Cléberson de Freitas Fernandes

Componentes primários e secundários do rendimento de óleo de pinhão-manso

The definition of the most suitable plant selection criteria is especially important for Jatropha curcas L. as subsidies the genetic progress estimates and the ideotype characterization, described in terms of the traits that are thought to enhance genetic yield potential. The objective of this study was to estimate the direct and indirect effects of seven vegetative traits and four seed quality traits on J. curcas oil yield. The traits grain yield and the weight of the nuts showed direct and positive effect on the oil yield. The tree volume showed the major indirect and positive effect on the main variable through grain yield. The development of J. curcas with higher oil yield in the evaluated population has to consider the selection of plants with higher yield and tree volume, maintaining the variability of the grain quality traits.

Inhibition of the Myotoxicity Induced by Bothrops jararacussu Venom and Isolated Phospholipases A2 by Specific Camelid Single-Domain Antibody Fragments

Antivenoms, produced using animal hyperimmune plasma, remains the standard therapy for snakebites. Although effective against systemic damages, conventional antivenoms have limited efficacy against local tissue damage. Additionally, the hypersensitivity reactions, often elicited by antivenoms, the high costs for animal maintenance, the difficulty of producing homogeneous lots, and the instability of biological products instigate the search for innovative products for antivenom therapy. In this study, camelid antibody fragments (VHH) with specificity to Bothropstoxin I and II (BthTX-I and BthTX-II), two myotoxic phospholipases from Bothrops jararacussu venom, were selected from an immune VHH phage display library. After biopanning, 28 and 6 clones recognized BthTX-I and BthTX-II by ELISA, respectively. Complementarity determining regions (CDRs) and immunoglobulin frameworks (FRs) of 13 VHH-deduced amino acid sequences were identified, as well as the camelid hallmark amino acid substitutions in FR2. Three VHH clones (KF498607, KF498608, and KC329718) were capable of recognizing BthTX-I by Western blot and showed affinity constants in the nanomolar range against both toxins. VHHs inhibited the BthTX-II phospholipase A2 activity, and when tested for cross-reactivity, presented specificity to the Bothrops genus in ELISA. Furthermore, two clones (KC329718 and KF498607) neutralized the myotoxic effects induced by B. jararacussu venom, BthTX-I, BthTX-II, and by a myotoxin from Bothrops brazili venom (MTX-I) in mice. Molecular docking revealed that VHH CDRs are expected to bind the C-terminal of both toxins, essential for myotoxic activity, and to epitopes in the BthTX-II enzymatic cleft. Identified VHHs could be a biotechnological tool to improve the treatment for snake envenomation, an important and neglected world public health problem.

Camelid Single-Domain Antibodies (VHHs) against Crotoxin: A Basis for Developing Modular Building Blocks for the Enhancement of Treatment or Diagnosis of Crotalic Envenoming

Toxic effects triggered by crotalic envenoming are mainly related to crotoxin (CTX), composed of a phospholipase A2 (CB) and a subunit with no toxic activity (CA). Camelids produce immunoglobulins G devoid of light chains, in which the antigen recognition domain is called VHH. Given their unique characteristics, VHHs were selected using Phage Display against CTX from Crotalus durissus terrificus. After three rounds of biopanning, four sequence profiles for CB (KF498602, KF498603, KF498604, and KF498605) and one for CA (KF498606) were revealed. All clones presented the VHH hallmark in FR2 and a long CDR3, with the exception of KF498606. After expressing pET22b-VHHs in E. coli, approximately 2 to 6 mg of protein per liter of culture were obtained. When tested for cross-reactivity, VHHs presented specificity for the Crotalus genus and were capable of recognizing CB through Western blot. KF498602 and KF498604 showed thermostability, and displayed affinity constants for CTX in the micro or nanomolar range. They inhibited in vitro CTX PLA2 activity, and CB cytotoxicity. Furthermore, KF498604 inhibited the CTX-induced myotoxicity in mice by 78.8%. Molecular docking revealed that KF498604 interacts with the CA–CB interface of CTX, seeming to block substrate access. Selected VHHs may be alternatives for the crotalic envenoming treatment.

Acaricidal activity of extracts from different structures of Piper tuberculatum against larvae and adults of Rhipicephalus microplus

Estrategias de controle do carrapato dos bovinos, Rhipicephalus microplus, fundamentam-se na utilizacao de pesticidas. Os principais grupos quimicos utilizados atualmente mostram-se ineficazes devido ao surgimento de populacoes resistentes. A pesquisa de novas moleculas com eficiencia acaricida e uma necessidade para manutencao da produtividade dos rebanhos bovinos estabelecidos em regioes de clima tropical. Avaliamos a atividade de Piper tuberculatum para o controle de femeas ingurgitadas e larvas de R. microplus atraves de bioensaios de imersao de adultos e de pacotes de larvas. As concentracoes avaliadas foram de 50; 25; 12,50; 6,25; 3,12 e 1,56 mg mL-1 de extrato etanolico de folha, talo e fruto de P. tuberculatum. As concentracoes letais para 50% dos individuos (CL50) apos 24 horas de exposicao de larvas de R. microplus foram 3,62; 3,99 e 5,30 mg mL-1 para os extratos etanolicos de fruto, talo e folha, respectivamente. Para femeas ingurgitadas, a maior eficacia resultou da concentracao de 50 mg mL-1 de extrato de fruto (71.57%). O principal efeito dos extratos etanolicos de P. tuberculatum foi sobre a eclodibilidade, com uma reducao de 55.63% para extratos de fruto e folha. P. tuberculatum mostra-se promissora como fonte de moleculas candidatas a uso em pesticidas, em formulacoes destinadas ao controle das infestacoes de R. microplus.

Antifungal activity of Hedychium coronarium J. König essential oil against Fusarium oxysporum Schlecht and Thanatephorus cucumeris (A.B. Frank) Donk in vitro

Fusarium oxysporum Schlecht and Thanatephorus cucumeris (A.B. Frank) Donk are fungi which cause some diseases in important plant cultures in Brazil - like bananas and beans. Hedychium coronarium J. Konig is a native plant from Asia that currently occurs in Brazil, where it is used in folk medicine to treat infections in general. The objective of this work was to evaluate the effect of the H. coronarium essential oil from leaves on the in vitro growth of F. oxysporum and T. cucumeris colonies. Discs of 5 mm diameter from isolated cultures of each fungi were placed in the center of 90 mm diameter Petri dishes with potato dextrose agar. In the peripheral area of the dishes four discs of filter paper were placed with 10 µL of essential oil, extracted through distillation system. For a control treatment, discs without essential oil were used. A completely randomized design was used with four replications (of four dishes). Every 24 hours for 8 days the fungi growth was evaluated by measuring the diameter of the colonies. After 192 hours F. oxysporum colonies had 37.4 mm diameter in the treatment with essential oil, while in the control the diameter was 66.4 mm. T. cucumeris colonies reached 11.9 mm diameter with essential oil and covered the dishes in the control. The results showed that the essential oil from leaves of H. coronarium has an inhibitory effect on the in vitro growth of F. oxysporum and T. cucumeris , thus suggesting its potential use in agriculture, mainly with respect to banana and bean cultures. Keywords: phytopathology, medicinal plants, biological control.