Heterologous expression in Escherichia coli of Neurospora crassa neutral trehalase as an active enzyme

Neutral trehalase from Neurospora crassa was expressed in Escherichia coli as a polypeptide of 84 kDa in agreement with the theoretical size calculated from the corresponding cDNA. The recombinant neutral trehalase, purified by affinity chromatography exhibited a specific activity of 80–150 mU/mg protein. Optima of pH and temperature were 7.0 and 30 °C, respectively. The enzyme was absolutely specific for trehalose, and was quite sensitive to incubation at 40 °C. The recombinant enzyme was totally dependent on calcium, and was inhibited by ATP, copper, silver, aluminium and cobalt. K_M was 42 mM, and V_max was 30.6 nmol of glucose/min. The recombinant protein was phosphorylated by cAMP-dependent protein kinase, but not significantly activated. Immunoblotting with polyclonal antiserum prepared against the recombinant protein showed that neutral trehalase protein levels increased during exponential phase of N. crassa growth and dropped at the stationary phase. This is the first report of a neutral trehalase produced in E. coli with similar biochemical properties described for fungi native neutral trehalases, including calcium-dependence.     

DIATMOD: diatom predictive model for quality assessment of Portuguese running waters

A predictive model for diatoms based on an adaptation of the River Invertebrate Prediction and Classification System/Australian River Assessment System approaches was evaluated as an effective tool for measuring stream ecological quality. This type of model was originally developed in UK and later in Australia and is extensively used to obtain ecological quality assessments with macroinvertebrates. The first step for the model construction was the definition of six consistent reference biological groups (ANOSIM: Global R = 0.77; P < 0.001) after classification (UPGMA) and ordination (nMDS) of 120 reference sites containing 254 different diatom taxa (species and infra-specific rank). A set of five environmental variables (slope, hydrological regime, mean annual temperature, mean annual precipitation and alkalinity) correctly discriminated 67% of reference sites (stepwise forward discriminant analysis, Jackknifed classification). The model was statistically accurate (slope = 1.07, intercept = −0.68, R ² = 0.65) and was validated by an independent set of reference data (13 reference sites; 70% correct answers). In addition, the model was tested by running data from 113 potentially disturbed sites. The model (DIATMOD) was well correlated with a general abiotic degradation gradient (Spearman correlations, R ² = 0.53, P < 0.001; and PCA analysis) and also with several specific pressure variables such as nitrates, phosphates, urban area, connectivity and land use (P < 0.001). Most diatom indices assess chemical contamination and we showed here that through predictive modelling the potential of diatoms as bioindicators increases as they also responded to hydromorphological changes. Further investigation on model potential consists in: testing different probability levels for taxa inclusion (here it was >0.5 as the most common models); comparing with alternative classification systems; assessing the influence of substrate type and seasonal variation in assessments.     

Population structure and adaptation of a bacterial pathogen in California grapevines

Xylella fastidiosa subsp. fastidiosa causes Pierce's disease of grapevine (PD) and has been present in California for over a century. A singly introduced genotype spread across the state causing large outbreaks and damaging the grapevine industry. This study presents 122 X. fastidiosa subsp. fastidiosa genomes from symptomatic grapevines, and explores pathogen genetic diversity associated with PD in California. A total of 5218 single-nucleotide polymorphisms (SNPs) were found in the dataset. Strong population genetic structure was found; isolates split into five genetic clusters divided into two lineages. The core/soft-core genome constituted 41.2% of the total genome, emphasizing the high genetic variability of X. fastidiosa genomes. An ecological niche model was performed to estimate the environmental niche of the pathogen within California and to identify key climatic factors involved in dispersal. A landscape genomic approach was undertaken aiming to link local adaptation to climatic factors. A total of 18 non-synonymous polymorphisms found to be under selective pressures were correlated with at least one environmental variable highlighting the role of temperature, precipitation and elevation on X. fastidiosa adaptation to grapevines in California. Finally, the contribution to virulence of three of the genes under positive selective pressure and of one recombinant gene was studied by reverse genetics.     

Cultivable fungi present in deep-sea sediments of Antarctica: taxonomy,diversity, and bioprospecting of bioactive compounds

Extremophiles - We accessed the culturable mycobiota present in marine sediments at different depths in Antarctica Ocean. Acremonium fusidioides, Penicillium allii-sativi, Penicillium chrysogenum,...     

