Elba Verônica Matoso Maciel Carvalho

Characterization of acetylcholinesterase from the brain of the Amazonian tambaqui (Colossoma macropomum) and in vitro effect of organophosphorus and carbamate pesticides

In the present study, acetylcholinesterase (AChE) from the brain of the Amazonian fish tambaqui (Colossoma macropomum) was partially characterized and its activity was assayed in the presence of five organophosphates (dichlorvos, diazinon, chlorpyrifos, and tetraethyl pyrophosphate [TEPP]) and two carbamates (carbaryl and carbofuran) insecticides. Optimal pH and temperature were 7.0 to 8.0 and 45°C, respectively. The enzyme retained approximately 70% of activity after incubation at 50°C for 30 min. The insecticide concentration capable of inhibiting half of the enzyme activity (IC50) for dichlorvos, chlorpyrifos, and TEPP were calculated as 0.04 µmol/L, 7.6 µmol/L, and 3.7 µmol/L, respectively. Diazinon and temephos did not inhibit the enzyme. The IC50 values for carbaryl and carbofuran were estimated as 33.8 µmol/L and 0.92 µmol/L, respectively. These results suggest that AChE from the juvenile C. macropomum brain could be used as an alternative biocomponent of organophosphorus and carbamate biosensors in routine pesticide screening in the environment.

Comparative effect of pesticides on brain acetylcholinesterase in tropical fish.

Monitoring of pesticides based on acetylcholinesterase (AChE; EC 3.1.1.7) inhibition in vitro avoids interference of detoxification defenses and bioactivation of some of those compounds in non-target tissues. Moreover, environmental temperature, age and stress are able to affect specific enzyme activities when performing in vivo studies. Few comparative studies have investigated the inter-specific differences in AChE activity in fish. Screening studies allow choosing the suitable species as source of AChE to detect pesticides in a given situation. Brain AChE from the tropical fish: pirarucu (Arapaima gigas), cobia (Rachycentron canadum) and Nile tilapia (Oreochromis niloticus) were characterized and their activities were assayed in the presence of pesticides (the organophosphates: dichlorvos, diazinon, chlorpyrifos, temephos, tetraethyl pyrophosphate- TEPP and the carbamates: carbaryl and carbofuran). Inhibition parameters (IC₅₀ and Ki) for each species were found and compared with commercial AChE from electric eel (Electrophorus electricus). Optimal pH and temperature were found to be 8.0 and 35-45 °C, respectively. A. gigas AChE retained 81% of the activity after incubation at 50 °C for 30 min. The electric eel enzyme was more sensitive to the compounds (mainly carbofuran, IC₅₀ of 5 nM), excepting the one from A. gigas (IC₅₀ of 9 nM) under TEPP inhibition. These results show comparable sensitivity between purified and non-purified enzymes suggesting them as biomarkers for organophosphorus and carbamate detection in routine environmental and food monitoring programs for pesticides.

Impact of stress on Aeromonas diversity in tambaqui (Colossoma macropomum) and lectin level change towards a bacterial challenge

ABSTRACT Tambaqui (Colossoma macropomum) is among the most cultivated fish species in tropical countries. Stress is the main cause of disease in fish farms. The genus Aeromonas is a common causative agent of fish diseases. This work reports the identification of Aeromonas species colonizing gills of C. macropomum submitted or not to a confinement stress. We also evaluated changes in serum levels of lectins (carbohydrate-binding proteins that are components of fish immune system) in tambaqui submitted to a challenge using two isolated Aeromonas strains. Gill tissues from stressed and unstressed fishes were used to isolate Aeromonas. Then 72 Aeromonas strains were isolated, 97% being from stressed fishes. Among these, 63 were identified at species level and 6 were classified as atypical Aeromonas strains. The most prevalent species were Aeromonas bestiarum and Aeromonas caviae and their strains were used in bacterial challenges. The lectin serum levels significantly increased after 24 h of infection with A. bestiarum; however, no significant increase was found for infection with A. caviae. In conclusion, C. macropomum gills are susceptible to colonization by different Aeromonas species, mainly at confinement stressful conditions, and serum lectins may have a role in the acute immunological response towards infection by A. bestiarum.

Seasonality Influence on Biochemical and Hematological Indicators of Stress and Growth of Pirarucu (Arapaima gigas), an Amazonian Air-Breathing Fish

Environmental factors such as seasonal cycles are the main chronic stress cause in fish increasing incidence of disease and mortality and affecting productive performance. Arapaima gigas (pirarucu) is an Amazonian air-breathing and largest freshwater fish with scales in the world. The captivity development of pirarucu is expanding since it can fatten up over 1 kg per month reaching 10 kg body mass in the first year of fattening. This work was conducted in three periods (April to July 2010, August to November 2010, and December 2010 to March 2011) defined according to rainfall and medium temperatures. Seasonality effect analysis was performed on biochemical (lectin activity, lactate dehydrogenase, and alkaline phosphatase activities) and hematological (total count of red blood cells, hematocrit, hemoglobin, and hematimetric Wintrobe indexes) stress indicators, as well as on growth and wellbeing degree expressed by pirarucu condition factor developed in captivity. All biochemical and hematological stress indicators showed seasonal variations. However, the fish growth was allometrically positive; condition factor high values indicated good state of healthiness in cultivation. These results reinforce the robust feature of pirarucu and represent a starting point for understanding stress physiology and environmental changes during cultivation enabling identification and prevention of fish adverse health conditions.

Lectins as mitosis stimulating factors: Briefly reviewed

ABSTRACT Lectins are carbohydrate binding proteins that can stimulate cell proliferation. This property makes these biomolecules capable of being used as mitogen reagents to study the interaction with lymphocytes allowing evaluation of immunomodulatory action, since B and T lymphocytes are related to humoral and innate immunity, respectively. Isolated cells from spleen, which include lymphocytes, are widely applied as a model in screening lectin mitogenic capacity. This mitotic stimulus is initiated by interaction of the lectin with T‐cell receptor on cell surface. This brief review article aims to explain how cell proliferation, especially lymphocytes, can be achieved through lectin induction. Additionally, this work intends to highlight the main colorimetric and radiographic techniques to encourage the scientific community in searching for new mitogenic lectins.