This study investigates the effect of temperature (range, 20–35°C) on the growth and cylindrospermopsin (CYL) content of seven isolates of Cylindrospermopsis raciborskii grown in batch culture. The concentration of CYL, measured late in the exponential growth phase for cultures grown at 25°C, ranged from below detection (< 2 × 10−5% freeze-dried weight) for isolates from Lake Julius (20°08′S; 139°44′E) and an aquaculture pond in Townsville (19°16′S; 146°49′E) up to 0.46% for an isolate from Solomon Dam, Palm Island (18°45′S; 146°35′E). Maximum growth rates for all isolates occurred at temperatures between 25 and 30°C. Only one isolate failed to grow at 20°C. For isolates containing undetectable concentrations of CYL, the production of this compound could not be induced by an increase in temperature. For strains producing detectable concentrations, there was a strong negative correlation between CYL content and temperature. While all the isolates grew well at 35°C, none produced any detectable amount of CYL. When cultures of isolates that normally produce CYL were transferred from 35°C to a lower temperature, production of CYL was restored. In the cultured isolates, less than 10% of the total CYL was released from exponentially growing cells. In one CYL-producing isolate, extracellular CYL accounted for c. 50% of the total in stationary phase (20 day old) cultures.