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Tilapia were exposed to 0, 0.2, 2, 20, 200 µg/L methomyl for 30 days, and then transferred to methomyl-free water for 18 days. Caspase-8 in serum, apoptosis rate, microstructure and ultra-microstructure of testis were checked after methomyl exposure and at 18 days after transferring to methomyl-free water. There were no significant changes in Caspase-8 activity, apoptosis rate, and tissue structure in testis exposed to 0.2 and 2 µg/L compared with control. However, when tilapia exposed to 20 and 200 µg/L, the Caspase-8 activity and apoptosis rate were induced significantly, and tissue damage happened compared with the control. Thus it would appear 2 µg/L methomyl might be considered as the no observed adverse effect level. Recovery data showed that the effects produced by lower concentration of 20 µg/L were reversible but not at the higher 200 µg/L concentration.

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The discovery of aerobic and anammox bacteria capable of generating methane in bio-filters in freshwater aquaculture systems is generating interest in studies to understand the activity, diversity, distribution and roles of these environmental bacteria. In this study, we used microbial enrichment of bio-filters to assess their effect on water quality. Profiles of ammonia-oxidizing bacterial communities generated using nested PCR methods and DGGE were used to assess the expression of 16S rRNA genes using DNA sequencing. Five dominant ammonia-oxidizing bacterial strains-clones; KB.13, KB.15, KB.16, KB.17 and KB.18-were isolated and identified by phylogenetic analysis as environmental samples closely related to genera Methylobacillus, Stanieria, Nitrosomonas, and Heliorestis. The methyl ammonia-oxidizing microbes thereby found suggest a biochemical pathway involving electron donors and carbon sources, and all strains were functional in freshwater aquaculture systems. Environmental parameters including TN (2.69-20.43); COD (9.34-31.47); NH4+-N (0.44-11.78); NO2-N (0.00-3.67); NO3-N (0.05-1.82), mg/L and DO (1.47-10.31 µg/L) assessed varied in the ranges in the different tanks. Principal component analysis revealed that these water quality parameters significantly influenced the ammonia oxidizing microbial community composition. Temperature rises to about 40 °C significantly affected environmental characteristics-especially DO, TN and NH4+-N-and directly or indirectly affected the microbial communities. Although the nested PCR design was preferred due to its high sensitivity for amplifying specific DNA regions, a more concise method is recommended, as an equimolar mixture of degenerate PCR primer pairs, CTO189f-GC and CTO654r, never amplified only 16S rRNA of ammonia-oxidizing bacteria.

RESUMEN

Tilapia were exposed to sub-lethal concentrations of 0, 0.2, 2, 20 or 200 µg/L for 30 days, then transferred to methomyl-free water for 18 days. E2 , T, 11-KTand VTG in serum were examined. There were no significant changes in all the parameters in serum of tilapia exposed to 0.2 µg/L and 2 µg/L methomyl compared to the control. However, 20 µg/L and 200 µg/L have the potential to disrupt the endocrine system of male tilapia, as shown by its ability to increase VTG and E2 and decrease T and 11-KT in serum. Thus it would appear the no observed adverse effect level for sexual steroid hormones of methomyl is lower than 2 µg/L. Recovery data showed that the effects produced by 20µg/L were reversible but not at 200µg/L. Furthermore, the sensitivity of above parameters to methomyl followed the order of VTG>E2 >11-KT>T>GSI, suggesting VTG being the better biomarkers.

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Tilapia were exposed to sublethal methomyl concentrations of 0, 0.2, 2, 20 or 200 µg/L for 30 days, and then transferred to methomyl-free water for 18 days. The sexual steroid hormones 17ß-estradiol (E2), testosterone (T), and 11-ketotestosterone (11-KT) in tilapia testes were examined at 0, 6, 12, 18, 24 and 30 days after methomyl exposure, and at 18 days after fish were transferred to methomyl-free water. There were no significant changes in the hormone parameters in testes of tilapia exposed to low concentration 0.2 and 2 µg/L methomyl compared with the controls. However, high concentration 20 and 200 µg/L methomyl had the potential to disrupt the endocrine system of male tilapia, as shown by an increase in E2 and a decrease in T and 11-KT in the testes. Thus, it would appear that the 2 µg/L methomyl might be considered the no-observed-adverse-effect level. Recovery data showed that the effects produced by the lower concentration of 20 µg/L were reversible but the effects were not reversible at the higher concentration of 200 µg/L.

