Detection of Leishmania (Leishmania) infantum RNA in fleas and ticks collected from naturally infected dogs

The occurrence of the insect vector (sand flies) with low rates of Leishmania infection, as well as autochthonous transmission in the absence of the natural vector in dogs, have been reported. These unexpected data suggest a hypothesis of other arthropods as a possible way of Leishmania transmission. The prevalence of Leishmania (Leishmania) infantum in fleas and ticks collected from dogs with canine visceral leishmaniasis (CVL), as well as parasite viability, were evaluated herein. The presence of L. (L.) infantum was assayed by PCR and ELISA in ectoparasites and biological samples from 73 dogs living in a Brazilian endemic area. As the occurrence of Leishmania DNA in ticks and fleas is expected given their blood-feeding habits, we next investigated whether parasites can remain viable inside ticks. PCR and ELISA confirmed that 83% of the dogs had CVL. Fleas and ticks (nymphs, male and female adults) were collected in 55% and 63% of the 73 dogs, respectively. Out of the 60 dogs with CVL, 80% harbored ectoparasites infected with L. (L.) infantum. The infection rates of the ectoparasites were 23% and 50% for fleas and ticks, respectively. The RNA analysis of the extract from ticks left in laboratory conditions during 7 to 10 days after removal from CVL dogs showed that parasites were alive. In addition, live parasites were also detected inside adult ticks recently molted in laboratory conditions. These findings indicate a higher infection rate of L. (L.) infantum in ticks and fleas, but they do not conclusively demonstrate whether these ticks can act as vectors of CVL, despite the fact that their rates were higher than those previously described in Lutzomyia longipalpis. The presence of viable L. (L.) infantum in ticks suggests the possible importance of dog ectoparasites in CVL dissemination.

Biochemical characterization of serine transport in Leishmania (Leishmania) amazonensis

In addition to its role as a protein component in Leishmania, serine is also a precursor for the synthesis of both phosphatidylserine, which is a membrane molecule involved in parasite invasion and inactivation of macrophages, and sphingolipids, which are necessary for Leishmania to differentiate into its infective forms. We have characterized serine uptake in both promastigote and amastigote forms of Leishmania (Leishmania) amazonensis. In promastigotes, kinetic data show a single, saturable transport system, with a Km of 0.253 +/- 0.01 mM and a maximum velocity of 0.246 +/- 0.04 nmol/min per 107 cells. Serine transport increased linearly with temperature in the range from 20 degrees C to 45 degrees C, allowing the calculation of an activation energy of 7.09 kJ/mol. Alanine, cysteine, glycine, threonine, valine and ethanolamine competed with the substrate at a ten-fold excess concentration. Serine uptake was dependent on pH, with an optimum activity at pH 7.5. The characterization of the serine transport process in amastigotes revealed a transport system with a similar Km, energy of activation and pH response to that found in promastigotes, suggesting that the same transport system is active in both insect vector and mammalian host Leishmania stages. This could constitute an evolutionary mechanism that guarantees the provision of such an essential molecule during host change events, such as differentiation into amastigotes and macrophage invasion, as well as to ensure that the parasite maintains the infection in the mammalian host. (C) 2008 Elsevier B.V. All rights reserved.

Effects of the antimicrobial peptide gomesin on the global gene expression profile, virulence and biofilm formation of Xylella fastidiosa

In the xylem vessels of susceptible hosts, such as citrus trees, Xylella fastidiosa forms biofilm-like colonies that can block water transport, which appears to correlate to disease symptoms. Besides aiding host colonization, bacterial biofilms play an important role in resistance against antimicrobial agents, for instance antimicrobial peptides (AMPs). Here, we show that gomesin, a potent AMP from a tarantula spider, modulates X. fastidiosa gene expression profile upon 60 min of treatment with a sublethal concentration. DNA microarray hybridizations revealed that among the upregulated coding sequences, some are related to biofilm production. In addition, we show that the biofilm formed by gomesin-treated bacteria is thicker than that formed by nontreated cells or cells exposed to streptomycin. We have also observed that the treatment of X. fastidiosa with a sublethal concentration of gomesin before inoculation in tobacco plants correlates with a reduction in foliar symptoms, an effect possibly due to the trapping of bacterial cells to fewer xylem vessels, given the enhancement in biofilm production. These results warrant further investigation of how X. fastidiosa would respond to the AMPs produced by citrus endophytes and by the insect vector, leading to a better understanding of the mechanism of action of these molecules on bacterial virulence.

