Nataly Diniz de Lima Santos

Effect of Moringa oleifera lectin on development and mortality of Aedes aegypti larvae

Aedes aegypti larvae have developed tolerance to many insecticides used for mosquito control. Moringa oleifera seeds contain a water-soluble lectin (WSMoL) and this paper reports the effect of M. oleifera seed extracts (MoE(1-15)) and WSMoL on development and survival of A. aegypti larvae. WSMoL peptide from in-gel trypsin digestion is also described. MoE(1-15) showed hemagglutinating activity and WSMoL had similarity with flocculating proteins from M. oleifera seeds. MoE(1) and MoE(3) delayed larval development which stopped in the third instar (L3) in MoE(6) and MoE(15). Significant (p<0.0001) larval mortality was only detected in MoE(15). Native WSMoL showed larvicidal activity (LC(50) 0.197 mg mL(-1)) and heated lectin, without hemagglutinating activity, did not kill fourth instar (L4) larvae. Optical microscopy showed that live L4 from MoE(1) presented underlying epithelium, increased gut lumen and hypertrophic segments; dead L4 from WSMoL were absent of underlying epithelium, had increased gut lumen and hypertrophic segments. The presence of hemagglutinating activity in the extracts suggests that soluble lectin promotes the delay of larval development and mortality; furthermore, the absence of larvicidal activity in heat-denatured WSMoL strengthens the involvement of lectin in this activity mechanism.

Larvicidal activity of lectins from Myracrodruon urundeuva on Aedes aegypti

Aedes aegypti transmits etiologic agents of yellow fever and dengue. Vaccine for dengue virus is not available and vector control is essential to minimize dengue incidence. This report deals with the larvicidal activity of lectins isolated from Myracrodruon urundeuva bark (MuBL) and heartwood (MuHL). The lectins were isolated by ammonium sulphate treatment of crude extracts followed by chromatography on chitin. MuBL and MuHL were evaluated by electrophoresis under native (PAGE) and denaturing conditions (SDS-PAGE). Carbohydrate specificity of lectins was evaluated by hemagglutinating activity (HA) inhibition assay using N-acetyl-d-glucosamine and by affinity chromatography on N-acetyl-D-glucosamine immobilized in agarose gel. Larvicidal activity against A. aegypti was investigated with the extracts, salt fractions and isolated lectins. MuBL and MuHL were characterized by PAGE as basic proteins of molecular masses of 14.0 and 14.4 kDa, respectively. The interaction of lectins with N-acetylglucosamine was detected by inhibition of HA by monosaccharide and lectin adsorptions on N-acetyl-D-glucosamine matrix. All M. urundeuva preparations promoted larvae mortality. LC16, LC50 and LC84 values of 0.077, 0.125, 0.173 for MuBL and 0.03, 0.04 and 0.05 mg/mL for MuHL were obtained. To our knowledge this is the first report of larvicidal activity of lectins against A. aegypti.

Antibacterial and antifungal activities of Myracrodruon urundeuva heartwood

The aim of this work was to isolate a lectin from Myracrodruon urundeuva heartwood and to evaluate its antimicrobial activity against bacteria and fungi that attack plants, including woods. The lectin was isolated from heartwood through affinity chromatography on a chitin column monitored by hemagglutination assay. The lectin inhibited Gram-negative and Gram-positive bacteria and was more effective than antifungal Cercobin in growth inhibition of phytopathogenic fungi. The detected antimicrobial activity reveals the possible role of the lectin in the resistance of M. urundeuva heartwood against deteriorative biological agents. The M. urundeuva lectin is the first bioactive peptide found in heartwood, probably stored as a chemical protection against biodegradation.

Oviposition-Stimulant and Ovicidal Activities of Moringa oleifera Lectin on Aedes aegypti

Background Natural insecticides against the vector mosquito Aedes aegypti have been the object of research due to their high level of eco-safety. The water-soluble Moringa oleifera lectin (WSMoL) is a larvicidal agent against A. aegypti. This work reports the effects of WSMoL on oviposition and egg hatching of A. aegypti. Methodology/Principal Findings WSMoL crude preparations (seed extract and 0–60 protein fraction), at 0.1 mg/mL protein concentration, did not affect oviposition, while A. aegypti gravid females laid their eggs preferentially (73%) in vessels containing isolated WSMoL (0.1 mg/mL), compared with vessels containing only distilled water (control). Volatile compounds were not detected in WSMoL preparation. The hatchability of fresh eggs deposited in the solutions in the oviposition assay was evaluated. The numbers of hatched larvae in seed extract, 0–60 protein fraction and WSMoL were 45±8.7 %, 20±11 % and 55±7.5 %, respectively, significantly (p<0.05) lower than in controls containing only distilled water (75–95%). Embryos were visualized inside fresh control eggs, but not within eggs that were laid and maintained in WSMoL solution. Ovicidal activity was also assessed using stored A. aegypti eggs. The protein concentrations able to reduce the hatching rate by 50% (EC50) were 0.32, 0.16 and 0.1 mg/mL for seed extract, 0–60 protein fraction and WSMoL, respectively. The absence of hatching of stored eggs treated with WSMoL at 0.3 mg/mL (EC99) after transfer to medium without lectin indicates that embryos within the eggs were killed by WSMoL. The reduction in hatching rate of A. aegypti was not linked to decrease in bacteria population. Conclusions/Significance WSMoL acted both as a chemical stimulant cue for ovipositing females and ovicidal agent at a given concentration. The oviposition-stimulant and ovicidal activities, combined with the previously reported larvicidal activity, make WSMoL a very interesting candidate in integrated A. aegypti control.

