Complementary Sex Determination in the Parasitic Wasp Diachasmimorpha longicaudata

We studied the sex determination in Diachasmimorpha longicaudata, a parasitoid braconid wasp widely used as biological control agent of fruit pest tephritid flies. We tested the complementary sex determination hypothesis (CSD) known in at least 60 species of Hymenoptera. According to CSD, male or female development depends on the allelic composition of one sex locus (single-locus CSD) or multiple sex loci (multiple-locus CSD). Hemizygote individuals are normal haploid males, and heterozygotes for at least one sex locus are normal diploid females, but homozygotes for all the sex loci are diploid males. In order to force the occurrence of diploid males in D. longicaudata, we established highly inbred lines and examined their offspring using chromosome counting, flow cytometry, and sex ratio analysis. We found that when mother-son crosses were studied, this wasp produced about 20% of diploid males out of the total male progeny. Our results suggest that this parasitoid may represent the second genus with multiple-locus CSD in Hymenoptera. Knowledge about the sex determination system in D. longicaudata is relevant for the improvement of mass rearing protocols of this species. This information also provides the necessary background for further investigations on the underlying molecular mechanisms of sex determination in this species, and a better insight into the evolution of this pathway in Hymenoptera in particular and insects in general.     

Selective Expansion of Viral Variants following Experimental Transmission of a Reconstituted Feline Immunodeficiency Virus Quasispecies

Following long-term infection with virus derived from the pathogenic GL8 molecular clone of feline immunodeficiency virus (FIV), a range of viral variants emerged with distinct modes of interaction with the viral receptors CD134 and CXCR4, and sensitivities to neutralizing antibodies. In order to assess whether this viral diversity would be maintained following subsequent transmission, a synthetic quasispecies was reconstituted comprising molecular clones bearing envs from six viral variants and its replicative capacity compared in vivo with a clonal preparation of the parent virus. Infection with either clonal (Group 1) or diverse (Group 2) challenge viruses, resulted in a reduction in CD4+ lymphocytes and an increase in CD8+ lymphocytes. Proviral loads were similar in both study groups, peaking by 10 weeks post-infection, a higher plateau (set-point) being achieved and maintained in study Group 1. Marked differences in the ability of individual viral variants to replicate were noted in Group 2; those most similar to GL8 achieved higher viral loads while variants such as the chimaeras bearing the B14 and B28 Envs grew less well. The defective replication of these variants was not due to suppression by the humoral immune response as virus neutralising antibodies were not elicited within the study period. Similarly, although potent cellular immune responses were detected against determinants in Env, no qualitative differences were revealed between animals infected with either the clonal or the diverse inocula. However, in vitro studies indicated that the reduced replicative capacity of variants B14 and B28 in vivo was associated with altered interactions between the viruses and the viral receptor and co-receptor. The data suggest that viral variants with GL8-like characteristics have an early, replicative advantage and should provide the focus for future vaccine development.     

EscO,a Functional and Structural Analog of the Flagellar FliJ Protein,Is a Positive Regulator of EscN ATPase Activity of the Enteropathogenic Escherichia coli Injectisome

Type III secretion systems (T3SSs) are multiprotein molecular devices used by many Gram-negative bacterial pathogens to translocate effector proteins into eukaryotic cells. A T3SS is also used for protein export in flagellar assembly, which promotes bacterial motility. The two systems are evolutionarily related, possessing highly conserved components in their export apparatuses. Enteropathogenic Escherichia coli (EPEC) employs a T3SS, encoded by genes in the locus of enterocyte effacement (LEE) pathogenicity island, to colonize the human intestine and cause diarrheal disease. In the present work, we investigated the role of the LEE-encoded EscO protein (previously Orf15 or EscA) in T3SS biogenesis. We show that EscO shares similar properties with the flagellar FliJ and the Yersinia YscO protein families. Our findings demonstrate that EscO is essential for secretion of all categories of T3SS substrates. Consistent with its central role in protein secretion, it was found to interact with the ATPase EscN and its negative regulator, EscL, of the export apparatus. Moreover, we show that EscO stimulates EscN enzymatic activity; however, it is unable to upregulate ATP hydrolysis in the presence of EscL. Remarkably, EscO partially restored the swimming defect of a Salmonella flagellar fliJ mutant and was able to stimulate the ATPase activity of FliI. Overall, our data indicate that EscO is the virulence counterpart of the flagellar FliJ protein.     