First evidence of insect attraction by a Southern Hemisphere Splachnaceae: The case of Tayloria dubyi Broth. in the Reserve Biosphere Cape Horn, Chile

The moss Tayloria dubyi (Splachnaceae) is endemic to the subantarctic Magallanes ecoregion where it grows exclusively on bird dung and perhaps only on feces of the goose Chloephaga picta, a unique habitat among Splachnaceae. Some species of Splachnaceae from the Northern Hemisphere are known to recruit coprophilous flies as a vector to disperse their spores by releasing intense odors mimicking fresh clung or decaying corpses. The flies land on the capsule, and may get in contact with the protruding mass of spores that stick to the insect body. The dispersal strategy relies on the spores falling off when the insect reaches fresh droppings or carrion. Germination is thought to be rapid and a new population is quickly established over the entire substrate. The objectives of this investigation were to determine whether the coprophilous T. dubyi attracts flies and to assess the taxonomic diversity of the flies visiting this moss. For this, fly traps were set up above mature sporophyte bearing populations in two peatlands on Navarino Island. We captured 64 flies belonging to the Muscidae (Palpibracus chilensis), Tachinidae (Dasyuromyia sp) and Sarcophagidae (not identified to species) above sporophytes of T. dubyi, whereas no flies were captured in control traps set up above Sphagnum mats nearby.

Molecular analysis of a mutant Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) shows an interruption of an inhibitor of apoptosis gene (iap-3) by a new class-II piggyBac-related insect transposon

A new piggyBac-related transposable element (TE) was found in the genome of a mutant Anticarsia gemmatalis multiple nucleopolyhedrovirus interrupting an inhibitor of apoptosis gene. This mutant virus induces apoptosis upon infection of an Anticarsia gemmatalis cell line, but not in a Trichoplusia ni cell line. The sequence of the new TE (which was named IDT for iap disruptor transposon) has 2531 bp with two DNA sequences flanking a putative Transposase (Tpase) ORF of 1719 bp coding for a protein with 572 amino acids. These structural features are similar to the piggyBac TE, also reported for the first time in the genome of a baculovirus. We have also isolated variants of this new TE from different lepidopteran insect cells and compared their Tpase sequences.

Spatial expansion and population structure of the neotropical malaria vector, Anopheles darlingi (Diptera: Culicidae)

Extensive population structuring is known to occur in Anopheles darlingi, the primary malaria vector of the Neotropics. We analysed the phylogeographic structure of the species using the mitochondrial cytochrome oxidase I marker. Diversity is divided into six main population groups in South America: Colombia, central Amazonia, southern Brazil, south-eastern Brazil, and two groups in north-east Brazil. The ancestral distribution of the taxon is hypothesized to be central Amazonia, and there is evidence of expansion from this region during the late Pleistocene. The expansion was not a homogeneous front, however, with at least four subgroups being formed due to geographic barriers. As the species spread, populations became isolated from each other by the Amazon River and the coastal mountain ranges of south-eastern Brazil and the Andes. Analyses incorporating distances around these barriers suggest that the entire South American range of An. darlingi is at mutation-dispersal-drift equilibrium. Because the species is distributed throughout such a broad area, the limited dispersal across some landscape types promotes differentiation between otherwise proximate populations. Moreover, samples from the An. darlingi holotype location in Rio de Janeiro State are substantially derived from all other populations, implying that there may be additional genetic differences of epidemiological relevance. The results obtained contribute to our understanding of gene flow in this species and allow the formulation of human mosquito health protocols in light of the potential population differences in vector capacity or tolerance to control strategies. (C) 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 97, 854-866.

