mean reduction in total cost was $420.00 USD (a 23.4% reduction) per procedure in the test group. CONCLUSIONS: Rapid-closure DC without watertight duraplasty is a safe procedure. It is not associated with a higher incidence of surgical complications (CSF leak, wound infection, brain abscess, or subgaleal fluid collections), and it decreased surgical time by 31 minutes on average. There was also a hospital cost reduction of $420.00 USD (23.4% reduction) per procedure. Clinical trial registration no.: NCT02594137 (clinicaltrials.gov).

Normal limits of the electrocardiogram derived from a large database of Brazilian primary care patients.

BACKGROUND: Knowledge of the normal limits of the electrocardiogram (ECG) is mandatory for establishing which patients have abnormal ECGs. No studies have assessed the reference standards for a Latin American population. Our aim was to establish the normal ranges of the ECG for pediatric and adult Brazilian primary care patients. METHODS: This retrospective observational study assessed all the consecutive 12-lead digital electrocardiograms of primary care patients at least 1 year old in Minas Gerais state, Brazil, recorded between 2010 and 2015. ECGs were excluded if there were technical problems, selected abnormalities were present or patients with selected self-declared comorbidities or on drug therapy. Only the first ECG from patients with multiple ECGs was accepted. The University of Glasgow ECG analysis program was used to automatically interpret the ECGs. For each variable, the 1st, 2nd, 50th, 98th and 99th percentiles were determined and results were compared to selected studies. RESULTS: A total of 1,493,905 ECGs were recorded. 1,007,891 were excluded and 486.014 were analyzed. This large study provided normal values for heart rate, P, QRS and T frontal axis, P and QRS overall duration, PR and QT overall intervals and QTc corrected by Hodges, Bazett, Fridericia and Framingham formulae. Overall, the results were similar to those from other studies performed in different populations but there were differences in extreme ages and specific measurements. CONCLUSIONS: This study has provided reference values for Latinos of both sexes older than 1 year. Our results are comparable to studies performed in different populations.

D-mannitol sensor based on molecularly imprinted polymer on electrode modified with reduced graphene oxide decorated with gold nanoparticles.

An electrochemical sensor for D-mannitol based on molecularly imprinted polymer on electrode modified with reduced graphene oxide decorated with gold nanoparticles was developed in this present work. The sensor was constructed for the first time via the electropolymerization of o-phenylenediamine (o-PD) over a surface containing reduced graphene oxide (RGO) and gold nanoparticles (AuNP) in the presence of D-mannitol molecules. The surface modification with AuNP/RGO-GCE facilitated the charge transfer processes of [Fe(CN)6]3-/4-, which was used as an electrochemical probe. It also contributed meaningfully towards the increase in the surface/volume ratio, creating more locations for imprinting, and providing greater sensitivity to the sensor. The MIP/AuNP/RGO-GCE sensor was characterized by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), scanning electron microscope (SEM), atomic force microscope (AFM) and X-ray Photoelectron Spectroscopy (XPS). Important parameters that exert control over the performance of the molecularly imprinted sensor (such as number of cycles, pH, monomer and template concentration and extraction and rebinding conditions) were investigated and optimized. The imprinting factor was 4.9, showing greater response to the D-mannitol molecule compared to the interfering molecules. The limit of detection, limit of quantification and amperometric sensitivity were 7.7×10-13molL-1, 2.6×10-12molL-1 and 3.9×1010µALmol-1 (n=3) respectively. The MIP/AuNP/RGO-GCE sensor was successfully applied towards the selective determination of D-mannitol in sugarcane vinasse, thus making it, in essence, a valuable tool for the accurate and reliable determination of this molecule.

Population genetics of self-incompatibility in a clade of relict cliff-dwelling plant species.

The mating systems of species in small or fragmented populations impact upon their persistence. Small self-incompatible (SI) populations risk losing S allele diversity, responsible for the SI response, by drift thereby limiting mate availability and leading to population decline or SI system breakdown. But populations of relict and/or endemic species have resisted these demographic conditions over long periods suggesting their mating systems have adapted. To address a lack of empirical data on this topic, we studied the SI systems of three relict cliff-dwelling species of Sonchus section Pustulati (Asteraceae): S. masguindalii, S. fragilis and S. pustulatus in the western Mediterranean region. We performed controlled pollinations within and between individuals to measure index of SI (ISI) expression and identify S alleles in multiple population samples. Sonchus masguindalii and S. pustulatus showed strong SI (ISI = 0.6-1.0) compared to S. fragilis (ISI = 0.1-0.7). Just five S alleles were estimated for Spanish S. pustulatus and a moderate 11-15 S alleles for Moroccan S. pustulatus and S. fragilis, respectively. The fact that autonomous fruit set was generally improved by active self-pollination in self-compatible S. fragilis suggests that individuals with weak SI can show a wide range of outcrossing levels dependent on the degree of self or outcross pollen that pollinators bear. We conclude that frequent S allele dominance interactions that mask the incompatibility interactions of recessive S alleles leading to higher mate availability and partial breakdown of SI leading to mixed mating, both contribute to reproductive resilience in this group.

Phylogeography of cliff-dwelling relicts with a highly narrow and disjunct distribution in the western Mediterranean.

PREMISE OF THE STUDY: The high biodiversity in the Baetic-Rifan hotspot of Mediterranean region is shaped by complex geological and climatic histories and has been a subject of recent intensive studies. However, very little is known about phylogenetic and biogeographic history of three rare and critically endangered cliff-dwelling species of Sonchus in section Pustulati in this region. METHODS: We investigated the genetic variation and phylogenetic relationships of populations based on nuclear (ITS/ETS) and plastid (3'trnL-ndhJ/psaI-accD) DNA sequences, and amplified fragment length polymorphisms (AFLPs). We performed a Bayesian relaxed molecular clock analysis with ITS data to estimate divergence times for major lineages. KEY RESULTS: ITS/ETS and AFLP phylogenies showed high concordance and contrasted with cpDNA data. The divergence between S. masguindalii and S. fragilis was dated at 5.48 Ma, between S. fragilis and S. pustulatus at 3.89 Ma, and between the Baetic and Rifan S. pustulatus at 1.18 Ma. Within each distribution area, AFLP data showed a relatively high genetic structuring and moderate genetic diversity, the latter being impoverished in the Baetic populations. CONCLUSIONS: Our results further confirm the hybrid origin of S. pustulatus, a critically endangered species. The origin and diversification of lineages appear to have occurred on the temporary land bridge that joined Iberian and North Africa during the Messinian Salinity Crisis (5.96-5.33 Ma) and the subsequent Zanclean flood that progressively refilled the Mediterranean Basin (5.33-3.60 Ma). The only Baetic populations of S. pustulatus most likely originated from the Rifan ones.