Roberta Pereira Miranda Fernandes

Leishmanicidal activity of carvacrol-rich essential oil from Lippia sidoides Cham

Leishamaniasis is a disease that affects more than 2 million people worldwide, whose causative agent is Leishmania spp. The current therapy for leishmaniasis is far from satisfactory. All available drugs, including pentavalent antimony, require parenteral administration and are potentially toxic. Moreover, an increase in clinical resistance to these drugs has been reported. In this scenario, plant essential oils used traditionally in folk medicine are emerging as alternative sources for chemotherapeutic compounds. In this study, in vitro leishmanicidal effects of a thymol- and a carvacrol-rich essential oil from leaves of Lippia sidoides Cham. were investigated. The essential oils were extracted and their constituents were characterized by gas chromatography coupled to mass spectrometry (GC/MS). Both essential oils showed significant activity against promastigote forms of Leishmania chagasi. However, we found that carvacrol-rich essential oil was more effective, with IC50/72 h of 54.8 μg/mL compared to 74.1 μg/mL for thymol-rich oil. Carvacrol also showed lower IC50 than thymol. Our data suggest that L. sidoides essential oils are indeed promising sources of leishmanicidal compounds.

Identification of Suitable Reference Genes for Real Time Quantitative Polymerase Chain Reaction Assays on Pectoralis major Muscle in Chicken ( Gallus gallus )

Thirteen reference genes were investigated to determine their stability to be used as a housekeeping in gene expression studies in skeletal muscle of chickens. Five different algorithms were used for ranking of reference genes and results suggested that individual rankings of the genes differed among them. The stability of the expression of reference genes were validated using samples obtained from the Pectoralis major muscle in chicken. Samples were obtained from chickens in different development periods post hatch and under different nutritional diets. For gene expression calculation the ΔΔCt approach was applied to compare relative expression of pairs of genes within each of 52 samples when normalized to mitochondrially encoded cytochrome c oxidase II (MT-CO2) target gene. Our findings showed that hydroxymethylbilane synthase (HMBS) and hypoxanthine phosphoribosyl transferase 1 (HPRT1) are the most stable reference genes while transferrin receptor (TFRC) and beta-2-microglobulin (B2M) ranked as the least stable genes in the Pectoralis major muscle of chickens. Moreover, our results revealed that HMBS and HPRT1 gene expression did not change due to dietary variations and thus it is recommended for accurate normalization of RT-qPCR data in chicken Pectoralis major muscle.

Seleção de rizóbios nativos para guandu, caupi e feijão-de-porco nos tabuleiros costeiros de Sergipe

Abstract€–€The inoculation of rhizobial strains in jackbean ( Canavalia ensiformis ), cowpea ( Vignaunguiculata ) and pigeonpea ( Cajanus cajan ), recommended for other regions of Brazil, has not resultedin increases of biological nitrogen fixation rates and plant growth in soils of the coastal tableland ofSergipe (Brazilian Northeast). The objectives of this work were to evaluate the symbiotic effectivenessof indigenous rhizobia from that coastal tableland associated to these three legumes and their toleranceto stresses. Seventeen rhizobia strains were isolated and evaluated in a greenhouse. Four strains wereselected for pigeonpea; seven for cowpea and three for jackbean. Nodules number and dry weight wererelated to shoot dry weight, leaf area and leaf N€content, in all three legumes. The same strains wereefficient for both cowpea and pigeonpea. Three pigeonpea (R35, R43 and R45) and two cowpea (R10and R17) rhizobia were characterized in€vitro and all showed tolerance to high levels of nalidix acid,chloramphenicol and tetracycline; however, they were sensitive to streptomycin and kanamycin. Allstrains were able to grow at€35oC and, except for R17, were tolerant to aluminium (10€mg€L

Fatty acid profiles in Leishmania spp. isolates with natural resistance to nitric oxide and trivalent antimony