A new type of anti-ganglioside antibodies present in neurological patients

High titers of anti-GA1 antibodies have been associated with neurological syndromes. In most cases, these antibodies cross-react with the structurally related glycolipids GM1 and GD1b, although specific anti-GA1 antibodies have also been reported. The role of specific anti-GA1 antibodies is uncertain since the presence of GA1 in the human nervous system has not been clarified. A rabbit was immunized with GD1a and its sera were screened for antibody reactivity by standard immunoassay methods (HPTLC-immunostaining and ELISA). Anti-GD1a antibodies were not detected but, unexpectedly, anti-GA1 IgG-antibodies were found. Antibody binding to GA1 was inhibited by soluble GA1 but also by GD1a. These results indicate that the rabbit produced antibodies that recognize epitopes present on the glycolipids, that are absent or not exposed on solid phase adsorbed GD1a. We investigated the presence of these unusual anti-ganglioside antibodies in normal and neurological patient sera. Approximately, 10% of normal human sera contained low titer of specific anti-GA1 IgG-antibodies but none of them recognized soluble GD1a. High titers of IgG-antibodies reacting only with GA1 were detected in 12 patient sera out of 325 analyzed. Of these, 6 sera showed binding that was inhibited by soluble GD1a and four of them also by GM1. This new type of anti-ganglioside antibodies should be considered important elements for understanding of the pathogenesis of these diseases as well as their diagnosis.     

Helminth Antigens Enable CpG-Activated Dendritic Cells to Inhibit the Symptoms of Collagen-induced Arthritis through Foxp3+ Regulatory T Cells

Dendritic cells (DC) have the potential to control the outcome of autoimmunity by modulating the immune response. In this study, we tested the ability of Fasciola hepatica total extract (TE) to induce tolerogenic properties in CpG-ODN (CpG) maturated DC, to then evaluate the therapeutic potential of these cells to diminish the inflammatory response in collagen induced arthritis (CIA). DBA/1J mice were injected with TE plus CpG treated DC (T/C-DC) pulsed with bovine collagen II (CII) between two immunizations with CII and clinical scores CIA were determined. The levels of CII-specific IgG2 and IgG1 in sera, the histological analyses in the joints, the cytokine profile in the draining lymph node (DLN) cells and in the joints, and the number, and functionality of CD4+CD25+Foxp3+ T cells (Treg) were evaluated. Vaccination of mice with CII pulsed T/C-DC diminished the severity and incidence of CIA symptoms and the production of the inflammatory cytokine, while induced the production of anti-inflammatory cytokines. The therapeutic effect was mediated by Treg cells, since the adoptive transfer of CD4+CD25+ T cells, inhibited the inflammatory symptoms in CIA. The in vitro blockage of TGF-β in cultures of DLN cells plus CII pulsed T/C-DC inhibited the expansion of Treg cells. Vaccination with CII pulsed T/C-DC seems to be a very efficient approach to diminish exacerbated immune response in CIA, by inducing the development of Treg cells, and it is therefore an interesting candidate for a cell-based therapy for rheumatoid arthritis (RA).     

Differential regulation of p53 function by protein kinase C isoforms revealed by a yeast cell system

The complexity of the mammalian p53 pathway and protein kinase C (PKC) family has hampered the discrimination of the effect of PKC isoforms on p53 activity. Using yeasts co-expressing the human wild-type p53 and a mammalian PKC-α, -δ, -ε or -ζ, we showed a differential regulation of p53 activity and phosphorylation state by PKC isoforms. Whereas PKC-α reduced the p53-induced yeast growth inhibition and cell cycle arrest, PKC-δ and -ε enhanced the p53 activity through p53 phosphorylation, and PKC-ζ had no effect on p53. This work identified positive and negative p53 regulators which represent promising pharmacological targets in anti-cancer therapy.     

