A new approach to dengue fatal cases diagnosis: NS1 antigen capture in tissues

Abstract/Background

Dengue is the most important arthropod borne viral disease worldwide in terms of morbidity and mortality and is caused by any of the four serotypes of dengue virus (DENV-1 to 4). Brazil is responsible for approximately 80% of dengue cases in the Americas, and since the introduction of dengue in 1986, a total of 5,944,270 cases have been reported including 21,596 dengue hemorrhagic fever and 874 fatal cases. DENV can infect many cell types and cause diverse clinical and pathological effects. The goal of the study was to investigate the usefulness of NS1 capture tests as an alternative tool to detect DENV in tissue specimens from previously confirmed dengue fatal cases (n = 23) that occurred in 2002 in Brazil.

Methodology/Principal Findings

A total of 74 tissue specimens were available: liver (n = 23), lung (n = 14), kidney (n = 04), brain (n = 10), heart (n = 02), skin (n = 01), spleen (n = 15), thymus (n = 03) and lymph nodes (n = 02). We evaluated three tests for NS1 antigen capture: first generation Dengue Early ELISA (PanBio Diagnostics), Platelia NS1 (BioRad Laboratories) and the rapid test NS1 Ag Strip (BioRad Laboratories). The overall dengue fatal case diagnosis based on the tissues analyzed by Dengue Early ELISA, Platelia NS1 and the NS1 Ag Strip was 34.7% (08/23), 60.8% (14/23) and 91.3% (21/23), respectively. The Dengue Early ELISA detected NS1 in 22.9% (17/74) of the specimens analyzed and the Platelia NS1 in 45.9% (34/74). The highest sensitivity (78.3%; 58/74) was achieved by the NS1 Ag Strip, and the differences in the sensitivities were statistically significant (p<0.05). The NS1 Ag Strip was the most sensitive in liver (91.3%; 21/23), lung (71.4%; 10/14), kidney (100%; 4/4), brain (80%; 8/10), spleen (66.6%, 10/15) and thymus (100%, 3/3) when compared to the other two ELISA assays.

Conclusions/Significance

This study shows the DENV NS1 capture assay as a rapid and valuable approach to postmortem dengue confirmation. With an increasing number of DHF and fatal cases, the availability of new approaches useful for cases confirmation plays an important tool for the disease surveillance.     

In silico and in vivo analysis of ABI3 and VAL2 genes during somatic embryogenesis of Coffea arabica: competence acquisition and developmental marker genes

Plant Cell, Tissue and Organ Culture (PCTOC) - The employment of biotechnology-based approaches such as somatic embryogenesis has been applied to several plants including Coffea sp. Despite the...     

Effects of Beta-Alanine Supplementation on Brain Homocarnosine/Carnosine Signal and Cognitive Function: An Exploratory Study

Objectives

Two independent studies were conducted to examine the effects of 28 d of beta-alanine supplementation at 6.4 g d^-1 on brain homocarnosine/carnosine signal in omnivores and vegetarians (Study 1) and on cognitive function before and after exercise in trained cyclists (Study 2).

Methods

In Study 1, seven healthy vegetarians (3 women and 4 men) and seven age- and sex-matched omnivores undertook a brain 1H-MRS exam at baseline and after beta-alanine supplementation. In study 2, nineteen trained male cyclists completed four 20-Km cycling time trials (two pre supplementation and two post supplementation), with a battery of cognitive function tests (Stroop test, Sternberg paradigm, Rapid Visual Information Processing task) being performed before and after exercise on each occasion.

Results

In Study 1, there were no within-group effects of beta-alanine supplementation on brain homocarnosine/carnosine signal in either vegetarians (p = 0.99) or omnivores (p = 0.27); nor was there any effect when data from both groups were pooled (p = 0.19). Similarly, there was no group by time interaction for brain homocarnosine/carnosine signal (p = 0.27). In study 2, exercise improved cognitive function across all tests (P<0.05), although there was no effect (P>0.05) of beta-alanine supplementation on response times or accuracy for the Stroop test, Sternberg paradigm or RVIP task at rest or after exercise.

Conclusion

28 d of beta-alanine supplementation at 6.4g d^-1 appeared not to influence brain homocarnosine/carnosine signal in either omnivores or vegetarians; nor did it influence cognitive function before or after exercise in trained cyclists.     

