Long-term acclimation temperature effects on biomarkers of oxidative stress, metabolic stress, expression of heat shock proteins (Hsps), and warm-temperature acclimation related 65-kDa protein (Wap65) were evaluated in the threatened chocolate mahseer (Neolissochilus hexagonolepis). Fifteen-day-old larvae were acclimated to different water temperatures (15, 19, 23-control group, 27, and 31 degrees C) for 60 days prior to the sampling for quantification of mRNA, enzyme, nitric oxide, and malondialdehyde (MDA) content. Acclimation to 31 degrees C increased the basal mRNA level of glutathione S-transferase alpha 1 (GSTa1), and activities of catalase (CAT), glutathione reductase (GR), and GST enzymes and but downregulated the expression of superoxide dismutase 1 (SOD1) in the whole-body homogenate. Other antioxidant genes, i.e., CAT and GPx1a, were unaffected at 31 degrees C, and nitric oxide (NO) concentration was significantly lower. In contrast, fish acclimated to 15 degrees C showed an upregulated transcript level of all the antioxidant genes and no significant difference in the CAT, GR, and GST enzymes. Activities of the metabolic enzymes, aspartate transaminase (AST) and alanine transaminase (ALT), were significantly lower at 15 degrees C. The expression of Hsp47 was upregulated at both 15 and 31 degrees C groups, whereas Hsp70 was elevated at 27 and 31 degrees C groups. Wap65-1 transcription did not show significant variation in treatment groups compared to control. Fish in the high (31 degrees C) and low-temperature (15 degrees C) acclimation groups were capable of maintaining oxidative stress by modulating their antioxidant transcripts, enzymes, and Hsps.