Fatty acids, especially those from phospholipids (PLFA), are essential membrane components that are present in relatively constant proportions in biological membranes under natural conditions. However, under harmful growth conditions, such as diseases, environmental changes, and chemical exposure, the fatty acid proportions might vary. If such changes could be identified and revealed to be specific for adverse situations, they could be used as biomarkers. Such biomarkers could facilitate the identification of virulence and resistance mechanisms to particular chemotherapeutic agents. Therefore, specific biomarkers could lead to better therapeutic decisions that would, in turn, enhance treatment effectiveness. The objective of this study was to compare the fatty acid profiles of trivalent antimony and nitric oxide (NO)-resistant and -sensitive Leishmania chagasi and Leishmania amazonensis isolates. Fatty acid methyl esters (FAMEs) were obtained from total lipids (MIDI), ester-linked lipids (ELFA), and ester-linked phospholipids (PLFA). FAMEs were analyzed by chromatography and mass spectrometry. Species- or resistance-associated differences in FAME profiles were assessed by nonmetric multidimensional scaling, multiresponse permutation procedures, and indicator species analyses. The isolate groups had different MIDI-FAME profiles. However, neither the ELFA nor PLFA profiles differed between the sensitive and resistant isolates. Levels of the fatty acid 18:1 Δ9c were increased in sensitive isolates (p < 0,001), whereas the fatty acid 20:4 Δ5,8,11,14 showed the opposite trend (p < 0.01). We conclude that these two fatty acids are potential biomarkers for NO and antimony resistance in L. chagasi and L. amazonensis and that they could be helpful in therapeutic diagnoses.

Antimicrobial potential of Actinomycetes by NRPS and PKS-I pathways

BackgroundActinomycetes may account for 10 to 30% of the totalsoil rhizosphere microorganisms. The attention given tothe actinomycetes in biotechnological applications is aresult of their metabolic versatility that is accompaniedby the production of primary and secondary metabolitesof economic importance, which are a promising source ofproducts (e.g., antibiotics, enzyme inhibitors, antiparasiticand anticancer agents) [1,2]. Included in this range ofcompounds are secondary metabolites synthesized bypolyketide synthase (PKS) and non-ribosomal peptidesynthetase (NRPS) pathways. An effective method forassessing the presence of these biosynthetic pathways isthe detection of PKS and NRPS genes by PCR [3,4].Thus, this study was based on targeted analyses of 31 soilisolate actinomycetes aiming to evaluate their antimicro-bial potential through the NRPS and PKS-I pathways.MethodsThe antimicrobial activity was evaluated by the antagon-ism test against two economically important phyto-pathogens, the bacterium Xanthomonas campestris andthe fungus Thielaviopsis paradoxa,usingthetechniqueof double layer. TheX. campestris and T. paradoxawere propagated at 28°C in YM (yeast malt) pH 6.0 andPDB (potato dextrose broth) respectively. The resultswere statistically analyzed using the Bonferroni test.The presence of genes PKS and NRPS was evaluatedby PCR, using degenerate primers for highly conservedregions encoding enzymes associated with biosynthesisof polyketides and peptides.Results and conclusionsAccording to the experimental results, 52% of the iso-lates showed antimicrobial activity against at least oneof the target bacterial pathogens tested. Among theseactive isolates, some belong to rare families. Thus, thisfinding can be a source of novel biomolecules with anti-microbial activity. From those isolates that presentedone of the NRPS and PKS-I genes, 75% of them showedantagonistic activity against one of the phytopathogensevaluated. Preliminary data on this screening demon-strate the importance of the biotechnological potentialof these actinomycetes due to the antagonistic activityagainst plant pathogens of economic interest and thepossibility of be used as biocontrol, besides offering astrong area for metabolic research [2,5].