Environmental influences on and antimicrobial activity of the skin microbiota of Proceratophrys boiei (Amphibia,Anura) across forest fragments

The composition of the skin microbiota of amphibians is related to the biology of host species and environmental microbial communities. In this system, the environment serves as a microbial source and can modulate the hosted community. When habitats are fragmented and the environment disturbed, changes in the structure of this microbial community are expected. One important potential consequence of fragmentation is a compromised protective function of the microbiota against pathogenic microorganisms. In this study, the skin microbiota of the amphibian Proceratophrys boiei was characterized, evaluated for relationships with environmental variables and environmental sources of microbial communities, and its diversity evaluated for frog populations from fragmented and continuous forests. In addition, the antimicrobial activity of this skin community was studied in frogs from both forest types. Culture methods and 16S rRNA high‐throughput gene sequencing were used to characterize the microbial community and demonstrated that the skin microbiota of P. boiei is more closely related to the soil microbial communities than those inhabiting water bodies or fragment matrix, the unforested area around the forested fragment. The microbial diversity and abundance of P. boiei skin microbiota are different between continuous forests and fragments. This community is correlated with environmental variables, especially with temperature of microhabitat and distance to human dwelling. All individuals of P. boiei harbored bacteria capable of inhibiting the growth of pathogenic bacteria and different strains of the pathogenic fungus Batrachochytrium dendrobatidis, and a total of 27 bacterial genera were detected. The results of this study indicate that the persistence of populations of this species will need balanced and sustained interactions among host, microorganisms, and environment.     

Development of a panel of genome-wide ancestry informative markers to study admixture throughout the Americas

Most individuals throughout the Americas are admixed descendants of Native American, European, and African ancestors. Complex historical factors have resulted in varying proportions of ancestral contributions between individuals within and among ethnic groups. We developed a panel of 446 ancestry informative markers (AIMs) optimized to estimate ancestral proportions in individuals and populations throughout Latin America. We used genome-wide data from 953 individuals from diverse African, European, and Native American populations to select AIMs optimized for each of the three main continental populations that form the basis of modern Latin American populations. We selected markers on the basis of locus-specific branch length to be informative, well distributed throughout the genome, capable of being genotyped on widely available commercial platforms, and applicable throughout the Americas by minimizing within-continent heterogeneity. We then validated the panel in samples from four admixed populations by comparing ancestry estimates based on the AIMs panel to estimates based on genome-wide association study (GWAS) data. The panel provided balanced discriminatory power among the three ancestral populations and accurate estimates of individual ancestry proportions (R2 > 0.9 for ancestral components with significant between-subject variance). Finally, we genotyped samples from 18 populations from Latin America using the AIMs panel and estimated variability in ancestry within and between these populations. This panel and its reference genotype information will be useful resources to explore population history of admixture in Latin America and to correct for the potential effects of population stratification in admixed samples in the region.     

Synthesis and potent antitumor activity of new arylamino derivatives of nor-beta-lapachone and nor-alpha-lapachone

Several arylamino derivatives of nor-beta-lapachone were synthesized in moderate to high yields and found to show very potent cytotoxicity against six neoplastic cancer cells: SF-295 (central nervous system), HCT-8 (colon), MDAMB-435 (breast), HL-60 (leukaemia), PC-3 (prostate), and B-16 (murine melanoma), with IC(50) below 1 microg/mL. Their cytotoxicities were compared to doxorubicin and with their synthetic precursors, beta-lapachone and nor-beta-lapachone. The activity against a normal murine fibroblast L-929 showed that some of the compounds were selective against cancer cells. The absence of hemolytic activity (EC(50)>200 microg/mL), performed with erythrocyte suspensions, suggests that the cytotoxicity of the compounds was not related to membrane damage of mouse erythrocytes. For comparison purposes, one isomeric compound based on nor-alpha-lapachone was also synthesized and showed lower activity than the related ortho-derivative. The modified arylamino quinones appear as interesting new lead compounds in anti-cancer drug development.     

