Toxicon 54 (2009) 208–216 Contents lists available at ScienceDirect Toxicon journal homepage: www.elsevier.com/locate/toxicon Effects of dietary cyanobacteria of two different sources on growth and recovery of hybrid tilapia (Oreochromis niloticus  O. aureus) Guifang Dong a,b, Xiaoming Zhu a,*, Dong Han a, Yunxia Yang a, Lirong Song a, Shouqi Xie a,c a State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei 430072, China b Graduate University of Chinese Academy of Sciences, Beijing 100039, China c Aquaculture Divisions, E-Institute of Shanghai Universities, Shanghai, China article info abstract Article history: A 115 days feeding trial was conducted to evaluate the effect of dietary cyanobacteria on Received 22 December 2008 growth, microcystins (MCs) accumulation in hybrid tilapia (Oreochromis niloticus  O. Received in revised form 28 March 2009 aureus) and the recovery when the fish were free of cyanobacteria. Three experimental Accepted 30 March 2009 diets were formulated: the control (cyanobacteria free diet); one test diet with cyano- Available online 15 April 2009 bacteria from Lake Taihu (AMt, 80.0 mg MCs gÀ1 diet) and one with cyanobacteria from Lake Dianchi (AMd, 410.0 mg MCs gÀ1 diet). Each diet was fed to fish for 60 days and then Keywords: all fish were free of cyanobacteria for another 55 days. Cyanobacteria Growth A significant increase in feeding rate (FR) was observed in fish fed AMd diet after a first st Recovery 30-day exposure (1 EP), and in fish fed both AMt diet and AMd diet after a second 30-day nd Hybrid tilapia exposure (2 EP). Specific growth rates (SGR) of fish fed AMt diet and AMd diet were both obviously affected after the first 30-day exposure, but SGR was only significantly affected in fish fed AMt diet after the second 30-day exposure. After a 55-day recovery, there were no significant differences among diets in the indices mentioned above. Much higher concentrations of MCs were accumulated in tissues of all fish exposed to cyanobacteria. After the 55-day recovery, MC concentrations in fish tissues were significantly lower than those on day 60. Ó 2009 Elsevier Ltd. All rights reserved. 1. Introduction serious cyanobacterial blooms have occurred frequently in many freshwater lakes (e.g. Lake Dianchi, Lake Taihu, Lake Eutrophication in freshwater has induced the occurrence Chaohu, etc.) and most blooms produced MCs at high of dense cyanobacterial blooms (Carmichael, 1994). Some concentrations (Li et al., 2001; Song et al., 1998). Previous cyanobacteria including Anabaena, Aphanocapsa, Hapalosi- studies indicate that MCs accumulated in tissues of aquatic phon, Nostoc, Pseudanabaena, Planktothrix and Microcystis animals such as snails (Bellamya aeruginosa), shrimps could produce cyclic peptide toxins – microcystins (MCs) (de (Palaemon modestus and Macrobrachium nipponensis), and Figueiredo et al., 2004; Izaguirre et al., 2007) – which can fishes in those freshwater lakes (Chen and Xie, 2005; Chen cause liver failure in animals, livestock and aquatic life et al., 2005; Xie et al., 2005; Chen et al., 2007). (Carmichael,1994; Sivonen and Jones,1999), and even cause The acute effects of cyanobacteria or MCs on fish species human diseases or death (Azevedo et al., 2002). have been well investigated (Bury et al., 1995; Li et al., Fish deaths have been reported to be related to severe 2005). Generally, aquatic animals might be exposed to MCs cyanobacterial blooms (Rodger et al., 1994). In China, via the consumption of toxic cyanobacteria (Soares et al., 2004) or through gills (Zimba et al., 2001). Cyanobacteria could also be an important dietary component for many * Corresponding author. Tel.: þ86 27 68780060; fax: þ86 27 68780667. fish species including tilapia (Zurawell et al., 2005). E-mail address: [email protected] (X. Zhu). However, there is still a lack of studies on the chronic toxic 0041-0101/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.toxicon.2009.03.031 G. Dong et al. / Toxicon 54 (2009) 208–216 209 effects of dietary cyanobacteria on fishes (Zhao et al., 2005, Table 1 À1 2006a). It is also unclear if or how much the fish could Formulation and chemical composition of experimental diets (g 100 g in dry matter). recover when they are free of cyanobacteria. MCs not only affect the growth of fish, but also accu- Control AMt AMd mulate in their tissues (Magalha˜es et al., 2003; Mohamed Ingredients et al., 2003; Soares et al., 2004; Deblois et al., 2008). MCs Algae meal 0.0 43.6 50.8 contaminated fish may be a risk to human health from the White fishmeal (USA) 20.0 20.0 20.0 Soybean meal (oil-extracted) 49.9 0.0 0.0 food chain. To reduce potential risks, a tolerable daily Corn starch 10.2 10.2 10.2 À1 intake (TDI) of 0.04 mgkg total MCs per kilogram body Fish oil 5.3 5.8 6.0 weight per day as fish meat was used as a provisional a-Starch 6.0 6.0 6.0 a guideline (Chorus and Bartram, 1999). Mineral premix 0.4 0.4 0.4 Vitamin premixb 5.0 5.0 5.0 Previous studies in our laboratory demonstrated that Choline chloride 0.1 0.1 0.1 chronic exposure to lower levels of dietary cyanobacteria Cr2O3 0.5 0.5 0.5 increased the growth rate of Nile tilapia without impact on Cellulose 2.7 8.4 1.0 food conversion efficiency but high toxins were found to Chemical composition (g 100 gÀ1 in dry matter) accumulate in fish muscle and liver (Zhao et al., 2006). The Crude protein (%) 41.4 43.1 44.0 toxin level reported by Zhao et al. (2006) was even higher Crude lipid (%) 7.8 7.9 7.9 than the tolerable daily intake. If these contaminated fish Crude ash (%) 10.7 10.2 11.0 Gross energy (kJ gÀ1) 19.7 20.8 20.6 could recover when they were free of toxins is very Microcystins (mggÀ1) 0.0 80.0 410.0 important for food safety. a À1 $ The purpose of the present study was to investigate the Mineral premix (mg kg diet, H440): NaCl, 500; MgSO4 7H2O, 7500; NaH PO $2H O, 12,500; KH PO , 16,000; Ca(H PO ) $2H O,10,000; FeSO , chronic effects of dietary cyanobacteria on hybrid tilapia 2 4 2 2 4 2 4 2 4 1250; C6H10CaO6$5H2O, 1750; ZnSO4$7H2O,176.5; MnSO4.$4H2O, 81; and if or how much the fish could recover when they were CuSO4.$5H2O, 15.5; CoSO4$6H2O, 0.5; KI, 1.5; starch, 225. free of cyanobacteria in their diet. b Vitamin premix (mg kgÀ1 diet, NRC, 1993): Thiamin, 20; riboflavin, 20; pyridoxine, 20; cyanocobalamine, 2; folic acid, 5; calcium patotheniate, 50; inositol, 100; niacin, 100; biotin, 5; starch, 3226; vitamin A (ROVIMIX 2. Materials and methods A-1000), 110; vitamin D3, 20; vitamin E, 100; vitamin K3, 10. 2.1. Fish, cyanobacteria and experimental diets transferred from the rearing tank to the flow-through Hybrid tilapia (Oreochromis niloticus  O. aureus)were system for acclimation. At the beginning of the trial, fish obtained from the tilapia hatchery farm in Puqi, Hubei, were fasted for 1 day to empty the gut. Fifty fish of similar China and acclimated in two cylindrical fiberglass tanks size (initial body weight 2.2 g) were randomly selected, (diameter 150 cm  height 120 cm, water volume 1500 L) weighed and stocked in each tank. Six tanks were randomly for 20 days prior to the experiment. During the acclimation assigned to each diet. During the experiment, aeration was period, fish were fed to satiation twice daily (09:00 and provided to each tank to maintain dissolved oxygen level 15:00) with the control diet. above 7 mg LÀ1; the photoperiod was 12 h light:12 h dark Two batches of fresh cyanobacteria were collected from with the light period from 08:00 to 20:00. Light intensity at Lake Taihu in Jiangsu and Lake Dianchi in Yunnan, China. the water surface was around 200 lx. Water temperature The cyanobacteria from Lake Taihu were composed of 50% was recorded daily and was maintained at 26 Æ 2 C, pH Microcystis aeruginosa and 50% Microcystis wesenbegii was about 7.0. Ammonia-N was monitored once a week and whereas the cyanobacteria from Lake Dianchi mainly con- was less than 0.5 mg LÀ1 and residual chloride was less sisted of Microcystis aeruginosa (95%). The fresh cyanobac- than 0.01 mg LÀ1. Fish were hand-fed to apparent satiation teria were air-dried before use. twice daily (09:00 and 15:00). The daily food supplied was Three experimental diets were formulated to be recorded and uneaten diets were siphoned 1 h after approximately isonitrogenous (crude protein: 42%) and feeding, dried to constant weight at 70 C and reweighed. isocaloric (gross energy: 20 kJ gÀ1)(Table 1). In the control Leaching rates (potential loss of uneaten diet) of uneaten diet, 20% fishmeal and 49.9% soybean meal were used as the diets were estimated by placing weighed food in tanks protein source while 43.6% of cyanobacteria from Lake without fish for 1 h and then collecting, drying and Taihu were used to replace soybean meal to formulate AMt reweighing. Leaching rate was used to calibrate the and 50.8% of cyanobacteria from Lake Dianchi were used to uneaten diets. The accurate total food intake was calculated replace soybean meal to formulate AMd. The MCs concen- from the difference between food intake and the calibrated trations in the AMt and AMd diets were 80.0 mg MCs gÀ1 diet uneaten diet.