Phenolic Composition and Leishmanicidal Activity of Red Propolis and Dalbergia ecastaphyllum (L.) Taub (Fabaceae) Extracts from Sergipe, Brazil

Leishmaniasis is a parasitic disease caused by protozoa of the Leishmania genus. It may manifest in visceral and tegumentary forms, and pentavalent antimonials are the first choice drugs used for the treatment. Frequently these drugs show low efficiency and high toxicity to mammalian host. The present study describes the chemical profile and the in vitro leishmanicidal effects of red propolis and Dalbergia ecastaphyllum extracts from Sergipe, Brazil, in Leishmania chagasi and Leishmania amazonensis promastigotes. The phenolic composition of the extracts was evaluated by direct infusion electrospray ionization mass spectrometry (ESI-MS) fingerprinting. The leishmanicidal effect was evaluated by the Resazurin colorimetric method. Similar composition profiles have been found for D. ecastaphyllum and propolis samples. The isoflavones formononetin, biochanin A, daidzein and pinocembrin were identified in both extracts. Propolis extract showed leishmanicidal activity in both L. chagasi and L. amazonensis, with IC50 values of 21.54 and 9.73 μg/mL, respectively. The D. ecastaphyllum extract presented activity only in L. amazonensis, with IC50 of 53.42 μg/mL. These results suggest that red propolis extract from Sergipe has the leguminosae D. ecastaphyllum as botanical origin, and that it presents potential leishmanicidal activity, which may be associated with the presence of the phenolic compounds found in its composition.

The role of cinnamon as a modulator of the expression of genes related to antioxidant activity and lipid metabolism of laying quails

Since cinnamon has vitamins and minerals in addition to antioxidants compounds in its chemical composition studies have shown the potential of cinnamon supplementation on some important characteristics in the performance of birds. Thus, this study was conducted under the hypothesis that the inclusion of cinnamon in the laying quail diet could influence the performance of the birds through the expression of genes related to antioxidant activity and lipid metabolism. To test this hypothesis, 144 Japanese quail (Coturnix japonica) with an initial age of 18 weeks and average weight of 133g were distributed in a completely randomized design with two treatments: no cinnamon supplementation (NCS—control group) and with supplementation of 9g/kg of cinnamon powder (CPS). The experiment lasted for 84 days. At the end of the experimental period, six animals from each treatment were euthanized by cervical dislocation, blood was collected and organs weighed. Liver tissue was collected for gene expression and biochemical analyses. We observed a significant effect of cinnamon inclusion on the weight of the pancreas (P = 0.0418), intestine (P = 0.0209) and ovary (P = 0.0389). Lower weights of the pancreas and intestine, and a higher ovary weight was observed in birds receiving the CPS diet. Quails fed with cinnamon supplementation also had better feed conversion per egg mass (2.426 g /g, P = 0.0126), and higher triglyceride (1516.60 mg/dL, P = 0.0207), uric acid (7.40 mg/dL, P = 0.0003) and VLDL (300.40 mg/dL, P = 0.0252) contents. A decreased content of thiobarbituric acid reactive substances (TBARS) and lower catalase activity was observed in the liver of quails from the CPS diet (0.086 nmoles/mg PTN, and 2.304 H2O2/min/mg PTN, respectively). Quails from the CPS group presented significantly greater expression of FAS (fatty acid synthase, 36,03 AU), ACC (Acetyl-CoA Carboxylase, 31.33 AU), APOAI (apolipoprotein A-I, 803,9 AU), ESR2 (estrogen receptor 2, 0.73 AU) SOD (superoxide dismutase, 4,933.9 AU) and GPx7 (glutathione peroxidase 7, 9.756 AU) than quails from the control group. These results allow us to suggest that cinnamon powder supplementation in the diet of laying quails can promote balance in the metabolism and better performance through the modulation of antioxidant activity and the expression of genes related to lipid metabolism.

Cytokinin Dehydrogenase activity in primary roots and characterization of primary metabolites from leaves and rootlets of Ricinus Communis

The aim of this study was to determine the activity of cytokinin dehydrogenase (CKX) and to measure other biochemical components in the primary leaves and radicles of castor seedlings (BRS Energia) in the initial phase of growth. The crude protein extract obtained after a 1-h extraction from the root tissues of seedlings showed no detectable CKX enzymatic activity when incubated with the substrate isopentenyl adenine for 1 h. However, after precipitation with ammonium sulfate at 70% saturation, the pellet showed CKX activity. The peroxidase enzyme activity was higher in the leaves than in the radicles. The total and reducing sugar content was 1.5 times higher in the leaves than in the radicles. The amino acid and protein contents were 6.4 and 9.2 times higher in the leaves than in the radicles, respectively.