Phytophagous insect fauna tracks host plant responses to exotic grass invasion

The high dependence of herbivorous insects on their host plants implies that plant invaders can affect these insects directly, by not providing a suitable habitat, or indirectly, by altering host plant availability. In this study, we sampled Asteraceae flower heads in cerrado remnants with varying levels of exotic grass invasion to evaluate whether invasive grasses have a direct effect on herbivore richness independent of the current disturbance level and host plant richness. By classifying herbivores according to the degree of host plant specialization, we also investigated whether invasive grasses reduce the uniqueness of the herbivorous assemblages. Herbivorous insect richness showed a unimodal relationship with invasive grass cover that was significantly explained only by way of the variation in host plant richness. The same result was found for polyphagous and oligophagous insects, but monophages showed a significant negative response to the intensity of the grass invasion that was independent of host plant richness. Our findings lend support to the hypothesis that the aggregate effect of invasive plants on herbivores tends to mirror the effects of invasive plants on host plants. In addition, exotic plants affect specialist insects differently from generalist insects; thus exotic plants affect not only the size but also the structural profile of herbivorous insect assemblages.

Alboserpin, a Factor Xa Inhibitor from the Mosquito Vector of Yellow Fever, Binds Heparin and Membrane Phospholipids and Exhibits Antithrombotic Activity

The molecular mechanism of factor Xa (FXa) inhibition by Alboserpin, the major salivary gland anticoagulant from the mosquito and yellow fever vector Aedes albopictus, has been characterized. cDNA of Alboserpin predicts a 45-kDa protein that belongs to the serpin family of protease inhibitors. Recombinant Alboserpin displays stoichiometric, competitive, reversible and tight binding to FXa (picomolar range). Binding is highly specific and is not detectable for FX, catalytic site-blocked FXa, thrombin, and 12 other enzymes. Alboserpin displays high affinity binding to heparin (K(D) similar to 20 nM), but no change in FXa inhibition was observed in the presence of the cofactor, implying that bridging mechanisms did not take place. Notably, Alboserpin was also found to interact with phosphatidylcholine and phosphatidylethanolamine but not with phosphatidylserine. Further, annexin V (in the absence of Ca(2+)) or heparin outcompetes Alboserpin for binding to phospholipid vesicles, suggesting a common binding site. Consistent with its activity, Alboserpin blocks prothrombinase activity and increases both prothrombin time and activated partial thromboplastin time in vitro or ex vivo. Furthermore, Alboserpin prevents thrombus formation provoked by ferric chloride injury of the carotid artery and increases bleeding in a dose-dependent manner. Alboserpin emerges as an atypical serpin that targets FXa and displays unique phospholipid specificity. It conceivably uses heparin and phosphatidylcholine/phosphatidylethanolamine as anchors to increase protein localization and effective concentration at sites of injury, cell activation, or inflammation.

Transcriptome and gene expression profile of ovarian follicle tissue of the triatomine bug Rhodnius prolixus

Insect oocytes grow in close association with the ovarian follicular epithelium (OFE), which escorts the oocyte during oogenesis and is responsible for synthesis and secretion of the eggshell. We describe a transcriptome of OFE of the triatomine bug Rhodnius prolixus, a vector of Chagas disease, to increase our knowledge of the role of FE in egg development. Random clones were sequenced from a cDNA library of different stages of follicle development. The transcriptome showed high commitment to transcription, protein synthesis, and secretion. The most abundant cDNA was a secreted (S) small, proline-rich protein with maximal expression in the vitellogenic follicle, suggesting a role in oocyte maturation. We also found Rp45, a chorion protein already described, and a putative chitin-associated cuticle protein that was an eggshell component candidate. Six transcripts coding for proteins related to the unfolded-protein response (UPR) by were chosen and their expression analyzed. Surprisingly, transcripts related to UPR showed higher expression during early stages of development and downregulation during late stages, when transcripts coding for S proteins participating in chorion formation were highly expressed. Several transcripts with potential roles in oogenesis and embryo development are also discussed. We propose that intense protein synthesis at the FE results in reticulum stress (RS) and that lowering expression of a set of genes related to cell survival should lead to degeneration of follicular cells at oocyte maturation. This paradoxical suppression of UPR suggests that ovarian follicles may represent an interesting model for studying control of RS and cell survival in professional S cell types. (C) 2011 Elsevier Ltd. All rights reserved.

