Background: Asthma prevalence may be reduced in large families. The hygiene hypothesis suggests that older siblings protect their younger siblings from asthma through a modulating effect on the still-maturing immune system. If the hygiene hypothesis is correct, asthma prevalence should be inversely related to birth order. The objective of this study was to examine the relationship between asthma prevalence, and family size and birth order. Methods: The medical records of 531,116 Israeli military conscripts were reviewed. The association between number of children in the family and the prevalence of asthma, and between birth order and the prevalence of asthma was assessed. Odds ratios for asthma by birth order and family size, adjusted for each other, were calculated. Results: Asthma was diagnosed in 26,833 male subjects (8.6%) and 15,079 female subjects (6.9%). Asthma prevalence was inversely related to the number of children in the family (p < 0.001). Among subjects who were the only child in the family, the prevalence of asthma was 7.3%. The prevalence increased to 8.95% among subjects from families with three siblings, and then progressively decreased as the number of siblings increased, and reached a trough of 0.58% in conscripts from families of 15 to 20 siblings. Asthma prevalence was similar for all birth orders. Conclusions: In families with four or more children, asthma prevalence is inversely related to the number of children in the family. Asthma prevalence is similar for all birth orders. The similar asthma prevalence for all birth orders challenges the hygiene hypothesis as the mechanism for the decreased asthma prevalence in large families.