Symptomatic and asymptomatic astrovirus infection was prospectively determined in a 3-year birth cohort of Mayan infants. Stool samples from 271 infants and 268 older siblings were tested for astrovirus, adenovirus 40/41, rotavirus and Salmonella, Shigella and Campylobacter species. Concurrent diarrhea, vomiting, fever, or anorexia were noted. Astrovirus was detected in 164 infants (61%) and 20 siblings (7%). Rotavirus (4%) and adenovirus 40/41 (13%) were isolated less frequently. Of all diarrheal episodes reported at a visit, 26% (78/305) were associated with astrovirus; 17% (78/452) of astrovirus infections were associated with diarrhea and 9% with other symptoms. Only diarrhea was associated with astrovirus infection (odds ratio, 1.4; 95% confidence interval [CI], 1.07-1.92; P = .01). Of infants with astrovirus, 70% shed at multiple visits over a period of 2-17 weeks (median, 5). The point prevalence of astrovirus infection was significantly higher among infants than siblings (relative risk, 6.18; 95% CI, 3.93-9.72; P < .0001, chi2). Astrovirus was identified throughout the year, peaked in March and May, and decreased in September. In this population, astrovirus was the most common enteric pathogen isolated; symptomatic infection was prevalent among infants.