Development of an adenoviral vector with robust expression driven by p53

Here we introduce a new adenoviral vector where transgene expression is driven by p53. We first developed a synthetic promoter, referred to as PGTx beta containing a p53-responsive element, a minimal promoter and the first intron of the rabbit P-globin gene. Initial assays using plasmid-based vectors indicated that expression was tightly controlled by p53 and was 5-fold stronger than the constitutive CMV immediate early promoter/enhancer. The adenoviral vector, AdPG, was also shown to offer p53-responsive expression in prostate carcinoma cells LNCaP (wt p53), DU-145 (temperature sensitive mutant of p53) and PC3 (p53-null, but engineered to express temperature-sensitive p53 mutants). AdPG served as a sensor of p53 activity in LNCaP cells treated with chemotherapeutic agents. Since p53 can be induced by radiotherapy and chemotherapy, this new vector could be further developed for use in combination with conventional therapies to bring about cooperation between the genetic and pharmacologic treatment modalities. (c) 2007 Elsevier Inc. All rights reserved.

C(k)-solvability near the characteristic set for a class of planar complex vector fields of infinite type

This paper deals with semi-global C(k)-solvability of complex vector fields of the form L = partial derivative/partial derivative t + x(r) (a(x) + ib(x))partial derivative/partial derivative x, r >= 1, defined on Omega(epsilon) = (-epsilon, epsilon) x S(1), epsilon > 0, where a and b are C(infinity) real-valued functions in (-epsilon, epsilon). It is shown that the interplay between the order of vanishing of the functions a and b at x = 0 influences the C(k)-solvability at Sigma = {0} x S(1). When r = 1, it is permitted that the functions a and b of L depend on the x and t variables, that is, L = partial derivative/partial derivative t + x(a(x, t) + ib(x, t))partial derivative/partial derivative x, where (x, t) is an element of Omega(epsilon).

THE POINCARE-BENDIXSON THEOREM ON THE KLEIN BOTTLE FOR CONTINUOUS VECTOR FIELDS

We present a version of the Poincare-Bendixson Theorem on the Klein bottle K(2) for continuous vector fields. As a consequence, we obtain the fact that K(2) does not admit continuous vector fields having a omega-recurrent injective trajectory.

Phase portraits of quadratic polynomial vector fields having a rational first integral of degree 3

In this paper, we classify all the global phase portraits of the quadratic polynomial vector fields having a rational first integral of degree 3. (C) 2008 Elsevier Ltd. All rights reserved.

Invariant theory and reversible-equivariant vector fields

In this paper we present results for the systematic study of reversible-equivariant vector fields - namely, in the simultaneous presence of symmetries and reversing symmetries - by employing algebraic techniques from invariant theory for compact Lie groups. The Hilbert-Poincare series and their associated Molien formulae are introduced,and we prove the character formulae for the computation of dimensions of spaces of homogeneous anti-invariant polynomial functions and reversible-equivariant polynomial mappings. A symbolic algorithm is obtained for the computation of generators for the module of reversible-equivariant polynomial mappings over the ring of invariant polynomials. We show that this computation can be obtained directly from a well-known situation, namely from the generators of the ring of invariants and the module of the equivariants. (C) 2008 Elsevier B.V, All rights reserved.

Nonexistence of global solutions for a class of complex vector fields on two-torus

The goal of this paper is study the global solvability of a class of complex vector fields of the special form L = partial derivative/partial derivative t + (a + ib)(x)partial derivative/partial derivative x, a, b epsilon C(infinity) (S(1) ; R), defined on two-torus T(2) congruent to R(2)/2 pi Z(2). The kernel of transpose operator L is described and the solvability near the characteristic set is also studied. (c) 2008 Elsevier Inc. All rights reserved.