A proteomic overview of the major venom components from Tityus championi from Panama.

In recent years, morbidity caused by scorpion sting of the species Tityus championi has increased in Panama. Therefore, the LD50 was determined by intravenous injection in 2.9 mg/kg and the venom of T. championi was separated using a HPLC system and their fractions were tested for biological activities in mice to identify the most toxic fractions to mammals. In addition, the venom fractions were also tested against invertebrates to look for insect-specific toxin peptides. The most toxic fractions were analyzed by MS/MS spectrometry. The primary structures of T. championi venom peptides with the most relevant activity were obtained, and the primary structure of one of most neurotoxic peptides was found at least in other four species of Tityus from Panama. This neurotoxin is quite important to be used as a protein target to be neutralized if developing antivenoms against the sting of this Panamanian scorpion or other relevant species of genera Tityus in the country.

Heterologous expression of four recombinant toxins from Panamanian scorpions of the genus Tityus and Centruroides for production of antivenom.

BACKGROUND: The development of more effective antivenoms remains a necessity for countries where scorpionism is a public health problem. Also, the regionalization of antivenoms may be important for some countries with special scorpionism characteristics. OBJECTIVE: Production of antibodies capable of neutralizing the lethal effect of the venom of three scorpion species from Panama. METHODS: The primary structures of two neurotoxins from T. pachyurus, one from T. cerroazul and another from C. bicolor were elucidated using N-terminal amino acid degradation and Sanger gene cloned sequencing. The obtained mRNA transcripts were cloned and expressed using E. coli vectors. Different bacterial expression conditions were tested and the best culture conditions for each expressed protein is reported. The expressed scorpion toxins were purified by chromatographic methods and used as immunogens in rabbits. RESULTS: The antibodies produced under the reported immunization scheme show better neutralization (ED50) than other reported commercial antivenoms used to neutralize similar species scorpion venoms under similar LD50 conditions. CONCLUSION: The information reported here shows the proof of concept for selecting recombinant immunogens with the ability to produce antibodies for neutralizing the lethal effects of the most important medical species of scorpions in Panama.

Venoms of Centruroides and Tityus species from Panama and their main toxic fractions.

The scorpionism in Panama is notorious for the confluence and coexistence of buthid scorpions from the genera Centruroides and Tityus. This communication describes an overview of the larger representative toxic venom fractions from eight dangerous buthid scorpion species of Panama: Centruroides (C. granosus, C. bicolor, C. limbatus and C. panamensis) and Tityus (T. (A.) asthenes, T. (A.) festae, T. (T.) cerroazul and T. (A.) pachyurus). Their venoms were separated by HPLC and the corresponding sub-fractions were tested for lethality effects on mice and insects. Many fractions toxic to either mice or insects, or both, were found and have had their molecular masses determined by mass spectrometry analysis. The great majority of the lethal components had a molecular mass close to 7000 Da, assumed to be peptides that recognize Na+-channels, responsible for the toxicity symptoms observed in other buthids scorpion venoms. A toxic peptide isolated from the venom of T. pachyurus was sequenced by Edman degradation, allowing the synthesis of nucleotide probe for cloning the correspondent gene. The mature toxin based on the cDNA sequencing has the C-terminal residue amidated, contains 62 amino acid packed by 4 disulfide linkages, with molecular mass of 7099.1 Da. This same toxic peptide seems to be present in scorpions of the species T. pachyurus collected in 5 different regions of Panama, although the overall HPLC profile is quite different. The most diverse neurotoxic venom components from the genus Centruroides were found in the species C. panamensis, whereas T. cerroazul was the one from the genus Tityus. The most common neurotoxins were observed in the venoms of T. festae, T. asthenes and T. pachyurus with closely related molecular masses of 7099.1 and 7332 Da. The information reported here is considered very important for future generation of a neutralizing antivenom against scorpions from Panama. Furthermore, it will contribute to the growing interest in using bioactive toxins from scorpions for drug discovery purposes.

Geographical variability of the venoms of four populations of Bothrops asper from Panama: Toxicological analysis and neutralization by a polyvalent antivenom.

