Colony organization via caste specialization in the Formosan subterranean termite, Coptotermes formosanus Shiraki, as in other species of termites, seems to be influenced by external factors such as nutrition and temperature. Here, we report on the influence of nutrition and temperature and the associated juvenile hormone levels on the regulation of the soldier caste. Two nutritional levels of food sources or five different temperatures were provided to C. formosanus workers in the laboratory. Juvenile hormone titers of these workers were monitored on 0, 12, 24, 36, 48, and 60 d after treatment. Termite survival and soldier-to-worker proportions also were recorded at each sample date. Workers fed a high-quality food source produced significantly more soldiers and this was associated with high worker juvenile hormone (JH) titers. JH titers in workers maintained at 20 and 36°C remained constant throughout the observational period with no presoldier or soldier formation. Under 24, 28, and 32°C temperature regimes, JH titers in workers climbed to a peak at day 48 and then sharply dropped at day 60 when soldier proportion increased to a specific value. At each sample date, JH titers in workers demonstrated a positive and statistically significant relationship to temperatures within the range of 24–32°C, but upon inclusion of temperatures below 24 and above 32°C, the correlation is no longer significant. The results show that favorable extrinsic conditions of temperature and nutrition allow for a higher soldier proportion threshold through the regulation of JH levels and caste differentiation in workers.