Immunohistochemical Approach to Study Cylindrospermopsin Distribution in Tilapia (Oreochromis niloticus) under Different Exposure Conditions

Abstract

Cylindrospermopsin (CYN) is a cytotoxic cyanotoxin produced by several species of freshwater cyanobacteria (i.e., Aphanizomenon ovalisporum). CYN is a tricyclic alkaloid combined with a guanidine moiety. It is well known that CYN inhibits both protein and glutathione synthesis, and also induces genotoxicity and the alteration of different oxidative stress biomarkers. Although the liver and kidney appear to be the main target organs for this toxin based on previous studies, CYN also affects other organs. In the present study, we studied the distribution of CYN in fish (Oreochromis niloticus) under two different exposure scenarios using immunohistochemical (IHC) techniques. In the first method, fish were exposed acutely by intraperitoneal injection or by gavage to 200 µg pure CYN/Kg body weight (bw), and euthanized after 24 h or five days of exposure. In the second method, fish were exposed by immersion to lyophilized A. ovalisporum CYN-producing cells using two concentration levels (10 or 100 µg/L) for two different exposure times (7 or 14 days). The IHC was carried out in liver, kidney, intestine, and gills of fish. Results demonstrated a similar pattern of CYN distribution in both experimental methods. The organ that presented the most immunopositive results was the liver, followed by the kidney, intestine, and gills. Moreover, the immunolabeling signal intensified with increasing time in both assays, confirming the delayed toxicity of CYN, and also with the increment of the dose, as it is shown in the sub-chronic assay. Thus, IHC is shown to be a valuable technique to study CYN distribution in these organisms.

Effects of cylindrospermopsin on the phagocytic cells of the common carp (Cyprinus carpio L.)

“…Fish may be exposed to the toxin via several routes, including by dermal exposure through skin and gills, or by oral uptake. After entering the body, CYN accumulates in various tissues (Guzmán‐Guillén et al ., ; Kinnear, ), in fish mainly in the liver and kidneys (Guzmán‐Guillén et al ., ). As a highly water‐soluble molecule, CYN also has a high affinity for blood and lymph (Kinnear, ).…”

Evidence of cylindrospermopsin uptake and clearance in fish (Oreochromis niloticus) under laboratory conditions

“…In recent years, a number of studies have been conducted on the acute effects of CYN on fish. In vivo and in vitro research studies have shown that CYN toxicity might be mediated by reactive oxygen species (ROS) (Liebel et al ., 2011; Silva et al ., 2017) and, as a result, lead to alterations, in areas such as the antioxidant defence system, lipid peroxidation, protein carbonilation and DNA oxidation (Guzmán‐Guillén et al ., 2014, 2015a; Puerto et al ., 2011, 2014). In addition, it has had biological effects ( e.g ., developmental toxicity and a high mortality rate) on embryo fish ( Danio rerio ).…”

DNA damage induced by cylindrospermopsin on different tissues of the biomonitor fish Poecilia reticulata

“…In O. niloticus , animals sampled at fish pounds in Egypt accumulated up to 1.5 μg g −1 of CYN in their livers 20 . For this same species, a strongest immunolocalization of CYN in the liver was detected, and the immunolabeling was intensified over time after exposure, demonstrating the biocamulation and toxicity of CYN in the liver of animals after acute and sub‐chronic CYN‐exposure 26 . For H. malabaricus , in animals exposed to a single intraperitomial injection (50 μg k −1 ) of aqueous extract of CYN‐producing cyanobacteria, on average 6.79 and 5.61 ng g −1 of CYN was detected in the liver, after 7 and 14 days of exposition, respectively 24 …”

“…Although in vivo studies evaluating the toxic effects of CYN are related to a few fish species, 21‐27 they have demonstrated a broad action of CYN in several target organs. However, there are no data about the putative genotoxicity and mutagenicity caused by CYN in different tissues from fish.…”