Do mosquitoes filter the access of Plasmodium cytochrome b lineages to an avian host?

Many parasites show fidelity to a set of hosts in ecological time but not evolutionary time and the determinants of this pattern are poorly understood. Malarial parasites use vertebrate hosts for the asexual stage of their life cycle but use Dipteran hosts for the sexual stage. Despite the potential evolutionary importance of Dipteran hosts, little is known of their role in determining a parasite's access to vertebrate hosts. Here, we use an avian malarial system in Panama to explore whether mosquitoes act as an access filter that limits the range of vertebrate hosts used by particular parasite lineages. We amplified and sequenced Plasmodium mitochondrial DNA (mtDNA) from Turdus grayi (clay-coloured robin) and from mosquitoes at the same study site. We trapped and identified to species 123 141 female mosquitoes and completed polymerase chain reaction (PCR) screening for Plasmodium parasites in 435 pools of 20 mosquitoes per pool (8700 individuals total) spanning the 11 most common mosquito species. Our primers amplified nine Plasmodium lineages, whose sequences differed by 1.72%–10.0%. Phylogenetic analyses revealed partial clustering of lineages that co-occurred in mosquito hosts. However PAN3 and PAN6, the two primary parasite lineages of T. grayi , exhibited sequence divergence of 8.59% and did not cluster in the phylogeny. We detected these two lineages exclusively in mosquitoes from different genera — PAN3 was found only in Culex (Melanoconion) ocossa , and PAN6 was found only in Aedeomyia squamipennis . Furthermore, each of these two parasite lineages co-occurred in mosquitoes with other Plasmodium lineages that were not found in the vertebrate host T. grayi . Together, this evidence suggests that parasite–mosquito associations do not restrict the access of parasites to birds but instead may actually facilitate the switching of vertebrate hosts that occurs over evolutionary time.     

Control in vitro tick (Boophilus microplus; Acari: Ixodidae) through plant extracts

Control in vitro tick (Boophilus microplus; Acari: Ixodidae) through plant extracts. Hydro-alcohol extracts of ten tropical plants were tested by the immersion method on mature ticks (Boophilus microplus Canestrini 1887). The biological evaluation was made with raw hydro alcoholic extracts (R-OH) to determine the most promissory compounds, which later were fractioned in their polar and non-polar parts. The polar parts showed a significantly inferior effect. The species that produced the highest mortality were Zizigium aromaticum, Morus alba, Piper nigrum and the mixture of Allium sativum- Z. aromaticum (all non-polar); on oviposition: M. alba, P. nigrum and the mixture of A. sativum - Z. aromaticum (all non-polar); on eclosion: P. nigrum, Z. aromaticum, Echinacea angustiofilia, Poligonum punctatum, M. alba and the mixture of A. sativum-Z. aromaticum (all non-polar): on control percentage: P. nigrum, E. angustifolia, P. punctatum and M. alba (all non-polar, except for M. alba in which both fractions showed important activity); and finally, on the percentage of oviposition inhibition: P. nigrum, E. angustifolia, P. punctatum, Gliricidia sepium and M. alba.     

Oleic acid induces intracellular calcium mobilization, MAPK phosphorylation, superoxide production and granule release in bovine neutrophils

Oleic acid (OA) is a nonesterified fatty acid that is released into the blood during lipomobilization at the time of calving in cows, a period where increased risk of infection and acute inflammation is observed. These data suggest potential OA-mediated regulation of innate immune responses. In the present study, we assessed the effects of OA on intracellular calcium release, ERK1/2 phosphorylation, superoxide production, CD11b expression and matrix metalloproteinase-9 (MMP-9) release in bovine neutrophils. Furthermore, the presence of GPR40, an OA receptor, was assessed by RT-PCR, immunoblotting and confocal microscopy. OA induced, in a dose-dependent manner, intracellular calcium mobilization, superoxide production and CD11b expression in bovine neutrophils; these effects were reduced by the intracellular chelating agent BAPTA-AM. OA also induced ERK2 phosphorylation and MMP-9 release. RT-PCR analysis detected mRNA expression of a bovine ortholog of the GPR40 receptor. Using a polyclonal antibody against human GPR40, we detected a protein of 31 kDa by immunoblotting that was localized predominately in the plasma membrane. The selective agonist of GPR40, GW9508, induced intracellular calcium mobilization and ERK2 phosphorylation. In conclusion, OA can modulate bovine neutrophil responses in an intracellular calcium-dependent manner; furthermore, these responses could be induced by GPR40 activation.     