Mosquito community composition and abundance at contrasting sites in northern South Africa, 2014–2017

Most data on species associations and vector potential of mosquitoes in relation to arboviral infections in South Africa date back from the 1940s to late 1990s. Contextual information crucial for disease risk management and control, such as the sampling effort, diversity, abundance, and distribution of mosquitoes in large parts of South Africa still remains limited. Adult mosquitoes were collected routinely from two horse farms in Gauteng Province; two wildlife reserves in Limpopo Province, at Orpen Gate in Kruger National Park (KNP) and Mnisi Area in Mpumalanga Province between 2014–2017, using carbon dioxide‐baited light and tent traps. Mosquito diversity and richness are greater in untransformed natural and mixed rural settings. In untransformed wilderness areas, the most dominant species were Culex poicilipes, Anopheles coustani, and Aedes mcintoshi, while in mixed rural settings such as the Mnisi area, the two most abundant species were Cx. poicilipes and Mansonia uniformis. However, in peri‐urban areas, Cx. theileri, Cx. univittatus, and Cx. pipiens sensu lato were the most dominant. Aedes aegypti, Ae. mcintoshi, Ae. metallicus, Ae. vittatus, Cx. pipiens s.l., Cx. theileri, and Cx. univittatus had the widest geographical distribution in northern South Africa. Also collected were Anopheles arabiensis and An. vaneedeni, both known malaria vectors in South Africa. Arbovirus surveillance and vector control programs should be augmented in mixed rural and peri‐urban areas where the risk for mosquito‐borne disease transmission to humans and domestic stock is greater.     

Leprosy in a prison population: A new active search strategy and a prospective clinical analysis

BackgroundThis study evaluates an active search strategy for leprosy diagnosis based on responses to a Leprosy Suspicion Questionnaire (LSQ), and analyzing the clinical, immunoepidemiological and follow-up aspects for individuals living in a prison population.MethodsA cross-sectional study based on a questionnaire posing 14 questions about leprosy symptoms and signs that was distributed to 1,400 prisoners. This was followed by dermatoneurological examination, anti-PGL-I serology and RLEP-PCR. Those without leprosy were placed in the Non-leprosy Group (NLG, n = 1,216) and those diagnosed with clinical symptoms of leprosy were placed in the Leprosy Group (LG, n = 34).FindingsIn total, 896 LSQ were returned (64%), and 187 (20.9%) of the responses were deemed as positive for signs/symptoms, answering 2.7 questions on average. Clinically, 1,250 (89.3%) of the prisoners were evaluated resulting in the diagnosis of 34 new cases (LG), based on well-accepted clinical signs and symptoms, a new case detection rate of 2.7% within this population, while the NLG were comprised of 1,216 individuals. The confinement time medians were 39 months in the LG while it was 36 months in the NLG (p>0.05). The 31 leprosy cases who responded to the questionnaire (LSQ+) had an average of 1.5 responses. The symptoms “anesthetized skin area” and “pain in nerves” were most commonly mentioned in the LG while “tingling, numbness in the hands/feet”, “sensation of pricks and needles”, “pain in nerves” and “spots on the skin” responses were found in more than 30% of questionnaires in the NLG. Clinically, 88.2% had dysesthetic macular skin lesions and 97.1% presented some peripheral nerve impairment, 71.9% with some degree of disability. All cases were multibacillary, confirming a late diagnosis. Anti-PGL-I results in the LG were higher than in the NLG (p<0.0001), while the RLEP-PCR was positive in 11.8% of the patients.InterpretationOur findings within the penitentiary demonstrated a hidden prevalence of leprosy, although the individuals diagnosed were likely infected while living in their former communities and not as a result of exposure in the prison. The LSQ proved to be an important screening tool to help identify leprosy cases in prisons.     

Validation of a Dehydroepiandrosterone-Sulfate Assay in Three Platyrrhine Primates (Alouatta caraya,Aotus azarae infulatus,and Sapajus apella)

International Journal of Primatology - The hormone dehydroepiandrosterone (DHEA) and its sulfate ester (DHEAS) are the most abundant circulating steroids in human and some nonhuman primates, and...     

Antitumor activity via apoptotic cell death pathway of water soluble copper(II) complexes: effect of the diamino unit on selectivity against lung cancer NCI-H460 cell line

BioMetals - The cytotoxicity against five human tumor cell lines (THP-1, U937, Molt-4, Colo-205 and NCI-H460) of three water soluble copper(II) coordination compounds containing the ligands...