In South American countries, bovine trypanosomiasis is caused mainly by Trypanosoma vivax. Among the infected animals, bovines are considered more susceptible, and this susceptibility varies among breeds. To determine the occurrence of T. vivax in Maranhão State, Brazil, a total of 559 cattle blood samples were collected for molecular and parasitological studies. On São Luís Island, no parasites were observed in the parasitological exam; however, with the polymerase chain reaction (PCR) method, three samples (1.06%) were positive. In four municipalities that compose the Pedreiras Basin, 3.39% of the animals were positive on the parasitological exam, and 6.21% of animals were positive by PCR. This finding demonstrates that PCR is more sensitive and confirms that the method is very useful in epidemiological surveys. A further interesting point is that molecular studies clearly define the differences in surface glycoproteins and antigenic variants.