Bothrops asper is the medically most important venomous snake in Central America. In Panama, the country having the highest incidence of snakebites in Latin America, B. asper is widely distributed throughout the country and is responsible for the vast majority of snakebites. This study was performed to analyze whether there are variations in the toxicological profile and in some biochemical parameters between the venoms of B. asper from four different regions in Panama. The venoms showed a similar profile of lethal, hemorrhagic, in vitro coagulant, defibrinogenating, edema-forming, myotoxic and indirect hemolytic activities, with subtle quantitative variations between samples of some regions. The venoms also had similar SDS-PAGE patterns and reverse phase HPLC profiles. A polyvalent antivenom manufactured in Costa Rica, and regularly used in Panama, was effective in the neutralization of lethal activity of the venoms of the four populations, with Mean Effective Doses (ED50) ranging from 5.98 to 9.72 mg venom/mL antivenom. In agreement, a widespread pattern of cross-reactivity between this antivenom and the four venoms was observed by immunoblotting. Overall, results highlight the lack of marked differences between the venoms of the various populations of B. asper in Panama, and that the antivenom from Costa Rica is effective in neutralizing lethality.

Estudo químico e avaliação da atividade antimalárica dos galhos de Piranhea trifoliata / Chemical study and antimalarial activity evaluation of branches of Piranhea trifoliata

Este trabalho teve como objetivos realizar o estudo químico dos galhos de Piranhea trifoliata (Picrodendraceae) e avaliar o seu potencial antimalárico. Foram realizadas duas coletas no Estado do Pará. O material foi seco, moído e extraído com diclorometano (DCM), metanol (MeOH) e água (H2O). Os extratos DCM e MeOH foram submetidos à análise em cromatografia em camada delgada comparativa (CCDC), e apresentaram características principalmente de terpenos. Do estudo fitoquímico do extrato DCM da 2ª coleta, foi isolada: friedelan-3-ona e do extrato MeOH da 2ª coleta foi isolado: o triterpeno esqualeno, e mistura dos esteroides estigmasterol e ß-sitosterol. Quanto à atividade antimalárica, os extratos e as fases dos extratos MeOH foram avaliados in vitro frente ao Plasmodium falciparum, e os extratos MeOH da 1ª e 2ª coletas foram considerados ativos (CI50 = 13,7 µg/mL; CI50 = 5,8 µg/mL, respectivamente). Este trabalho contribuiu para o conhecimento biológico e químico da espécie Piranhea trifoliata e da família Picrodendraceae, sendo este o primeiro relato de atividades biológicas para a espécie em estudo.

Evaluation of anti-Bothrops asper venom activity of ethanolic extract of Brownea rosademonte leaves.

Significant inhibition of the coagulant and hemorrhagic effects of Bothrops asper venom was demonstrated by ethanolic extract prepared from the leaves of Brownea rosademonte. In vitro experiments preincubating 5.5 mg of extract kg-1 b.m. for 30 min with a minimum hemorrhagic dose of venom (273.8 ± 16.1 µg of venom kg-1 b.m.) lowered the hemorrhagic activity of the venom alone in CD-1 mice by 51.5 ± 2.6 %. Additionally, 1.7 mg extract L-1 plasma prolonged 5.1 times the plasma coagulation time. Fractionation of the extract led to the isolation of two compounds: ononitol (1) and quercetrin (2). The structure of compounds 1 and 2 was established by spectroscopic analyses, including APCI-HRMS and NMR (1H, 13C, HSQC, HMBC and COSY). A quercetrin concentration of 0.11 µmol L-1 prolonged the plasma coagulation time 2.6 times demonstrating that this compound was one of the active constituents of the Brownea rosademonte extract.

Genomagnetic electrochemical assays of DNA hybridization.

An electrochemical genomagnetic hybridization assay has been developed to take advantage of a new and efficient magnetic separation/mixing process, the amplification feature of enzyme labels, and single-use thick-film carbon transducers operated in the pulse-voltammetric mode. It represents the first example of coupling a magnetic isolation with electrochemical detection of DNA hybridization. The new protocol employs an enzyme-linked sandwich solution hybridization, with a magnetic-particle labeled probe hybridizing to a biotinylated DNA target that captures a streptavidin-alkaline phosphatase (AP). The alpha-naphthol product of the enzymatic reaction is quantitated through its well-defined, low-potential (+0.1 V vs. Ag/AgCl) differential pulse-voltammetric peak at the disposable screen-printed electrode. The efficient magnetic isolation is particularly attractive for electrical detection of DNA hybridization which is commonly affected by the presence of non-hybridized nucleic acid adsorbates. The new biomagnetic processing combines such magnetic separation with a low-volume magnetic mixing, and allows simultaneous handling of 12 samples. The attractive bioanalytical behavior of the new enzyme-linked genomagnetic electrical assay is illustrated for the detection of DNA segments related to the breast-cancer BRCA1 gene.

