Trypanosoma cruzi expresses a unique trans-sialidase that is responsible for the transfer of sialic acid from host glycoproteins and glycolipids to mucin-like glycoprotein acceptors on the parasite surface. The enzyme and the sialic acid acceptors are present in the mammalian forms of the parasite and in the parasite forms that grow in axenic cultures, which correspond to the developmental stages found in the insect vectors. Here we show that parasite forms growing in the vector Triatoma infestans express trans-sialidase in the hindgut portions of the insect. However, the sialic acid acceptors are poorly sialylated due to the low concentration of sialic acid donors in the gut lumen of T.infestans, which feeds exclusively on blood that is rich in sialic acid donors. These low levels of sialic acid donors are due to a novel sialidase activity present mainly in the anterior midgut with high specificity for alpha-2,3-sialyllactose, but not for alpha-2,6-sialyllactose. The activity is present in starved insects or insects fed with culture medium, indicating that it did not originate from the blood meal. Enzyme activity does not decrease in insects fed with antibiotics, is present in the salivary glands, and the few bacteria isolated from the gut and faeces of T.infestans did not display sialidase activity, indicating that the enzyme is not derived from a commensal organism. This novel activity could have a nutritional role in the gut of haematophagous insects and indicates that acquisition of sialic acid is not required for parasite development in the gut of T.infestans.