Lectin from inflorescences of ornamental crop Alpinia purpurata acts on immune cells to promote Th1 and Th17 responses, nitric oxide release, and lymphocyte activation

Alpinia purpurata is an ornamental crop known as a source of bioactive molecules. This is the first study to report isolation of a lectin (carbohydrate-binding protein) from A. purpurata inflorescences (ApuL). The immunomodulatory potential of ApuL was evaluated by investigating its effects on the production of cytokines and release of nitric oxide by human peripheral blood mononuclear cells (PBMCs). In addition, the differentiation and activation of lymphocytes treated with ApuL was evaluated by immunophenotyping assays. ApuL is an acidic and oligomeric protein with native molecular mass of 34kDa. The hemagglutinating activity (HA) of ApuL was inhibited by the glycoproteins fetuin and ovalbumin, was resistant to heating at 100°C and stimulated in the presence of calcium and magnesium ions. ApuL showed highest HA at pH 7.5 but failed to agglutinate erythrocytes at pH 8.0 and 9.0. ApuL induced the release of cytokines belonging to Th1 (IFN-γ, TNF-α, and IL-6) and Th17 (IL-17A) profiles as well as of nitric oxide, stimulating a pro-inflammatory environment. Moreover, ApuL also stimulated the production of IL-10, an anti-inflammatory cytokine with regulatory role. Incubation with lectin resulted in differentiation and activation of both T CD8+ and CD4+ subsets of lymphocytes, as evident from the expression of the CD28 costimulatory molecule. In conclusion, A. purpurata inflorescence is a source of an immunomodulatory lectin with potential immunoregulatory application, thereby adding biotechnological value to this ornamental crop.

Insect midgut structures and molecules as targets of plant-derived protease inhibitors and lectins: Insect midgut structures and molecules as targets of plant-derived protease inhibitors and lectins

The midgut of insects is involved in digestion, osmoregulation and immunity. Although several defensive strategies are present in this organ, its organization and function may be disturbed by some insecticidal agents, including bioactive proteins like lectins and protease inhibitors (PIs) from plants. PIs interfere with digestion, leading to poor nutrient absorption and decreasing amino acid bioavailability. Intake of PIs can delay development, cause deformities and reduce fertility. Ingestion of PIs may lead to changes in the set of proteases secreted in the insect gut, but this response is often insufficient and results in aggravation of the malnutrition status. Lectins are proteins that are able to interact with glycoconjugates, including those linked to cell surfaces. Their effects on the midgut include disruption of the peritrophic matrix, brush border and secretory cell layer; induction of apoptosis and oxidative stress; interference with nutrient absorption and transport proteins; and damaging effects on symbionts. In addition, lectins can cross the intestinal barrier and reach the hemolymph. The establishment of resistant insect populations due to selective pressure resulting from massive use of a bioactive protein is an actual possibility, but this can be minimized by the multiple mode-of-action of these proteins, mainly the lectins.

Antimicrobial potential of Alpinia purpurata lectin (ApuL): Growth inhibitory action, synergistic effects in combination with antibiotics, and antibiofilm activity

The Alpinia purpurata inflorescence contains a lectin (ApuL), which has immunomodulatory activities on human cells. In the present work, it was evaluated the antibacterial and antifungal effects of ApuL against human pathogens. ApuL showed bacteriostatic activity against non-resistant (UFPEDA-02) and an oxacillin-resistant isolate (UFPEDA-672) of Staphylococcus aureus with minimal inhibitory concentrations (MIC50) of 50 and 400 μg/mL, respectively. In addition, it showed bactericidal effect on the non-resistant isolate (minimal bactericidal concentration: 200 μg/mL). For Candida albicans and Candida parapsilosis, ApuL showed fungistatic effect (MIC50: 200 and 400 μg/mL, respectively). The lectin was able to impair the viability of the microorganism cells, as indicated by propidium iodide (PI) staining. Analysis of growth curves, protein leakage, and ultrastructural changes supported that ApuL acts through distinct mechanisms on S. aureus isolates. Ultrastructural analysis of ApuL-treated Candida cells revealed malformations with elongations and bulges. ApuL-oxacillin combination showed synergistic effect on the oxacillin-resistant isolates UFPEDA-670 and 671, which were not sensitive to lectin alone. Synergism was also detected for ApuL-ceftazidime against a multidrug-resistant isolate of Pseudomonas aeruginosa. Synergistic action of ApuL-fluconazole was detected for C. parapsilosis, which was insensitive to the drug alone. Biofilm formation by S. aureus non-resistant isolate and C. albicans was remarkably inhibited by ApuL at sub-inhibitory concentrations. In conclusion, ApuL showed differential effects on non-resistant and resistant bacterial isolates, was active against Candida species, and showed synergistic action in combination with antibiotics.