Nitrous oxide and methane emissions from soil and nitrogen uptake by eucalyptus fertilized with enhanced efficiency fertilizers

Plant and Soil - Evaluate which nitrogen fertilizer alone or in combination with additives for enhanced efficiency can reduce N2O and CH4 emissions of soil cultivated with eucalyptus and also...     

Effects of nitric oxide on sunflower seedlings: A balance between defense and development

Nitric oxide (NO) is a major plant signaling molecule that plays key roles during plant-pathogen interactions and plant development. Previous work showed the participation of NO in the development and lignin composition of sunflower roots. Thereby, we have hypothesized that NO applications could control the attack of the fungal pathogen Verticillium dahliae in sunflowers. Seedlings growing hydroponically were pretreated with NO donors and further inoculated with the fungus. Evaluation of disease symptoms showed that NO pretreatments could not reduce Verticillium wilt. Strikingly, NO donors appear to promote the fungal infection. These results indicate that NO applications were unable to protect sunflowers from Verticillium attack and highlight the role played by the fine tuning regulation of NO levels required to balance plant responses between development and defense.     

Correlating diet and digestive tract specialization: examples from the lizard family Liolaemidae

A range of digestive tract specializations were compared among dietary categories in the family Liolaemidae to test the hypothesis that herbivores require greater gut complexity to process plant matter. Additionally, the hypothesis that herbivory favors the evolution of larger body size was tested. Lastly, the association between diet and hindgut nematodes was explored. Herbivorous liolaemids were larger relative to omnivorous and insectivorous congeners and consequently had larger guts. In addition, small intestine length of herbivorous liolaemids was disproportionately longer than that of congeners. Significant interaction effects between diet and body size among organ dimensions indicate that increases in organ size occur to a greater extent in herbivores than other diet categories. For species with plant matter in their guts, there was a significant positive correlation between the percentage of plant matter consumed and small intestine length. Herbivorous liolaemids examined in this study lacked the gross morphological specializations (cecum and colonic valves) found in herbivores in the families Iguanidae and Agamidae. A significantly greater percentage of herbivorous species had nematodes in their gut. Of the species with nematodes, over 95% of herbivores had nematodes only in the hindgut. Prevalence of nematodes in the hindgut of herbivores was 2 x that of omnivores and 4 x that of insectivores.     

Fungi isolated from insects in strawberry crops act as potential biological control agents of <Emphasis Type="Italic">Duponchelia fovealis</Emphasis> (Lepidoptera: Crambidae)

Entomopathogenic fungi were collected from insects, belonging to orders Lepidoptera, Coleoptera, Hemiptera, Hymenoptera, and Diptera, with signs of infection. Single-spore colonies were identified based on morphological traits and ribosomal ITS sequences. The most commonly found fungi were Fusarium oxysporum Schlechtendal and Beauveria bassiana (Balsamo-Crivelli) Vuillemin. Isolation of Trichoderma atroviride Bisset, Aspergillus flavipes (Bainier and Sartory) Thom and Church, Aspergillus iizukae Sugiyama, Penicillium mallochii Rivera, Urb and Seifert, Penicillium adametzioides S. Abe ex G. Smith, and Mucor nidicola Madden, Stchigel, Guarro and Starks associated to insects is reported for the first time. Tests with B. bassiana isolates against Duponchelia fovealis Zeller (Lepidoptera: Crambidae), showed high larval mortality in vitro and in greenhouse, demonstrating its potential as biological control agent. Understanding the fungal microbiota from insects can provide promising isolates for use in integrated pest management programs. This is the first report characterizing the mycobiota in insects collected in strawberry crops, and evaluating the pathogenicity of B. bassiana against D. fovealis, a pest that causes severe losses to farmers.