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The changes of microbial community in aquaculture systems under the effects of stocking densities and seasonality were investigated in tilapia ponds. Total DNAs were extracted from the water samples, 16S rRNA gene was amplified and the bacterial community analyzed by Illumina high-throughput sequencing obtaining 3486 OTUs, from a total read of 715,842 sequences. Basing on the analysis of bacterial compositions, richness, diversity, bacterial 16S rRNA gene abundance, water sample comparisons and existence of specific bacterial taxa within three fish ponds in a 4 months period, the study conclusively observed that the dominant phylum in all water samples were similar, and they included; Proteobacteria, Cyanobacteria, Bacteroidetes, Actinobacteria, Planctomycetes and Chlorobi, distributed in different proportions in the different months and ponds. The seasonal changes had a more pronounced effect on the bacterioplankton community than the stocking densities; however some differences between the ponds were more likely caused by feed coefficient than by stocking densities. At the same time, most bacterial communities were affected by the nutrient input except phylum Cyanobacteria that was also affected by the feed control of tilapia.

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The chronic effect of methomyl on the antioxidant system in tilapia (Oreochromis niloticus) was investigated. Fish were exposed to sub-lethal concentrations of 0.2, 2, 20 and 200µgL(-1) for 30 days, and then transferred to methomyl-free water for 18 days. Hepatic antioxidant parameters, including Glutathione-S-transferase (GST), Glutathione peroxidase (GPx), Glutathione reductase (GR), Reduced glutathione (GSH) and oxidized glutathione (GSSG), were measured at 10min (day 0), 6, 12, 18, 24 and 30 days after starting the experiment and at 18 days after transferring to methomyl-free water. There were no significant changes in enzymatic activity and content of antioxidants in liver of tilapia exposed to 0.2µgL(-1) methomyl compared to controls. However, the results showed significant increases in activities of GST, GR, GPx and levels of GSSG accompanied by a decrease in GSH levels following methomyl exposure in tilapia to 2, 20 or 200µgL(-1) over the 30-day exposure period and the highest induction rates in GST, GR, GPx and GSSG were 150.87%, 163.21%, 189.76%, and 179.56% of the control respectively, and the highest inhibition rate in GSH was 50.67% of the control, suggesting the presence of oxidative stress. Thus it would appear that the 0.2µgL(-1) methomyl might be considered as the no observed adverse effect level (NOAEL). Recovery data showed that the effects produced by lower concentration of methomyl 20µgL(-1) were reversible but not at the higher 200µgL(-1) concentration.

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Tilapia were exposed to sublethal methomyl concentrations of 0, 0.2, 2, 20, or 200 µg/L for 30 d, and then were transferred to methomyl-free water for 18 d. Renal antioxidant parameters, including catalase (CAT), superoxide dismutase (SOD), glutathione S-transferase (GST), glutathione peroxidase (GPx) , glutathione reductase (GR), total glutathione (GSH), and reduced glutathione (GSSG), were examined in tilapia at d 0, 6, 12, 18, 24, and 30 after starting the experiment and at 18 d after transferring to methomyl-free water. There were no significant changes in enzymatic activity and content of antioxidants in kidney of tilapia exposed to 0.2 µg/L methomyl compared to controls. The results showed significant increases in SOD, CAT, GST, GR, GPx, and level of GSSG accompanied by a decrease in GSH levels following methomyl exposure in tilapia to 2, 20, or 200 µg/L over the 30-d exposure period, suggesting the presence of oxidative stress. Thus, it would appear the 0.2 µg/L methomyl might be considered the no-observed-adverse-effect level (NOAEL). Recovery data showed that the effects produced by lower concentration of methomyl at 20 µg/L were reversible but not at the higher 200 µg/L concentration.

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