Confirmation of Anopheles (Anopheles) calderoni Wilkerson, 1991 (Diptera: Culicidae) in Colombia and Ecuador through molecular and morphological correlation with topotypic material

The morphologically similar taxa Anopheles calderoni, Anopheles punctimacula, Anopheles malefactor and Anopheles guarao are commonly misidentified. Isofamilies collected in Valle de Cauca, Colombia, showed morphological characters most similar to An. calderoni, a species which has never previously been reported in Colombia. Although discontinuity of the postsubcostal pale spots on the costa (C) and first radial (R1) wing veins is purportedly diagnostic for An. calderoni, the degree of overlap of the distal postsubcostal spot on C and R1 were variable in Colombian specimens (0.003-0.024). In addition, in 98.2% of larvae, seta 1-X was located off the saddle and seta 3-C had 4-7 branches in 86.7% of specimens examined. Correlation of DNA sequences of the second internal transcribed spacer and mtDNA cytochrome c oxidase subunit I gene (COI) barcodes (658 bp of the COI gene) generated from Colombian progeny material and wild-caught mosquitoes from Ecuador with those from the Peruvian type series of An. calderoni confirmed new country records. DNA barcodes generated for the closely related taxa, An. malefactor and An. punctimacula are also presented for the first time. Examination of museum specimens at the University of the Valle, Colombia, revealed the presence of An. calderoni in inland localities across Colombia and at elevations up to 1113 m.     

Risk markers in subjects with high loads of Ascaris lumbricoides in a rural community of the Cojedes State , Venezuela

A total of 337 individuals from a rural community of Venezuela (Mapurite, Estado Cojedes) was studied. Stoll faecal examination was carried out and the total number of A. lumbricoides was recovered after the treatment of the individuals sampled with Pyrantel pamoate at a dose of 10 mg/kg body weight in children and 15 mg/kg in adults. A relationship between the high worm burden condition (individuals excreting more than 10,000 epg of A. lumbricoides) and the age of the hosts was found. Resulting with the major values of relative risk index (Rr) the 5-9 years old class (Rr = 3.2 t = 3.4). This relationship was not found between high worm burden condition (HWB) and the sex of the hosts. The following individuals have the the highest possibilities to become HWB: with blood group A (Rr = 3.05 t = 4.08), with haemoglobin type HbA HbS (Rr = 1.86 t = 2.13), individuals with the combinations A HbA HbF (Rr = 3.46 t = 2.31) and A HbA HbS (Rr = 3.11 t = 2.78). It was estimated that the selective treatment of the total HWB detected (72) with an effective product, determined a reduction of 95.4% of the environmental contamination with de eggs and 72.3% of the worm burden.     

Gold coated ferric oxide nanoparticles based disposable magnetic genosensors for the detection of DNA hybridization processes

In this article, a disposable magnetic DNA sensor using an enzymatic amplification strategy for the detection of specific hybridization processes, based on the coupling of streptavidin-peroxidase to biotinylated target sequences, has been developed. A thiolated 19-mer capture probe was attached to gold coated ferric oxide nanoparticles and hybridization with the biotinylated target was allowed to proceed. Then, a streptavidin-peroxide was attached to the biotinylated target and the resulting modified gold coated ferric oxide nanoparticles were captured by a magnetic field on the surface of a home-made carbon screen printed electrode (SPE). Using hydroquinone as a mediator, a square wave voltammetric procedure was chosen to detect the hybridization process after the addition of hydrogen peroxide. Different aspects concerning the assay protocol and nanoparticles fabrication were optimized in order to improve the sensitivity of the developed methodology. A low detection limit (31 pM) with good stability (RSD=7.04%, n=10) was obtained without the need of polymerase chain reaction (PCR) amplification.     

Effect of temperature, pH, and metals on the stability and activity of phenylalanine hydroxylase from Chromobacterium violaceum

Phenylalanine hydroxylase (PAH) is a non-heme iron dioxygenase catalyzing the conversion of phenylalanine to tyrosine and is present in both prokaryotic and eukaryotic organisms. A relatively simple PAH is expressed by Chromobacterium violaceum, a gram-negative bacterium found in tropical and subtropical regions. The effects of temperature, pH and metals on the stability and catalytic activity of Chromobacterium violaceum PAH were determined by steady-state kinetics, circular dichroism (CD) and differential scanning calorimetry (DSC). The kcat and KM for phenylalanine were determined between 7 and 40 degrees C. The KM remained constant between 20 and 40 degrees C but rapidly increased below 20 degrees C. The half-life of the enzyme at 47 degrees C is 66+/-4 min in the presence of Fe(II) and 8+/-1 min in the presence of EDTA. The melting temperature of the protein determined by CD and DSC is 53+/-2 degrees C in the presence of EDTA and 63+/-2 degrees C in the presence of Fe(II). Co(II) stabilizes the enzyme (Tm=63+/-2 degrees C) and inhibits the catalytic activity by displacing iron from the active site. The optimum pH for catalytic activity and stability is 7.4. In conclusion, PAH is adapted for optimal phenylalanine binding at temperatures above 20 degrees C and Fe(II) enhances the resistance of the enzyme to thermal denaturation.     

Late Pleistocene environmental changes lead to unstable demography and population divergence of Anopheles albimanus in the northern Neotropics

We investigated the historical demography of Anopheles albimanus using mosquitoes from five countries and three different DNA regions, the mitochondrial cytochrome oxidase subunit I gene (COI), the single copy nuclear white gene and the ribosomal internal transcribed spacer two (ITS2). All the molecular markers supported the taxonomic status of a single species of An. albimanus. Furthermore, agreement between the COI and the white genes suggested a scenario of Pleistocene geographic fragmentation (i.e., population contraction) and subsequent range expansion across southern Central America.