Ammonia sanitisation is a promising treatment alternative for inactivation of pathogens in fecal sludge intended for agricultural use. Inactivation of Ascaris eggs and Salmonella spp. was studied in fecal sludge at ≥28 °C treated with low doses of urea, and in fecal sludge at ≤17 °C treated with high doses of ammonia solution. The effect of ammonia and carbonate on Ascaris inactivation in buffer was also studied. Ascaris eggs and Salmonella spp. were inactivated in fecal sludge treated with 0.4% urea or more at ≥28 °C. With lower doses of urea, the pH of the fecal sludge decreased during the experiment, resulting in low NH3 concentrations and subsequently no inactivation of Ascaris eggs. Ascaris was successfully inactivated at 5 °C, but the NH3 concentrations required were 10-fold higher than at high temperatures and the storage time required was longer. The buffer study showed that carbonate (CO32−) had a statistically significant impact on Ascaris inactivation, but the effect was low compared with that of NH3. Thus for inactivation of Salmonella spp. with urea at low temperatures, CO32− is probably a more important factor than NH3.