NANOTECNOLOGY AS A NEW METHOD FOR ANTIBIOTIC MENAGEMENT IN AQUACULTURE.

Tons of drugs are produced annually worldwide for consumption by humans or use in the treatment of animals. The release and presence of drugs in the environment is a highly hot topic, which was firstly highlighted in the US in the 70's and almost a decade later in England. Yet, it was in the mid-90s, with the progress of analytical techniques, that the knowledge about environmental contamination caused by these compounds excelled considerably. Most of these drugs are poorly metabolized by animals after ingestion and a fraction ranging from 25% to 75% may be released into the environment after consumption. Recent studies have shown concentrations of different types of drugs in wastewater, surface waters, in sea water, groundwater and drinking water equal to mg-ng L^-1. Especially antibiotics and antiparasitic drugs are abundantly used for veterinary purposes. Oxytetracycline is one of the most commonly, wide spectrum, antibiotics used, typically administered orally, (incorporated into pellet feed) or via water. This antibiotic is characterized by a poor adsorption (25 to 30%) that often results in the production of large amounts of wastes. The release of these wastes into the aquatic environment can cause threat to aquatic organisms. For these reasons, a better management of antibiotic administration is considered of primary importance. At this regard, nanotechnologies, and in particular nanoparticles (NP) are regarded with great interest because of  their unique chemical-physical properties that make them ideal for several biomedical applications, including drug delivery. Adult Zebrafish were divided in 5 experimental groups of 150 fish each (in triplicate) and exposed to the following conditions for 28 days: Control; Treatment A: 50mg/L oxytetracycline added to the water; Treatment B: fed on food (2% bodyweight) added with 7.36g oxytetracycline/kg food; Treatment C: 100mg/L SAMN@oxytetracycline equivalent to a concentration of 4mg of oxytetracycline; Treatment D: 100mg/L SAMNs; Group E: 4mg/L oxytetracycline added to the water.

Samplings were performed at 0, 14 and 28 days after the beginning of the treatment, and water samples were collected once a week from all tanks to determine the presence of oxytetracycline in the water. Brain, liver, skin, digestive tract, gonads and whole animals were collected to determine the concentration of oxytetracycline through liquid chromatography assay. The anatomy of the digestive tract, gills and brain was performed through histological analysis(hematoxylin & eosin staining), and finally the relative   expression of genes involved in fish welfare (hsp70, hnf4a, sod1, sod2, gstal, nr3c1) growth (igf1, igf2a, mstnb), appetite (kiss1, kiss2, npy, gnrh3, cnr1, lepa) and reproduction (vtg, esr1, esr2b, esr2a),  were analyzed by Real time PCR.  Water and food analysis confirmed a stable concentration of oxytetracycline during the experiment. Results obtained from liquid chromatography revealed the accumulation of oxytetracycline in all the selected organs. Histological analysis showed effects on gills morphology causing thickening of the lamellae and hyperplasia of the epithelium. Livers collected from fish of treatment A presented hydropic degeneration while fish from treatment B showed hepatic steatosis. Finally, the digestive tract of treated fish showed several rodlet cells.    Molecular analysis performed by Real Time PCR revealed a growth promoting action by oxytetracycline. Finally oxytetracycline had no estrogenic effects on Zebrafish.

This study examined a possible application on nanotechnologies on oxytetracycline administration. Therapeutic concentrations can have some adverse effects on Zebrafish and thus a better administration through the use of nanoparticles can have positive effects both on fish and environment.