Effects of antibiotic residues on methane production in anaerobic digestion are commonly studied using the following two antibiotic addition methods: (1) adding manure from animals that consume a diet containing antibiotics, and (2) adding antibiotic-free animal manure spiked with antibiotics. This study used chlortetracycline (CTC) as a model antibiotic to examine the effects of the antibiotic addition method on methane production in anaerobic digestion under two different swine wastewater concentrations (0.55 and 0.22mg CTC/g dry manure). The results showed that CTC degradation rate in which manure was directly added at 0.55mg CTC/g (HSPIKE treatment) was lower than the control values and the rest of the treatment groups. Methane production from the HSPIKE treatment was reduced (p<0.05) by 12% during the whole experimental period and 15% during the first 7days. The treatments had no significant effect on the pH and chemical oxygen demand value of the digesters, and the total nitrogen of the 0.55mg CTC/kg manure collected from mediated swine was significantly higher than the other values. Therefore, different methane production under different antibiotic addition methods might be explained by the microbial activity and the concentrations of antibiotic intermediate products and metabolites. Because the primary entry route of veterinary antibiotics into an anaerobic digester is by contaminated animal manure, the most appropriate method for studying antibiotic residue effects on methane production may be using manure from animals that are given a particular antibiotic, rather than adding the antibiotic directly to the anaerobic digester.