Origen anómalo de la arteria coronaria izquierda: reporte de un caso / Anomalous origin of left coronary artery: case report

Se presenta el caso de una paciente de 8 años que acude a control médico por soplo cardiaco. El ecocardiogama evidencia valvulopatía mitral; recibe tratamiento con digoxina y enalapril que producen una leve mejoría, pero posteriomlente se manifiesta disnea que conduce a una nueva evaluación. Se realiza nuevo esquema diagnóstico por medio de tomografia helicoidal y cateterismo que permiten diagnosticar anomalía coronaria izquierda que nace en la arteria pulmonar. La paciente ingresó a esta institución de salud para proceder a la cOITección quirúrgica del origen anómalo de la arteria coronaria y plastia valvular. Fue dada de alta sin complicaciones y con tratamiento ambulatorio profiláctico para endocarditis bacteriana por patología valvular.

Asma en la unidad de terapia intensiva desde 1982 hasta 1988 / Asthma in the Intensive Care Unit from 1982 to 1988

Se realizó un estudio retrospectivo de siete años de atención, en la Unidad de Terapia Intensiva (UTI), de los pacientes con status asmiticus. Se encontró que a pesar del aumento general de hospitalizaciones por asma de 16.5 (por ciento) a 22 (por ciento), en UTI este aumento fue mínimo de 0.5 (por ciento) a 0.6 (por ciento). Los menores de 5 años representaron el 87 (por ciento) del grupo estudiado y las 9 defunciones tenían menos de 5 años. Hubo un predominio del sexo femenino, el 55 (por ciento). Los factores de riesgo para el ingreso a UTI fueron : el asma familiar en 75 (por ciento), un promedio de tres hospitalizaciones previas en 66 (por ciento). El inicio del asma antes del año de edad y la demanda tardía de atención en 58 (por ciento). El cuadro clínico del status asmaticus mejoró en las primeras 24 horas y el 50 (por ciento) de los pacientes egresaron de UTI a las 72 horas. Antes de 1985 se informaron 40 (por ciento) de convulsiones, generalmente por hiponatremia. El esquema de tratamiento utilizado desde 1986 comprendió inhalaciones de salbutanol antes de las 12 horas del ingreso a UTI. Se recomienda la educación permanente y participativa del público sobre las causas, la clínica y el tratamiento del asma (AU)

Asma en la unidad de terapia intensiva desde 1982 hasta 1988 / Asthma in the Intensive Care Unit from 1982 to 1988

Se realizó un estudio retrospectivo de siete años de atención, en la Unidad de Terapia Intensiva (UTI), de los pacientes con status asmiticus. Se encontró que a pesar del aumento general de hospitalizaciones por asma de 16.5 (por ciento) a 22 (por ciento), en UTI este aumento fue mínimo de 0.5 (por ciento) a 0.6 (por ciento). Los menores de 5 años representaron el 87 (por ciento) del grupo estudiado y las 9 defunciones tenían menos de 5 años. Hubo un predominio del sexo femenino, el 55 (por ciento). Los factores de riesgo para el ingreso a UTI fueron : el asma familiar en 75 (por ciento), un promedio de tres hospitalizaciones previas en 66 (por ciento). El inicio del asma antes del año de edad y la demanda tardía de atención en 58 (por ciento). El cuadro clínico del status asmaticus mejoró en las primeras 24 horas y el 50 (por ciento) de los pacientes egresaron de UTI a las 72 horas. Antes de 1985 se informaron 40 (por ciento) de convulsiones, generalmente por hiponatremia. El esquema de tratamiento utilizado desde 1986 comprendió inhalaciones de salbutanol antes de las 12 horas del ingreso a UTI. Se recomienda la educación permanente y participativa del público sobre las causas, la clínica y el tratamiento del asma