Background: The Australian Guide to Healthy Eating (AGHE) for pregnancy provides a number of food- and nutrition-related recommendations to assist pregnant women in optimizing their dietary behavior. However, there are limited data demonstrating pregnant women’s knowledge of the AGHE recommendations. This study investigated Australian pregnant women’s knowledge of the AGHE and related dietary recommendations for maintaining a healthy pregnancy. The variations in nutrition knowledge were compared with demographic characteristics. Methods: A cross-sectional study assessed eight different nutrition knowledge domains and the demographic characteristics of pregnant women. Four hundred women across Australia completed a multidimensional online survey based on validated and existing measures. Results: More than half of the pregnant women surveyed (65%) were not familiar with the AGHE recommendations. The basic recommendations to eat more fruit, vegetables, bread, and cereals but less meat were poorly understood. An in-depth investigation of knowledge of nutrition information revealed misconceptions in a range of areas, including standard serving size, nutrients content of certain foods, energy density of fat, and the importance of key nutrients in pregnancy. Univariate analysis revealed significant demographic variation in nutrition knowledge scores. Multiple regression analysis confirmed the significant independent effects on respondents’ nutrition knowledge score (P<0.000) of the education level, income, age, stage of pregnancy, language, and having a health/nutrition qualification. The model indicated that independent variables explained 33% (adjusted R2) of the variance found between respondents’ knowledge scores. Conclusion: Australian pregnant women’s knowledge regarding AGHE for pregnancy and other key dietary recommendations is poor and varies significantly with their demographic profile. The setting of dietary guidelines is not sufficient to ensure improvement in their nutrition knowledge. It is essential that women receive support to achieve optimal and healthy diets during pregnancy.