Vicilin from Anadenanthera colubrina Seeds: An alternative tool to combat Callosobruchus maculatus.

Vicilins are seed proteins, and they constitute 70-80% of the total protein in leguminous seeds; with amolecular mass between 150 and 190 kDa, they are composed of subunits without disulfide bridges, with high affinity for chitin-binding. They are also associated with seed defense against insect pests. The chitin-binding vicilin from Anadenanthera colubrina seeds was purified by ammonium sulfate, followed by affinity chromatography on a chitin column, molecular exclusion on Superdex 75 Tricorn in FPLC system and Phenomenex C8 chromatography in HPLC system. The A. colubrina vicilin, named AcV, is a tetrameric glycoprotein composed of 1.55% carbohydrates and molecular weight determined by SDS-PAGE, consisting of 70, 73, 43 and 41 kDa. The AcV homogeneity was confirmed in native PAGE, where it was observed to be a unique band with slow mobility in this gel, with approximately 230 kDa. AcV added to the Callosobruchus maculatus diet in the bioassays resulted in a strong effect on adult emergence (ED50 of 0.096%), and in larvae caused a marked reduction in mass (WD50 of 0.32%) and lethality (LD50 of 0.33%) (w:w). The digestibility of AcV was evaluated in vitro with the digestive enzymes of larvae of C. maculatus of fourth instar, showing major fragments of 10 and 30 kDa. AcV showed reactivity against the anti-EvV antibody from Erythrina velutina vicilin. The deleterious effects of AcV are likely to be associated with the chitin-binding fragments generated by proteolysis in the bruchid gut, similarly to that found for vicilins from other leguminous plant species, Enterolobium contortisiliquum and Vigna unguiculata. AcV might be a candidate protein for a possible bioinsecticidal control of the bruchid weevil, C. maculatus.

Intraocular pressure and pupil diameter in healthy cats anesthetized with isoflurane and pre-medicated with isolated acepromazine or in combination with tramadol / [Pressão intraocular e diâmetro pupilar em gatos saudáveis anestesiados com isoflurano e pré-medicados com acepromazina isolada ou em combinação com tramadol]

The objective of this study was to determine changes on intraocular pressure (IOP) and pupil diameter (PD) in healthy cats anesthetized with isoflurane, and premedicated with acepromazine alone or in combination with tramadol. Thirty cats were allocated in two groups (n=15/each) and were treated with acepromazine (AG) or acepromazine/tramadol (ATG). PD and IOP were assessed before and following 30 (PM1), and 40 minutes (PM2) of treatments. Anesthesia was induced with propofol, and IOP and DP were recorded (A10) at 10 minute intervals until the end of anesthesia (A40). IOP decreased in AG and ATG, when comparing baseline with PM1. IOP decreased only in AG, in comparisons between baseline and PM2. During anesthesia, IOP did not change within and between groups. Comparisons between baseline with those recorded at PM1 and 2 showed that PD increased in the ATG. During anesthesia, PD decreased significantly in AG and ATG. Both protocols maintained the IOP within the reference range to perform corneal or intraocular surgery in healthy cats but did not sustain pre-anesthetic pupil dilation observed in ATG.(AU)

O objetivo do presente artigo é determinar possíveis alterações na pressão intraocular (PIO) e no diâmetro pupilar (DP) em gatos saudáveis anestesiados com isoflurano e pré-medicados com acepromazina isolada ou em combinação com acepromazina/tramadol. Trinta gatos saudáveis foram distribuídos aleatoriamente em dois grupos (n=15/cada) e tratados com acepromazina (GA) ou acepromazina/tramadol (GAT). DP e PIO foram avaliadas antes (basal) e após 30 (PM1) e 40 minutos (PM2) dos tratamentos. A anestesia foi induzida com propofol, e a PIO e o DP foram registrados (A10) a cada 10 minutos até o final da anestesia com isoflurano (A40). Ao se compararem os valores obtidos no basal com PM1, a PIO diminuiu em GA e GAT; com PM2, a PIO reduziu apenas no GA. Durante a anestesia, a PIO não diferiu dentro e entre os grupos. Comparações entre os valores basais e os registrados em PM1 e em PM2 mostraram que a DP aumentou significativamente no GAT. Durante a anestesia, o DP diminuiu significativamente em GA e GAT. Ambos os protocolos mantêm a PIO dentro dos valores de referência para realizar cirurgias corneanas ou intraoculares em gatos saudáveis, mas não sustentam a dilatação pupilar pré-anestésica observada em GAT.(AU)

Impacto do grau de uveíte em diferentes tipos de úlceras de córneas em cães submetidos ao enxerto pediculado de conjuntiva bulbar - 34 casos / Impact of uveitis score on different types of corneal ulcers in dogs subjected to the bulbar conjunctival pedicle graft - 34 cases