Exploitation of mitochondrial nad6 as a complementary marker for studying population variability in Lepidoptera

The applicability of mitochondrial nad6 sequences to studies of DNA and population variability in Lepidoptera was tested in four species of economically important moths and one of wild butterflies. The genetic information so obtained was compared to that of cox1 sequences for two species of Lepidoptera. nad6 primers appropriately amplified all the tested DNA targets, the generated data proving to be as informative and suitable in recovering population structures as that of cox1. The proposal is that, to obtain more robust results, this mitochondrial region can be complementarily used with other molecular sequences in studies of low level phylogeny and population genetics in Lepidoptera.     

Effect of fertilization pulses on the production of <Emphasis Type="Italic">Gracilaria birdiae</Emphasis> seedlings under laboratory and field conditions

The genus Gracilaria is one of the most important sources of agar in the world. In Brazil, Gracilaria birdiae is the main commercially exploited species; however, overexploitation has contributed to the depletion of natural beds. In order to obtain more information so as to consolidate G. birdiae cultivation, studies under laboratory (indoor and outdoor) and field (sea and shrimp pond) conditions were conducted to evaluate the effects of fertilizer pulses on biomass and relative growth rate (RGR) of this species. The following nutrient sources used were (T₁) shrimp-pond effluent, (T₂) fertilizer for aquarium plants (Mbreda), and (T₃) fertilizer extract of Ascophyllum nodosum (Acadian). Significant differences for growth were recorded over time for all treatments in both outdoor and field conditions (p < 0.001). The highest RGRs were recorded for treatments that used pulses of commercial fertilizers (T₂ and T₃) and the lowest for treatment using shrimp-pond effluent pulses (T₁). The analysis of the nutrient content in tissue also showed a relationship between growth and nitrogen and phosphorus accumulation in the algal tissues. The N/P ratio indicated a significant effect on the growth of G. birdiae and the highest RGRs were registered for seedlings with a N/P ratio ≥16 (T₂ and T₃). In conclusion, the best results were recorded for the Mbreda and Acadian commercial fertilizers. However, although no significant differences were detected between growth and the two fertilizers (T₂ and T₃), the seedlings cultivated under Acadian pulses showed a better performance against environmental stress caused by reduced salinity.     

Cryptosexuality and the genetic diversity paradox in coffee rust, Hemileia vastatrix

Background

Despite the fact that coffee rust was first investigated scientifically more than a century ago, and that the disease is one of the major constraints to coffee production - constantly changing the socio-economic and historical landscape of the crop - critical aspects of the life cycle of the pathogen, Hemileia vastatrix, remain unclear. The asexual urediniospores are regarded as the only functional propagule: theoretically, making H. vastatrix a clonal species. However, the well-documented emergence of new rust pathotypes and the breakdown in genetic resistance of coffee cultivars, present a paradox.

Methods and Results

Here, using computer-assisted DNA image cytometry, following a modified nuclear stoichiometric staining technique with Feulgen, we show that meiosis occurs within the urediniospores. Stages of spore development were categorised based on morphology, from the spore-mother cell through to the germinating spore, and the relative nuclear DNA content was quantified statistically at each stage.

Conclusions

Hidden sexual reproduction disguised within the asexual spore (cryptosexuality) could explain why new physiological races have arisen so often and so quickly in Hemileia vastatrix. This could have considerable implications for coffee breeding strategies and may be a common event in rust fungi, especially in related genera occupying the same basal phylogenetic lineages.     

Targeting molecular interactions essential for Plasmodium sexual reproduction

Malaria remains one of the most devastating infectious diseases, killing up to a million people every year. Whereas much progress has been made in understanding the life cycle of the parasite in the human host and in the mosquito vector, significant gaps of knowledge remain. Fertilization of malaria parasites, a process that takes place in the lumen of the mosquito midgut, is poorly understood and the molecular interactions (receptor–ligand) required for Plasmodium fertilization remain elusive. By use of a phage display library, we identified FG1 (Female Gamete peptide 1), a peptide that binds specifically to the surface of female Plasmodium berghei gametes. Importantly, FG1 but not a scrambled version of the peptide, strongly reduces P. berghei oocyst formation by interfering with fertilization. In addition, FG1 also inhibits P. falciparum oocyst formation suggesting that the peptide binds to a molecule on the surface of the female gamete whose structure is conserved. Identification of the molecular interactions disrupted by the FG1 peptide may lead to the development of novel malaria transmission‐blocking strategies.