O presente trabalho objetivou avaliar os resultados de 34 olhos submetidos ao enxerto conjuntival pediculado (ECP) em ceratites ulcerativas profundas (n=5), em ceratites ulcerativas com colagenólise (n=2), descemetocele (n=7), perfuração corneal (n=15) e prolapso de íris (n=5). Os impactos do grau de uveíte e da integridade da córnea foram correlacionados com presença e ausência de visão por tabelas de contingência. Raças braquicefálicas foram acometidas em 91,11% dos casos. O número de córneas consideradas perfuradas [20/34 (58,82%)] foi maior que o de córneas íntegras [14/34 (41,17%)]. Ao 50º dia de pós-operatório, o número de olhos visuais que apresentavam córneas íntegras previamente às cirurgias (n=13) não diferiu significativamente dos olhos com córneas perfuradas (n=12) (P=0,05). Avaliações relativas ao grau de uveíte, revelaram que a metade dos casos foi considerada severa [17/34 (50%)] e na outra metade as uveítes foram consideradas discretas. Dos 17 casos que apresentaram uveíte severa, oito recuperaram a visão. Já nos 17 olhos onde a uveíte foi considerada leve, 15 mantiveram a visão ao final do período de avaliação. Apesar de a integridade da córnea não se correlacionar com a severidade da uveíte (P=0,48), constatou-se que o número de olhos visuais com uveíte discreta foi significativamente maior que os olhos com uveíte severa (P=0,006). Neste estudo, a taxa geral de sucesso visual após ECP foi de 73,52% e a integridade da córnea não exerceu impacto significativo sobre a manutenção da visão. Todavia, olhos acometidos por uveítes severas apresentaram menor chance de recuperar a visão.(AU)

The present study aimed to evaluate the results of 34 eyes of dogs presenting deep corneal ulcer (n=5), colagenolytic corneal ulcer (n=2), descemetocele (n=7), corneal perforation (n=15), and iris prolapse (n=5) that were corrected by the bulbar conjuntival pedicle graft (CPG). The uveitis score and the corneal integrity were correlated with the presence or absence of vision by contingency tables. Brachycephalic breeds accounted for 91.11% of all cases. The number of perforated corneas [20/34 (58.82%)] were higher than the non-perforated ones [14/34 (41.17%)]. At post-operative day 50, the number of visual eyes with non-perforated corneas before surgery (n=13) did not differ significantly from the eyes with perforated corneas (n=12) (P = 0.05). In half of the cases, uveitis score was considered severe [17/34 (50%)], and in the other half, mild. Eight out of 17 eyes classified with severe uveitis regained vision. In 17 eyes where uveitis score was considered mild, 16 regained vision at the end of the study. Although corneal integrity was non-correlated with the uveitis score (P = 0.48), the number of visual eyes scored with mild uveitis were significantly larger than the eyes with severe uveitis (P = 0.006). In this study, the overall success visual rate after CPG was 73,52% % and the corneal integrity did not play an important role regarding maintenance of vision. However, eyes presenting severe uveitis score had less chance to regain vision.(AU)

Molecular cloning and characterization of an α-amylase cDNA highly expressed in major feeding stages of the coffee berry borer, Hypothenemus hampei.

α-Amylases are common enzymes responsible for hydrolyzing starch. Insect-pests, whose larvae develop in seeds, rely obligatorily on α-amylase activity to digest starch, as their major food source. Considering the relevance of insect α-amylases and the natural α-amylase inhibitors present in seeds to protect from insect damage, we report here the molecular cloning and nucleotide sequence of the full-length AmyHha cDNA of the coffee berry borer, Hypothenemus hampei, a major insect-pest of coffee crops. The AmyHha sequence has 1879 bp, containing a 1458 bp open reading frame, which encodes a predicted protein with 485 amino acid residues, with a predicted molecular mass of 51.2 kDa. The deduced protein showed 55-79% identity to other insect α-amylases, including Anthonomus grandis, Ips typographus and Sitophilus oryzae α-amylases. In depth analysis revealed that the highly conserved three amino acid residues (Asp184, Glu220, and Asp285), which compose the catalytic site are also presented in AmyHha amylase. The AmyHha gene seems to be a single copy in the haploid genome and AmyHha transcription levels were found higher in L2 larvae and adult insects, both corresponding to major feeding phases. Modeling of the AmyHha predicted protein uncovered striking structural similarities to the Tenebrio molitor α-amylase also displaying the same amino acid residues involved in enzyme catalysis (Asp184, Glu220 and Asp285). Since AmyHha gene was mostly transcribed in the intestinal tract of H. hampei larvae, the cognate α-amylase could be considered a high valuable target to coffee bean insect control by biotechnological strategies.